Proteopedia - User contributions [en]

Sandbox Reserved 977

2015-05-01T23:18:59Z

Joseph P. Hoffmann:

Sandbox Reserved 982

2015-05-01T22:02:10Z

Joseph P. Hoffmann:

{{Telford_CHEM321}}
==Shiga Toxin==
<StructureSection load='1R4Q' size='340' side='right' caption='Crystal Structure for Shiga Toxin (1R4Q)' scene=''>

== Structural Highlights ==
Shiga toxin (stx) has multiple classifications. The shiga toxin produced by ''Shigella dysenteriae'' is classified as stx, while shiga toxin produced by Escherichia coli is classified as either stx1 or stx2. All three classifications vary slightly in structure but have identical functions. The A sub-unit of stx1 differs from stx by one residue: a serine in position 45 as opposed to threonine<ref name="Fraser">Fraser, M. E., Fujinaga, M., Cherney, M. M., Melton-Celsa, A. R., Twiddy, E. M., O’Brien, A. D., & James, M. N. G. (2004). Structure of shiga toxin type 2 (Stx2) from Escherichia coli O157:H7. The Journal of Biological Chemistry, 279(26), 27511–27517. doi:10.1074/jbc.M401939200</ref>.

Stx is a 70 kDa AB5 protein, meaning that it is composed of an A sub-unit bound to a B pentamer. A pentamer of B sub-units encircles the carboxy tail of the A sub-unit stabilized by a series of non-covalent interactions. <scene name='68/687332/Sub-unit_a/2'>The A sub-unit</scene> is 293 amino acids long with its active site being <scene name='68/687332/Active_site_for_stx/1'>glutamic acid 167 (1R4Q)</scene>. Though the active site contains a single residue, studies have shown that the first 239 residues are essential for the enzymatic activity of the A sub-unit. Residues 240-251 are essential for the A sub-unit’s translocation from the endoplasmic reticulum of an infected cell to the cytosol<ref name="Sandvig">Sandvig, K., & van Deurs, B. (2000). Entry of ricin and Shiga toxin into cells: molecular mechanisms and medical perspectives. The EMBO Journal, 19(22), 5943–5950. doi:10.1093/emboj/19.22.5943</ref>.

The A sub-unit can be broken down into A1 and A2 units. This break down occurs when a trypsin sensitive region, residues 248-251, is cleaved. This cleavage results in the only remaining link being a <scene name='68/687332/Disulfide_bridge/1'>disulfide bridge (1R4Q)</scene> between cysteine 242 and cysteine 261. If the disulfide bridge is reduced, the A1 and A2 fragments can completely separate. Interestingly, the disulfide bridge blocks the active site of the A sub-unit, so the A sub-unit is not enzymatically active unless it has been cleaved<ref name ="Fraser">Fraser, M. E., Fujinaga, M., Cherney, M. M., Melton-Celsa, A. R., Twiddy, E. M., O’Brien, A. D., & James, M. N. G. (2004). Structure of shiga toxin type 2 (Stx2) from Escherichia coli O157:H7. The Journal of Biological Chemistry, 279(26), 27511–27517. doi:10.1074/jbc.M401939200</ref>.

<scene name='68/687332/B_subunit/1'>The B sub-unit (2xsc)</scene> is a symmetrical pentamer that resembles a star and can be divided into five identical monomers. Each monomer is 69 amino acids in length. The purpose of the B sub-unit is to bind to globotriaosylceramide (GB3) which is a glycosphingolipid that is found on the lipid rafts of endothelial cells. Each monomer of the B sub-unit has three binding sites for GB3, binding to the <scene name='68/687332/Gb3_bound_to_subunit_b/1'>carbohydrate portion of GB3 (1cqf)</scene>. The B sub-unit has a millimolar affinity for GB3, and is actually one of the lowest recorded affinities for carbohydrate-protein interaction<ref>Jacobson, J. M., Yin, J., Kitov, P. I., Mulvey, G., Griener, T. P., James, M. N. G., … Bundle, D. R. (2014). The crystal structure of Shiga toxin type 2 with bound disaccharide guides the design of a heterobifunctional toxin inhibitor. Journal of Biological Chemistry, 289(2), 885–894. doi:10.1074/jbc.M113.518886</ref>. Some studies have shown that the level of saturation and length of the fatty acid on GB3 affects the strength of its interaction with the B sub-unit. Since the B sub-unit of stx has fifteen binding sites though, it significantly increases the strength of the interaction.<ref name="Melton">Melton-Celsa, Angela R. “Shiga Toxin (Stx) Classification, Structure, and Function.” Microbiology spectrum 2.2 (2014): 10.1128/microbiolspec.EHEC–0024–2013. PMC. Web</ref>.
== Function ==
The pathway of stx entering a cell begins with the B sub-unit’s binding to GB3. Once this occurs, the A sub-unit disconnects from the B sub-unit and enters the cell through endocytosis. Using retrograde transport the A sub-unit passes through the Golgi apparatus and the rough endoplasmic reticulum<ref name="Melton">Melton-Celsa, Angela R. “Shiga Toxin (Stx) Classification, Structure, and Function.” Microbiology spectrum 2.2 (2014): 10.1128/microbiolspec.EHEC–0024–2013. PMC. Web</ref> In the rough endoplasmic reticulum, the A sub-unit is cleaved into two parts, A1 and A2, through the cleavage of trypsin sensitive residues and the reduction of a disulfide bridge. A2 is degraded, but A1 freely enters the cytosol<ref name="Sandvig">Sandvig, K., & van Deurs, B. (2000). Entry of ricin and Shiga toxin into cells: molecular mechanisms and medical perspectives. The EMBO Journal, 19(22), 5943–5950. doi:10.1093/emboj/19.22.5943</ref>. Once in the cytosol, A1 acts as an N-glycosidase, hydrolyzing bonds that link sugars. With this enzymatic activity, A1 removes an adenine from the alpha-sarcin loop in the 28S RNA of the 60S ribosomal sub-unit<ref name="Melton">Melton-Celsa, Angela R. “Shiga Toxin (Stx) Classification, Structure, and Function.” Microbiology spectrum 2.2 (2014): 10.1128/microbiolspec.EHEC–0024–2013. PMC. Web</ref>. The removal of the adenine prevents elongation factors from associating with the ribosomal sub-unit. Without elongation factors, the ribosome can no longer synthesize proteins, leading to cell death<ref name="Melton">Melton-Cesla, A. (2012). Shiga toxin classification structure and function. Changes, 29(2), 997–1003</ref>.
== Disease and Pathogenesis ==
The shiga toxin is produced by ''Shigella dysenteriae'' and certain strains of ''Escherichia coli''. ''S. dysenteriae'' is a gram negative bacterium that causes disease in its host by entering into intestinal epithelial cells. It has a very low infectious dose of only ten to one hundred cells, and is contracted through the consumption of contaminated water or food. Once in intestines, the bacteria's proliferation and secretion of stx erodes the intestinal lining and leads to dysentery<ref>Zaidi, Mussaret Bano, and Teresa Estrada-García. "Shigella: A Highly Virulent and Elusive pathogen." Current Tropical Medicine Reports 1.2 (2014): 81-87</ref>. Those with dysentery can simply have diarrhea as a symptom, but if the infection progresses, it can lead to hemorrhagic colitis (bloody stool) or hemolytic uremic syndrome (HUS). HUS is characterized as acute renal failure, thrombocytopenia, and microangiopathic anemia<ref>Kaper, James B. and Alison D. O’Brien. “Overview and Historical Perspectives.” Microbiology spectrum 2.2 (2014): 10.1128/microbiolspec.EHEC–0028–2014. PMC. Web</ref>.

As stated previously, stx enters cells through its binding to GB3, but curiously many intestinal cells do not express GB3. Some researchers hypothesize it enters the cells through macropinocyosis, but when macropinocytotic function is removed from cells, stx still manages to enter through an as yet undefined mechanism.

The genes necessary to produce the toxin are interestingly not native to ''S. dysenteriae''. The genes for stx are carried by prophages: bacteria that have had viral DNA inserted into their genome. During certain conditions, the phage genes are expressed leading to the production of more bacteriophages as well as the shiga toxin. When the bacteria undergo lysis, bacteriophage and stx are released. It may seem as if there is no benefit for ''S. dysenteriae'' to carry the viral DNA, but studies have shown that while it is in the lysogenic cycle S. dysenteriae has increased acid tolerance and motility<ref>Krüger, Alejandra, and Paula María Alejandra Lucchesi. "Shiga toxins and stx-phages: highly diverse entities." Microbiology (2014): mic-0</ref>.

</StructureSection>
== References ==
<references/>

Sandbox Reserved 982

2015-05-01T20:44:59Z

Joseph P. Hoffmann:

{{Telford_CHEM321}}
==Shiga Toxin==
<StructureSection load='1R4Q' size='340' side='right' caption='Crystal Structure for Shiga Toxin (1R4Q)' scene=''>

== Structural Highlights ==
Shiga toxin (stx) has multiple classifications. The shiga toxin produced by ''Shigella dysenteriae'' is classified as stx, while shiga toxin produced by Escherichia coli is classified as either stx1 or stx2. All three classifications vary slightly in structure but have identical functions. The A sub-unit of stx1 differs from stx by one residue: a serine in position 45 as opposed to threonine<ref name="Fraser">Fraser, M. E., Fujinaga, M., Cherney, M. M., Melton-Celsa, A. R., Twiddy, E. M., O’Brien, A. D., & James, M. N. G. (2004). Structure of shiga toxin type 2 (Stx2) from Escherichia coli O157:H7. The Journal of Biological Chemistry, 279(26), 27511–27517. doi:10.1074/jbc.M401939200</ref>.

Stx is a 70 kDa AB5 protein, meaning that it is composed of an A sub-unit bound to a B pentamer. A pentamer of B sub-units encircles the carboxy tail of the A sub-unit stabilized by a series of non-covalent interactions. <scene name='68/687332/Sub-unit_a/2'>The A sub-unit</scene> is 293 amino acids long with its active site being <scene name='68/687332/Active_site_for_stx/1'>glutamic acid 167 (1R4Q)</scene>. Though the active site contains a single residue, studies have shown that the first 239 residues are essential for the enzymatic activity of the A sub-unit. Residues 240-251 are essential for the A sub-unit’s translocation from the endoplasmic reticulum of an infected cell to the cytosol<ref name="Sandvig">Sandvig, K., & van Deurs, B. (2000). Entry of ricin and Shiga toxin into cells: molecular mechanisms and medical perspectives. The EMBO Journal, 19(22), 5943–5950. doi:10.1093/emboj/19.22.5943</ref>.

The A sub-unit can be broken down into A1 and A2 units. This break down occurs when a trypsin sensitive region, residues 248-251, is cleaved. This cleavage results in the only remaining link being a <scene name='68/687332/Disulfide_bridge/1'>disulfide bridge (1R4Q)</scene> between cysteine 242 and cysteine 261. If the disulfide bridge is reduced, the A1 and A2 fragments can completely separate. Interestingly, the disulfide bridge blocks the active site of the A sub-unit, so the A sub-unit is not enzymatically active unless it has been cleaved<ref name ="Fraser">Fraser, M. E., Fujinaga, M., Cherney, M. M., Melton-Celsa, A. R., Twiddy, E. M., O’Brien, A. D., & James, M. N. G. (2004). Structure of shiga toxin type 2 (Stx2) from Escherichia coli O157:H7. The Journal of Biological Chemistry, 279(26), 27511–27517. doi:10.1074/jbc.M401939200</ref>.

<scene name='68/687332/B_subunit/1'>The B sub-unit (2xsc)</scene> is a symmetrical pentamer that resembles a star and can be divided into five identical monomers. Each monomer is 69 amino acids in length. The purpose of the B sub-unit is to bind to globotriaosylceramide (GB3) which is a glycosphingolipid that is found on the lipid rafts of endothelial cells. Each monomer of the B sub-unit has three binding sites for GB3, binding to the <scene name='68/687332/Gb3_bound_to_subunit_b/1'>carbohydrate portion of GB3 (1cqf)</scene>. The affinity between the B sub-unit and GB3 is incredibly low, and is actually one of the lowest recorded affinities for carbohydrate-protein interaction<ref>Jacobson, J. M., Yin, J., Kitov, P. I., Mulvey, G., Griener, T. P., James, M. N. G., … Bundle, D. R. (2014). The crystal structure of Shiga toxin type 2 with bound disaccharide guides the design of a heterobifunctional toxin inhibitor. Journal of Biological Chemistry, 289(2), 885–894. doi:10.1074/jbc.M113.518886</ref>. Some studies have shown that the level of saturation and length of the fatty acid on GB3 affects the strength of its interaction with the B sub-unit. Since the B sub-unit of stx has fifteen binding sites though, it has an incredibly high avidity<ref name="Melton">Melton-Celsa, Angela R. “Shiga Toxin (Stx) Classification, Structure, and Function.” Microbiology spectrum 2.2 (2014): 10.1128/microbiolspec.EHEC–0024–2013. PMC. Web</ref>.
== Function ==
The pathway of stx entering a cell begins with the B sub-unit’s binding to GB3. Once this occurs, the A sub-unit disconnects from the B sub-unit and enters the cell through endocytosis. Using retrograde transport the A sub-unit passes through the Golgi apparatus and the rough endoplasmic reticulum<ref name="Melton">Melton-Celsa, Angela R. “Shiga Toxin (Stx) Classification, Structure, and Function.” Microbiology spectrum 2.2 (2014): 10.1128/microbiolspec.EHEC–0024–2013. PMC. Web</ref> In the rough endoplasmic reticulum, the A sub-unit is cleaved into two parts, A1 and A2, through the cleavage of trypsin sensitive residues and the reduction of a disulfide bridge. A2 is degraded, but A1 freely enters the cytosol<ref name="Sandvig">Sandvig, K., & van Deurs, B. (2000). Entry of ricin and Shiga toxin into cells: molecular mechanisms and medical perspectives. The EMBO Journal, 19(22), 5943–5950. doi:10.1093/emboj/19.22.5943</ref>. Once in the cytosol, A1 acts as an N-glycosidase, hydrolyzing bonds that link sugars. With this enzymatic activity, A1 removes an adenine from the alpha-sarcin loop in the 28S RNA of the 60S ribosomal sub-unit<ref name="Melton">Melton-Celsa, Angela R. “Shiga Toxin (Stx) Classification, Structure, and Function.” Microbiology spectrum 2.2 (2014): 10.1128/microbiolspec.EHEC–0024–2013. PMC. Web</ref>. The removal of the adenine prevents elongation factors from associating with the ribosomal sub-unit. Without elongation factors, the ribosome can no longer synthesize proteins, leading to cell death<ref name="Melton">Melton-Cesla, A. (2012). Shiga toxin classification structure and function. Changes, 29(2), 997–1003</ref>.
== Disease and Pathogenesis ==
The shiga toxin is produced by ''Shigella dysenteriae'' and certain strains of ''Escherichia coli''. ''S. dysenteriae'' is a gram negative bacterium that causes disease in its host by entering into intestinal epithelial cells. It has a very low infectious dose of only ten to one hundred cells, and is contracted through the consumption of contaminated water or food. Once in intestines, the bacteria's proliferation and secretion of stx erodes the intestinal lining and leads to dysentery<ref>Zaidi, Mussaret Bano, and Teresa Estrada-García. "Shigella: A Highly Virulent and Elusive pathogen." Current Tropical Medicine Reports 1.2 (2014): 81-87</ref>. Those with dysentery can simply have diarrhea as a symptom, but if the infection progresses, it can lead to hemorrhagic colitis (bloody stool) or hemolytic uremic syndrome (HUS). HUS is characterized as acute renal failure, thrombocytopenia, and microangiopathic anemia<ref>Kaper, James B. and Alison D. O’Brien. “Overview and Historical Perspectives.” Microbiology spectrum 2.2 (2014): 10.1128/microbiolspec.EHEC–0028–2014. PMC. Web</ref>.

As stated previously, stx enters cells through its binding to GB3, but curiously many intestinal cells do not express GB3. Some researchers hypothesize it enters the cells through macropinocyosis, but when macropinocytotic function is removed from cells, stx still manages to enter through an as yet undefined mechanism.

The genes necessary to produce the toxin are interestingly not native to ''S. dysenteriae''. The genes for stx are carried by prophages: bacteria that have had viral DNA inserted into their genome. During certain conditions, the phage genes are expressed leading to the production of more bacteriophages as well as the shiga toxin. When the bacteria undergo lysis, bacteriophage and stx are released. It may seem as if there is no benefit for ''S. dysenteriae'' to carry the viral DNA, but studies have shown that while it is in the lysogenic cycle S. dysenteriae has increased acid tolerance and motility<ref>Krüger, Alejandra, and Paula María Alejandra Lucchesi. "Shiga toxins and stx-phages: highly diverse entities." Microbiology (2014): mic-0</ref>.

</StructureSection>
== References ==
<references/>

Sandbox Reserved 982

2015-05-01T20:41:33Z

Joseph P. Hoffmann:

{{Telford_CHEM321}}
==Shiga Toxin==
<StructureSection load='1R4Q' size='340' side='right' caption='Crystal Structure for Shiga Toxin (1R4Q)' scene=''>

== Structural Highlights ==
Shiga toxin (stx) has multiple classifications. The shiga toxin produced by ''Shigella dysenteriae'' is classified as stx, while shiga toxin produced by Escherichia coli is classified as either stx1 or stx2. All three classifications vary slightly in structure but have identical functions. The A sub-unit of stx1 differs from stx by one residue: a serine in position 45 as opposed to threonine<ref name="Fraser">Fraser, M. E., Fujinaga, M., Cherney, M. M., Melton-Celsa, A. R., Twiddy, E. M., O’Brien, A. D., & James, M. N. G. (2004). Structure of shiga toxin type 2 (Stx2) from Escherichia coli O157:H7. The Journal of Biological Chemistry, 279(26), 27511–27517. doi:10.1074/jbc.M401939200</ref>.

Stx is a 70 kDa AB5 protein, meaning that it is composed of an A sub-unit bound to a B pentamer. A pentamer of B sub-units encircles the carboxy tail of the A sub-unit stabilized by a series of non-covalent interactions. <scene name='68/687332/Sub-unit_a/2'>The A sub-unit</scene> is 293 amino acids long with its active site being <scene name='68/687332/Active_site_for_stx/1'>glutamic acid 167 (1R4Q)</scene>. Though the active site contains a single residue, studies have shown that the first 239 residues are essential for the enzymatic activity of the A sub-unit. Residues 240-251 are essential for the A sub-unit’s translocation from the endoplasmic reticulum of an infected cell to the cytosol<ref name="Sandvig">Sandvig, K., & van Deurs, B. (2000). Entry of ricin and Shiga toxin into cells: molecular mechanisms and medical perspectives. The EMBO Journal, 19(22), 5943–5950. doi:10.1093/emboj/19.22.5943</ref>.

The A sub-unit can be broken down into A1 and A2 units. This break down occurs when a trypsin sensitive region, residues 248-251, is cleaved. This cleavage results in the only remaining link being a <scene name='68/687332/Disulfide_bridge/1'>disulfide bridge (1R4Q)</scene> between cysteine 242 and cysteine 261. If the disulfide bridge is reduced, the A1 and A2 fragments can completely separate. Interestingly, the disulfide bridge blocks the active site of the A sub-unit, so the A sub-unit is not enzymatically active unless it has been cleaved<ref name ="Fraser">Fraser, M. E., Fujinaga, M., Cherney, M. M., Melton-Celsa, A. R., Twiddy, E. M., O’Brien, A. D., & James, M. N. G. (2004). Structure of shiga toxin type 2 (Stx2) from Escherichia coli O157:H7. The Journal of Biological Chemistry, 279(26), 27511–27517. doi:10.1074/jbc.M401939200</ref>.

<scene name='68/687332/B_sub-unit/1'>The B sub-unit (2XSC)</scene> is a symmetrical pentamer that resembles a star and can be divided into five identical monomers. Each monomer is 69 amino acids in length. The purpose of the B sub-unit is to bind to globotriaosylceramide (GB3) which is a glycosphingolipid that is found on the lipid rafts of endothelial cells. Each monomer of the B sub-unit has three binding sites for GB3, binding to the <scene name='68/687332/Gb3_bound_to_subunit_b/1'>carbohydrate portion of GB3</scene>. The affinity between the B sub-unit and GB3 is incredibly low, and is actually one of the lowest recorded affinities for carbohydrate-protein interaction<ref>Jacobson, J. M., Yin, J., Kitov, P. I., Mulvey, G., Griener, T. P., James, M. N. G., … Bundle, D. R. (2014). The crystal structure of Shiga toxin type 2 with bound disaccharide guides the design of a heterobifunctional toxin inhibitor. Journal of Biological Chemistry, 289(2), 885–894. doi:10.1074/jbc.M113.518886</ref>. Some studies have shown that the level of saturation and length of the fatty acid on GB3 affects the strength of its interaction with the B sub-unit. Since the B sub-unit of stx has fifteen binding sites though, it has an incredibly high avidity<ref name="Melton">Melton-Celsa, Angela R. “Shiga Toxin (Stx) Classification, Structure, and Function.” Microbiology spectrum 2.2 (2014): 10.1128/microbiolspec.EHEC–0024–2013. PMC. Web</ref>.
== Function ==
The pathway of stx entering a cell begins with the B sub-unit’s binding to GB3. Once this occurs, the A sub-unit disconnects from the B sub-unit and enters the cell through endocytosis. Using retrograde transport the A sub-unit passes through the Golgi apparatus and the rough endoplasmic reticulum<ref name="Melton">Melton-Celsa, Angela R. “Shiga Toxin (Stx) Classification, Structure, and Function.” Microbiology spectrum 2.2 (2014): 10.1128/microbiolspec.EHEC–0024–2013. PMC. Web</ref> In the rough endoplasmic reticulum, the A sub-unit is cleaved into two parts, A1 and A2, through the cleavage of trypsin sensitive residues and the reduction of a disulfide bridge. A2 is degraded, but A1 freely enters the cytosol<ref name="Sandvig">Sandvig, K., & van Deurs, B. (2000). Entry of ricin and Shiga toxin into cells: molecular mechanisms and medical perspectives. The EMBO Journal, 19(22), 5943–5950. doi:10.1093/emboj/19.22.5943</ref>. Once in the cytosol, A1 acts as an N-glycosidase, hydrolyzing bonds that link sugars. With this enzymatic activity, A1 removes an adenine from the alpha-sarcin loop in the 28S RNA of the 60S ribosomal sub-unit<ref name="Melton">Melton-Celsa, Angela R. “Shiga Toxin (Stx) Classification, Structure, and Function.” Microbiology spectrum 2.2 (2014): 10.1128/microbiolspec.EHEC–0024–2013. PMC. Web</ref>. The removal of the adenine prevents elongation factors from associating with the ribosomal sub-unit. Without elongation factors, the ribosome can no longer synthesize proteins, leading to cell death<ref name="Melton">Melton-Cesla, A. (2012). Shiga toxin classification structure and function. Changes, 29(2), 997–1003</ref>.
== Disease and Pathogenesis ==
The shiga toxin is produced by ''Shigella dysenteriae'' and certain strains of ''Escherichia coli''. ''S. dysenteriae'' is a gram negative bacterium that causes disease in its host by entering into intestinal epithelial cells. It has a very low infectious dose of only ten to one hundred cells, and is contracted through the consumption of contaminated water or food. Once in intestines, the bacteria's proliferation and secretion of stx erodes the intestinal lining and leads to dysentery<ref>Zaidi, Mussaret Bano, and Teresa Estrada-García. "Shigella: A Highly Virulent and Elusive pathogen." Current Tropical Medicine Reports 1.2 (2014): 81-87</ref>. Those with dysentery can simply have diarrhea as a symptom, but if the infection progresses, it can lead to hemorrhagic colitis (bloody stool) or hemolytic uremic syndrome (HUS). HUS is characterized as acute renal failure, thrombocytopenia, and microangiopathic anemia<ref>Kaper, James B. and Alison D. O’Brien. “Overview and Historical Perspectives.” Microbiology spectrum 2.2 (2014): 10.1128/microbiolspec.EHEC–0028–2014. PMC. Web</ref>.

As stated previously, stx enters cells through its binding to GB3, but curiously many intestinal cells do not express GB3. Some researchers hypothesize it enters the cells through macropinocyosis, but when macropinocytotic function is removed from cells, stx still manages to enter through an as yet undefined mechanism.

The genes necessary to produce the toxin are interestingly not native to ''S. dysenteriae''. The genes for stx are carried by prophages: bacteria that have had viral DNA inserted into their genome. During certain conditions, the phage genes are expressed leading to the production of more bacteriophages as well as the shiga toxin. When the bacteria undergo lysis, bacteriophage and stx are released. It may seem as if there is no benefit for ''S. dysenteriae'' to carry the viral DNA, but studies have shown that while it is in the lysogenic cycle S. dysenteriae has increased acid tolerance and motility<ref>Krüger, Alejandra, and Paula María Alejandra Lucchesi. "Shiga toxins and stx-phages: highly diverse entities." Microbiology (2014): mic-0</ref>.

</StructureSection>
== References ==
<references/>

Sandbox Reserved 982

2015-05-01T20:23:13Z

Joseph P. Hoffmann:

{{Telford_CHEM321}}
==Shiga Toxin==
<StructureSection load='1R4Q' size='340' side='right' caption='Crystal Structure for Shiga Toxin (1R4Q)' scene=''>

== Structural Highlights ==
Shiga toxin (stx) has multiple classifications. The shiga toxin produced by ''Shigella dysenteriae'' is classified as stx, while shiga toxin produced by Escherichia coli is classified as either stx1 or stx2. All three classifications vary slightly in structure but have identical functions. The A sub-unit of stx1 differs from stx by one residue: a serine in position 45 as opposed to threonine<ref name="Fraser">Fraser, M. E., Fujinaga, M., Cherney, M. M., Melton-Celsa, A. R., Twiddy, E. M., O’Brien, A. D., & James, M. N. G. (2004). Structure of shiga toxin type 2 (Stx2) from Escherichia coli O157:H7. The Journal of Biological Chemistry, 279(26), 27511–27517. doi:10.1074/jbc.M401939200</ref>.

Stx is a 70 kDa AB5 protein, meaning that it is composed of an A sub-unit bound to a B pentamer. A pentamer of B sub-units encircles the carboxy tail of the A sub-unit stabilized by a series of non-covalent interactions. <scene name='68/687332/Sub-unit_a/2'>The A sub-unit</scene> is 293 amino acids long with its active site being <scene name='68/687332/Active_site_for_stx/1'>glutamic acid 167 (1R4Q)</scene>. Though the active site contains a single residue, studies have shown that the first 239 residues are essential for the enzymatic activity of the A sub-unit. Residues 240-251 are essential for the A sub-unit’s translocation from the endoplasmic reticulum of an infected cell to the cytosol<ref name="Sandvig">Sandvig, K., & van Deurs, B. (2000). Entry of ricin and Shiga toxin into cells: molecular mechanisms and medical perspectives. The EMBO Journal, 19(22), 5943–5950. doi:10.1093/emboj/19.22.5943</ref>.

The A sub-unit can be broken down into A1 and A2 units. This break down occurs when a trypsin sensitive region, residues 248-251, is cleaved. This cleavage results in the only remaining link being a <scene name='68/687332/Disulfide_bridge/1'>disulfide bridge (1R4Q)</scene> between cysteine 242 and cysteine 261. If the disulfide bridge is reduced, the A1 and A2 fragments can completely separate. Interestingly, the disulfide bridge blocks the active site of the A sub-unit, so the A sub-unit is not enzymatically active unless it has been cleaved<ref name ="Fraser">Fraser, M. E., Fujinaga, M., Cherney, M. M., Melton-Celsa, A. R., Twiddy, E. M., O’Brien, A. D., & James, M. N. G. (2004). Structure of shiga toxin type 2 (Stx2) from Escherichia coli O157:H7. The Journal of Biological Chemistry, 279(26), 27511–27517. doi:10.1074/jbc.M401939200</ref>.

<scene name='68/687332/B_sub-unit/1'>The B sub-unit (2XSC)</scene> is a symmetrical pentamer that resembles a star and can be divided into five identical monomers. Each monomer is 69 amino acids in length. The purpose of the B sub-unit is to bind to globotriaosylceramide (GB3) which is a glycosphingolipid that is found on the lipid rafts of endothelial cells. Each monomer of the B sub-unit has three binding sites for GB3, binding to the <scene name='68/687332/Gb3_bound_to_sub-unit_b/1'> carbohydrate portion of GB3 (1CQF)</scene>. The affinity between the B sub-unit and GB3 is incredibly low, and is actually one of the lowest recorded affinities for carbohydrate-protein interaction<ref>Jacobson, J. M., Yin, J., Kitov, P. I., Mulvey, G., Griener, T. P., James, M. N. G., … Bundle, D. R. (2014). The crystal structure of Shiga toxin type 2 with bound disaccharide guides the design of a heterobifunctional toxin inhibitor. Journal of Biological Chemistry, 289(2), 885–894. doi:10.1074/jbc.M113.518886</ref>. Some studies have shown that the level of saturation and length of the fatty acid on GB3 affects the strength of its interaction with the B sub-unit. Since the B sub-unit of stx has fifteen binding sites though, it has an incredibly high avidity<ref name="Melton">Melton-Celsa, Angela R. “Shiga Toxin (Stx) Classification, Structure, and Function.” Microbiology spectrum 2.2 (2014): 10.1128/microbiolspec.EHEC–0024–2013. PMC. Web</ref>.
== Function ==
The pathway of stx entering a cell begins with the B sub-unit’s binding to GB3. Once this occurs, the A sub-unit disconnects from the B sub-unit and enters the cell through endocytosis. Using retrograde transport the A sub-unit passes through the Golgi apparatus and the rough endoplasmic reticulum<ref name="Melton">Melton-Celsa, Angela R. “Shiga Toxin (Stx) Classification, Structure, and Function.” Microbiology spectrum 2.2 (2014): 10.1128/microbiolspec.EHEC–0024–2013. PMC. Web</ref> In the rough endoplasmic reticulum, the A sub-unit is cleaved into two parts, A1 and A2, through the cleavage of trypsin sensitive residues and the reduction of a disulfide bridge. A2 is degraded, but A1 freely enters the cytosol<ref name="Sandvig">Sandvig, K., & van Deurs, B. (2000). Entry of ricin and Shiga toxin into cells: molecular mechanisms and medical perspectives. The EMBO Journal, 19(22), 5943–5950. doi:10.1093/emboj/19.22.5943</ref>. Once in the cytosol, A1 acts as an N-glycosidase, hydrolyzing bonds that link sugars. With this enzymatic activity, A1 removes an adenine from the alpha-sarcin loop in the 28S RNA of the 60S ribosomal sub-unit<ref name="Melton">Melton-Celsa, Angela R. “Shiga Toxin (Stx) Classification, Structure, and Function.” Microbiology spectrum 2.2 (2014): 10.1128/microbiolspec.EHEC–0024–2013. PMC. Web</ref>. The removal of the adenine prevents elongation factors from associating with the ribosomal sub-unit. Without elongation factors, the ribosome can no longer synthesize proteins, leading to cell death<ref name="Melton">Melton-Cesla, A. (2012). Shiga toxin classification structure and function. Changes, 29(2), 997–1003</ref>.
== Disease and Pathogenesis ==
The shiga toxin is produced by ''Shigella dysenteriae'' and certain strains of ''Escherichia coli''. ''S. dysenteriae'' is a gram negative bacterium that causes disease in its host by entering into intestinal epithelial cells. It has a very low infectious dose of only ten to one hundred cells, and is contracted through the consumption of contaminated water or food. Once in intestines, the bacteria's proliferation and secretion of stx erodes the intestinal lining and leads to dysentery<ref>Zaidi, Mussaret Bano, and Teresa Estrada-García. "Shigella: A Highly Virulent and Elusive pathogen." Current Tropical Medicine Reports 1.2 (2014): 81-87</ref>. Those with dysentery can simply have diarrhea as a symptom, but if the infection progresses, it can lead to hemorrhagic colitis (bloody stool) or hemolytic uremic syndrome (HUS). HUS is characterized as acute renal failure, thrombocytopenia, and microangiopathic anemia<ref>Kaper, James B. and Alison D. O’Brien. “Overview and Historical Perspectives.” Microbiology spectrum 2.2 (2014): 10.1128/microbiolspec.EHEC–0028–2014. PMC. Web</ref>.

As stated previously, stx enters cells through its binding to GB3, but curiously many intestinal cells do not express GB3. Some researchers hypothesize it enters the cells through macropinocyosis, but when macropinocytotic function is removed from cells, stx still manages to enter through an as yet undefined mechanism.

The genes necessary to produce the toxin are interestingly not native to ''S. dysenteriae''. The genes for stx are carried by prophages: bacteria that have had viral DNA inserted into their genome. During certain conditions, the phage genes are expressed leading to the production of more bacteriophages as well as the shiga toxin. When the bacteria undergo lysis, bacteriophage and stx are released. It may seem as if there is no benefit for ''S. dysenteriae'' to carry the viral DNA, but studies have shown that while it is in the lysogenic cycle S. dysenteriae has increased acid tolerance and motility<ref>Krüger, Alejandra, and Paula María Alejandra Lucchesi. "Shiga toxins and stx-phages: highly diverse entities." Microbiology (2014): mic-0</ref>.

</StructureSection>
== References ==
<references/>

Sandbox Reserved 982

2015-05-01T19:51:43Z

Joseph P. Hoffmann:

{{Telford_CHEM321}}
==Shiga Toxin==
<StructureSection load='1R4Q' size='340' side='right' caption='Crystal Structure for Shiga Toxin (1R4Q)' scene=''>

== Structural Highlights ==
Shiga toxin (stx) has multiple classifications. The shiga toxin produced by ''Shigella dysenteriae'' is classified as stx, while shiga toxin produced by Escherichia coli is classified as either stx1 or stx2. All three classifications vary slightly in structure but have identical functions. The A sub-unit of stx1 differs from stx by one residue: a serine in position 45 as opposed to threonine<ref name="Fraser">Fraser, M. E., Fujinaga, M., Cherney, M. M., Melton-Celsa, A. R., Twiddy, E. M., O’Brien, A. D., & James, M. N. G. (2004). Structure of shiga toxin type 2 (Stx2) from Escherichia coli O157:H7. The Journal of Biological Chemistry, 279(26), 27511–27517. doi:10.1074/jbc.M401939200</ref>.

Stx is a 70 kDa AB5 protein, meaning that it is composed of an A sub-unit bound to a B pentamer. A noncovalent interaction causes the association between the A and B sub-units, as the carboxy terminal tail of the A sub-unit is surrounded by the B pentamer. <scene name='68/687332/Sub-unit_a/2'>The A sub-unit</scene> is 293 amino acids long with its active site being <scene name='68/687332/Active_site_for_stx/1'>glutamic acid 167 (1R4Q)</scene>. Though the active site contains a single residue, studies have shown that the first 239 residues are essential for the enzymatic activity of the A sub-unit. Residues 240-251 are essential for the A sub-unit’s translocation from the endoplasmic reticulum of an infected cell to the cytosol<ref name="Sandvig">Sandvig, K., & van Deurs, B. (2000). Entry of ricin and Shiga toxin into cells: molecular mechanisms and medical perspectives. The EMBO Journal, 19(22), 5943–5950. doi:10.1093/emboj/19.22.5943</ref>.

The A sub-unit can be broken down into A1 and A2 units. This break down occurs when a trypsin sensitive region, residues 248-251, is cleaved. This cleavage results in the only remaining link being a <scene name='68/687332/Disulfide_bridge/1'>disulfide bridge (1R4Q)</scene> between cysteine 242 and cysteine 261. If the disulfide bridge is reduced, the A1 and A2 fragments can completely separate. Interestingly, the disulfide bridge blocks the active site of the A sub-unit, so the A sub-unit is not enzymatically active unless it has been cleaved<ref name ="Fraser">Fraser, M. E., Fujinaga, M., Cherney, M. M., Melton-Celsa, A. R., Twiddy, E. M., O’Brien, A. D., & James, M. N. G. (2004). Structure of shiga toxin type 2 (Stx2) from Escherichia coli O157:H7. The Journal of Biological Chemistry, 279(26), 27511–27517. doi:10.1074/jbc.M401939200</ref>.

<scene name='68/687332/B_sub-unit/1'>The B sub-unit (2XSC)</scene> is a symmetrical pentamer that resembles a star and can be divided into five identical monomers. Each monomer is 69 amino acids in length. The purpose of the B sub-unit is to bind to globotriaosylceramide (GB3) which is a glycosphingolipid that is found on the lipid rafts of endothelial cells. Each monomer of the B sub-unit has three binding sites for GB3, binding to the <scene name='68/687332/Gb3_bound_to_sub-unit_b/1'> carbohydrate portion of GB3 (1CQF)</scene>. The affinity between the B sub-unit and GB3 is incredibly low, and is actually one of the lowest recorded affinities for carbohydrate-protein interaction<ref>Jacobson, J. M., Yin, J., Kitov, P. I., Mulvey, G., Griener, T. P., James, M. N. G., … Bundle, D. R. (2014). The crystal structure of Shiga toxin type 2 with bound disaccharide guides the design of a heterobifunctional toxin inhibitor. Journal of Biological Chemistry, 289(2), 885–894. doi:10.1074/jbc.M113.518886</ref>. Some studies have shown that the level of saturation and length of the fatty acid on GB3 affects the strength of its interaction with the B sub-unit. Since the B sub-unit of stx has fifteen binding sites though, it has an incredibly high avidity<ref name="Melton">Melton-Celsa, Angela R. “Shiga Toxin (Stx) Classification, Structure, and Function.” Microbiology spectrum 2.2 (2014): 10.1128/microbiolspec.EHEC–0024–2013. PMC. Web</ref>.
== Function ==
The pathway of stx entering a cell begins with the B sub-unit’s binding to GB3. Once this occurs, the A sub-unit disconnects from the B sub-unit and enters the cell through endocytosis. Using retrograde transport the A sub-unit passes through the Golgi apparatus and the rough endoplasmic reticulum<ref name="Melton">Melton-Celsa, Angela R. “Shiga Toxin (Stx) Classification, Structure, and Function.” Microbiology spectrum 2.2 (2014): 10.1128/microbiolspec.EHEC–0024–2013. PMC. Web</ref> In the rough endoplasmic reticulum, the A sub-unit is cleaved into two parts, A1 and A2, through the cleavage of trypsin sensitive residues and the reduction of a disulfide bridge. A2 is degraded, but A1 freely enters the cytosol<ref name="Sandvig">Sandvig, K., & van Deurs, B. (2000). Entry of ricin and Shiga toxin into cells: molecular mechanisms and medical perspectives. The EMBO Journal, 19(22), 5943–5950. doi:10.1093/emboj/19.22.5943</ref>. Once in the cytosol, A1 acts as an N-glycosidase, hydrolyzing bonds that link sugars. With this enzymatic activity, A1 removes an adenine from the alpha-sarcin loop in the 28S RNA of the 60S ribosomal sub-unit<ref name="Melton">Melton-Celsa, Angela R. “Shiga Toxin (Stx) Classification, Structure, and Function.” Microbiology spectrum 2.2 (2014): 10.1128/microbiolspec.EHEC–0024–2013. PMC. Web</ref>. The removal of the adenine prevents elongation factors from associating with the ribosomal sub-unit. Without elongation factors, the ribosome can no longer synthesize proteins, leading to cell death<ref name="Melton">Melton-Cesla, A. (2012). Shiga toxin classification structure and function. Changes, 29(2), 997–1003</ref>.
== Disease and Pathogenesis ==
The shiga toxin is produced by ''Shigella dysenteriae'' and certain strains of ''Escherichia coli''. ''S. dysenteriae'' is a gram negative bacterium that causes disease in its host by entering into intestinal epithelial cells. It has a very low infectious dose of only ten to one hundred cells, and is contracted through the consumption of contaminated water or food. Once in intestines, the bacteria's proliferation and secretion of stx erodes the intestinal lining and leads to dysentery<ref>Zaidi, Mussaret Bano, and Teresa Estrada-García. "Shigella: A Highly Virulent and Elusive pathogen." Current Tropical Medicine Reports 1.2 (2014): 81-87</ref>. Those with dysentery can simply have diarrhea as a symptom, but if the infection progresses, it can lead to hemorrhagic colitis (bloody stool) or hemolytic uremic syndrome (HUS). HUS is characterized as acute renal failure, thrombocytopenia, and microangiopathic anemia<ref>Kaper, James B. and Alison D. O’Brien. “Overview and Historical Perspectives.” Microbiology spectrum 2.2 (2014): 10.1128/microbiolspec.EHEC–0028–2014. PMC. Web</ref>.

As stated previously, stx enters cells through its binding to GB3, but curiously many intestinal cells do not express GB3. Some researchers hypothesize it enters the cells through macropinocyosis, but when macropinocytotic function is removed from cells, stx still manages to enter through an as yet undefined mechanism.

The genes necessary to produce the toxin are interestingly not native to ''S. dysenteriae''. The genes for stx are carried by prophages: bacteria that have had viral DNA inserted into their genome. During certain conditions, the phage genes are expressed leading to the production of more bacteriophages as well as the shiga toxin. When the bacteria undergo lysis, bacteriophage and stx are released. It may seem as if there is no benefit for ''S. dysenteriae'' to carry the viral DNA, but studies have shown that while it is in the lysogenic cycle S. dysenteriae has increased acid tolerance and motility<ref>Krüger, Alejandra, and Paula María Alejandra Lucchesi. "Shiga toxins and stx-phages: highly diverse entities." Microbiology (2014): mic-0</ref>.

</StructureSection>
== References ==
<references/>

Sandbox Reserved 982

2015-05-01T19:40:56Z

Joseph P. Hoffmann:

{{Telford_CHEM321}}
==Shiga Toxin==
<StructureSection load='1R4Q' size='340' side='right' caption='Crystal Structure for Shiga Toxin (1R4Q)' scene=''>

== Structural Highlights ==
Shiga toxin (stx) has multiple classifications. The shiga toxin produced by ''Shigella dysenteriae'' is classified as stx, while shiga toxin produced by Escherichia coli can be classified as either stx1 or stx2. All three classifications very slightly in structure but have identical functions. The A sub-unit of stx1 differs from stx by only one residue: a serine in position 45 as opposed to threonine<ref name="Fraser">Fraser, M. E., Fujinaga, M., Cherney, M. M., Melton-Celsa, A. R., Twiddy, E. M., O’Brien, A. D., & James, M. N. G. (2004). Structure of shiga toxin type 2 (Stx2) from Escherichia coli O157:H7. The Journal of Biological Chemistry, 279(26), 27511–27517. doi:10.1074/jbc.M401939200</ref>.

Stx is a 70 kDa AB5 protein, meaning that it is composed of an A sub-unit bound to a B pentamer. A noncovalent interaction causes the association between the A and B sub-units, as the carboxy terminal tail of the A sub-unit is surrounded by the B pentamer. <scene name='68/687332/Sub-unit_a/2'>The A sub-unit</scene> is 293 amino acids long with its active site being <scene name='68/687332/Active_site_for_stx/1'>glutamic acid 167 (1R4Q)</scene>. Though the active site is a single residue, studies have shown that the first 239 residues are essential for the enzymatic activity of the A sub-unit. Residues 240-251 are essential for the A sub-unit’s translocation from the endoplasmic reticulum of an infected cell to the cytosol<ref name="Sandvig">Sandvig, K., & van Deurs, B. (2000). Entry of ricin and Shiga toxin into cells: molecular mechanisms and medical perspectives. The EMBO Journal, 19(22), 5943–5950. doi:10.1093/emboj/19.22.5943</ref>.

The A sub-unit can be broken down into A1 and A2 units. This break down occurs when a trypsin sensitive region, residues 248-251, is cleaved. This cleavage results in the only remaining link being a <scene name='68/687332/Disulfide_bridge/1'>disulfide bridge (1R4Q)</scene> between cysteine 242 and cysteine 261. If the disulfide bridge is reduced, the A1 and A2 fragments can completely separate. Interestingly, the disulfide bridge blocks the active site of the A sub-unit, so the A sub-unit is not enzymatically active unless it has been cleaved<ref name ="Fraser">Fraser, M. E., Fujinaga, M., Cherney, M. M., Melton-Celsa, A. R., Twiddy, E. M., O’Brien, A. D., & James, M. N. G. (2004). Structure of shiga toxin type 2 (Stx2) from Escherichia coli O157:H7. The Journal of Biological Chemistry, 279(26), 27511–27517. doi:10.1074/jbc.M401939200</ref>.

<scene name='68/687332/B_sub-unit/1'>The B sub-unit (2XSC)</scene> is a symmetrical pentamer that resembles a star and can be divided into five identical monomers. Each of the monomers is 69 amino acids in length. The purpose of the B sub-unit is to bind to globotriaosylceramide (GB3) which is a glycosphingolipid that is found on the lipid rafts of endothelial cells. Each monomer of the B sub-unit has three binding sites for GB3, binding to the <scene name='68/687332/Gb3_bound_to_sub-unit_b/1'> carbohydrate portion of GB3 (1CQF)</scene>. The affinity between the B sub-unit and GB3 is incredibly low, and is actually one of the lowest recorded affinities for carbohydrate-protein interaction<ref>Jacobson, J. M., Yin, J., Kitov, P. I., Mulvey, G., Griener, T. P., James, M. N. G., … Bundle, D. R. (2014). The crystal structure of Shiga toxin type 2 with bound disaccharide guides the design of a heterobifunctional toxin inhibitor. Journal of Biological Chemistry, 289(2), 885–894. doi:10.1074/jbc.M113.518886</ref>. Some studies have shown that the level of saturation and length of the fatty acid on GB3 affects the strength of its interaction with the B sub-unit. Since the B sub-unit of stx has fifteen binding sites though, it has an incredibly high avidity<ref name="Melton">Melton-Celsa, Angela R. “Shiga Toxin (Stx) Classification, Structure, and Function.” Microbiology spectrum 2.2 (2014): 10.1128/microbiolspec.EHEC–0024–2013. PMC. Web</ref>.
== Function ==
The pathway of stx entering a cell begins with the B sub-unit’s binding to GB3. Once this occurs, the A sub-unit disconnects from the B sub-unit and enters the cell through endocytosis. Using retrograde transport the A sub-unit passes through the Golgi apparatus and the rough endoplasmic reticulum<ref name="Melton">Melton-Celsa, Angela R. “Shiga Toxin (Stx) Classification, Structure, and Function.” Microbiology spectrum 2.2 (2014): 10.1128/microbiolspec.EHEC–0024–2013. PMC. Web</ref> In the rough endoplasmic reticulum, the A sub-unit is split into two parts called A1 and A2 through the cleavage of trypsin sensitive residues and the reduction of a disulfide bridge. A2 is degraded, but A1 freely enters the cytosol<ref name="Sandvig">Sandvig, K., & van Deurs, B. (2000). Entry of ricin and Shiga toxin into cells: molecular mechanisms and medical perspectives. The EMBO Journal, 19(22), 5943–5950. doi:10.1093/emboj/19.22.5943</ref>. Once in the cytosol, A1 acts as an N-glycosidase, which is an enzyme that hydrolyzes bonds that link sugars. With this enzymatic activity, A1 removes an adenine from the alpha-sarcin loop in the 28S RNA of the 60S ribosomal sub-unit<ref name="Melton">Melton-Celsa, Angela R. “Shiga Toxin (Stx) Classification, Structure, and Function.” Microbiology spectrum 2.2 (2014): 10.1128/microbiolspec.EHEC–0024–2013. PMC. Web</ref>. The removal of the adenine prevents elongation factors from associating with the ribosomal sub-unit. Without elongation factors, the ribosome can no longer synthesize proteins, leading to cell death<ref name="Melton">Melton-Cesla, A. (2012). Shiga toxin classification structure and function. Changes, 29(2), 997–1003</ref>.
== Disease and Pathogenesis ==
The shiga toxin is produced by ''Shigella dysenteriae'' and certain strains of ''Escherichia coli''. ''S. dysenteriae'' is a gram negative bacterium that causes disease in its host by entering into intestinal epithelial cells. It has a very low infectious dose of only ten to one hundred cells, and is contracted through the consumption of contaminated water or food. One it begins replicating in the intestines, its proliferation and secretion of stx erodes the intestinal lining and leads to dysentery<ref>Zaidi, Mussaret Bano, and Teresa Estrada-García. "Shigella: A Highly Virulent and Elusive pathogen." Current Tropical Medicine Reports 1.2 (2014): 81-87</ref>. Those with dysentery can simply have diarrhea as a symptom, but if the infection progresses, it can lead to hemorrhagic colitis (bloody stool) or hemolytic uremic syndrome (HUS). HUS is characterized as acute renal failure, thrombocytopenia, and microangiopathic anemia<ref>Kaper, James B. and Alison D. O’Brien. “Overview and Historical Perspectives.” Microbiology spectrum 2.2 (2014): 10.1128/microbiolspec.EHEC–0028–2014. PMC. Web</ref>.

As stated previously, stx enters cells through its binding to GB3, but curiously many intestinal cells do not express GB3. Some researchers hypothesize it enters the cells through macropinocyosis, but when macropinocytotic function is removed from cells, stx still manages to enter through an as yet undefined mechanism.

The genes necessary to produce the toxin are interestingly not native to ''S. dysenteriae''. The genes for stx are carried by prophages: bacteria that have had viral DNA inserted into their genome. During certain conditions, the phage genes are expressed leading to the production of more bacteriophages as well as the shiga toxin. When the bacteria undergo lysis, the bacteriophages and stx are released. It may seem as if there is no benefit for ''S. dysenteriae'' to carry the viral DNA, but studies have shown that while it is in the lysogenic cycle S. dysenteriae has increased acid tolerance and motility<ref>Krüger, Alejandra, and Paula María Alejandra Lucchesi. "Shiga toxins and stx-phages: highly diverse entities." Microbiology (2014): mic-0</ref>.

</StructureSection>
== References ==
<references/>

Sandbox Reserved 982

2015-05-01T19:38:54Z

Joseph P. Hoffmann:

{{Telford_CHEM321}}
==Shiga Toxin==
<StructureSection load='1R4Q' size='340' side='right' caption='Crystal Structure for Shiga Toxin (1R4Q)' scene=''>

== Structural Highlights ==
Shiga toxin (stx) has multiple classifications. The shiga toxin produced by ''Shigella dysenteriae'' is classified as stx, while shiga toxin produced by Escherichia coli can be classified as either stx1 or stx2. All three classifications very slightly in structure but have identical functions. The A sub-unit of stx1 differs from stx by only one residue: a serine in position 45 as opposed to threonine<ref name="Fraser">Fraser, M. E., Fujinaga, M., Cherney, M. M., Melton-Celsa, A. R., Twiddy, E. M., O’Brien, A. D., & James, M. N. G. (2004). Structure of shiga toxin type 2 (Stx2) from Escherichia coli O157:H7. The Journal of Biological Chemistry, 279(26), 27511–27517. doi:10.1074/jbc.M401939200</ref>.

Stx is a 70 kDa AB5 protein, meaning that it is composed of an A sub-unit bound to a B pentamer. A noncovalent interaction causes the association between the A and B sub-units, as the carboxy terminal tail of the A sub-unit is surrounded by the B pentamer. <scene name='68/687332/Sub-unit_a/2'>The A sub-unit</scene> is 293 amino acids long with its active site being <scene name='68/687332/Active_site_for_stx/1'>glutamic acid 167 (1R4Q)</scene>. Though the active site is a single residue, studies have shown that the first 239 residues are essential for the enzymatic activity of the A sub-unit. Residues 240-251 are essential for the A sub-unit’s translocation from the endoplasmic reticulum of an infected cell to the cytosol<ref name="Sandvig">Sandvig, K., & van Deurs, B. (2000). Entry of ricin and Shiga toxin into cells: molecular mechanisms and medical perspectives. The EMBO Journal, 19(22), 5943–5950. doi:10.1093/emboj/19.22.5943</ref>.

The A sub-unit can be broken down into A1 and A2 units. This break down occurs when a trypsin sensitive region, residues 248-251, is cleaved. This cleavage results in the only remaining link being a <scene name='68/687332/Disulfide_bridge/1'>disulfide bridge (1R4Q)</scene> between cysteine 242 and cysteine 261. If the disulfide bridge is reduced, the A1 and A2 fragments can completely separate. Interestingly, the disulfide bridge blocks the active site of the A sub-unit, so the A sub-unit is not enzymatically active unless it has been cleaved<ref name ="Fraser">Fraser, M. E., Fujinaga, M., Cherney, M. M., Melton-Celsa, A. R., Twiddy, E. M., O’Brien, A. D., & James, M. N. G. (2004). Structure of shiga toxin type 2 (Stx2) from Escherichia coli O157:H7. The Journal of Biological Chemistry, 279(26), 27511–27517. doi:10.1074/jbc.M401939200</ref>.

<scene name='68/687332/B_sub-unit/1'>The B sub-unit (2XSC)</scene> is a symmetrical pentamer that resembles a star and can be divided into five identical monomers. Each of the monomers is 69 amino acids in length. The purpose of the B sub-unit is to bind to globotriaosylceramide (GB3) which is a glycosphingolipid that is found on the lipid rafts of endothelial cells. Each monomer of the B sub-unit has three binding sites for GB3, binding to the <scene name='68/687332/Gb3_bound_to_sub-unit_b/1'> carbohydrate portion of GB3 (1CQF)</scene>. The affinity between the B sub-unit and GB3 is incredibly low, and is actually one of the lowest recorded affinities for carbohydrate-protein interaction<ref>Jacobson, J. M., Yin, J., Kitov, P. I., Mulvey, G., Griener, T. P., James, M. N. G., … Bundle, D. R. (2014). The crystal structure of Shiga toxin type 2 with bound disaccharide guides the design of a heterobifunctional toxin inhibitor. Journal of Biological Chemistry, 289(2), 885–894. doi:10.1074/jbc.M113.518886</ref>. Some studies have shown that the level of saturation and length of the fatty acid on GB3 affects the strength of its interaction with the B sub-unit. Since the B sub-unit of stx has fifteen binding sites though, it has an incredibly high avidity<ref name="Melton">Melton-Celsa, Angela R. “Shiga Toxin (Stx) Classification, Structure, and Function.” Microbiology spectrum 2.2 (2014): 10.1128/microbiolspec.EHEC–0024–2013. PMC. Web</ref>.
== Function ==
The pathway of stx entering a cell begins with the B sub-unit’s binding to GB3. Once this occurs, the A sub-unit disconnects from the B sub-unit and enters the cell through endocytosis. Using retrograde transport the A sub-unit passes through the Golgi apparatus and the rough endoplasmic reticulum<ref name="Melton">Melton-Celsa, Angela R. “Shiga Toxin (Stx) Classification, Structure, and Function.” Microbiology spectrum 2.2 (2014): 10.1128/microbiolspec.EHEC–0024–2013. PMC. Web</ref> In the rough endoplasmic reticulum, the A sub-unit is split into two parts called A1 and A2 through the cleavage of trypsin sensitive residues and the reduction of a disulfide bridge. A2 is degraded, but A1 freely enters the cytosol<ref name="Sandvig">Sandvig, K., & van Deurs, B. (2000). Entry of ricin and Shiga toxin into cells: molecular mechanisms and medical perspectives. The EMBO Journal, 19(22), 5943–5950. doi:10.1093/emboj/19.22.5943</ref>. Once in the cytosol, A1 acts as an N-glycosidase, which is an enzyme that hydrolyzes bonds that link sugars. With this enzymatic activity, A1 removes an adenine from the alpha-sarcin loop in the 28S RNA of the 60S ribosomal sub-unit<ref name="Melton">Melton-Celsa, Angela R. “Shiga Toxin (Stx) Classification, Structure, and Function.” Microbiology spectrum 2.2 (2014): 10.1128/microbiolspec.EHEC–0024–2013. PMC. Web</ref>. The removal of the adenine prevents elongation factors from associating with the ribosomal sub-unit. Without elongation factors, the ribosome can no longer synthesize proteins, leading to cell death<ref name="Melton">Melton-Cesla, A. (2012). Shiga toxin classification structure and function. Changes, 29(2), 997–1003</ref>.
== Disease and Pathogenesis ==
The shiga toxin is produced by ''Shigella dysenteriae'' and certain strains of ''Escherichia coli''. ''S. dysenteriae'' is a gram negative bacterium that causes disease in its host by entering into intestinal epithelial cells. It has a very low infectious dose of only ten to one hundred cells, and is contracted through the consumption of contaminated water or food. One it begins replicating in the intestines, its proliferation and secretion of stx erodes the intestinal lining and leads to dysentery<ref>Zaidi, Mussaret Bano, and Teresa Estrada-García. "Shigella: A Highly Virulent and Elusive pathogen." Current Tropical Medicine Reports 1.2 (2014): 81-87</ref>. Those with dysentery can simply have diarrhea as a symptom, but if the infection progresses, it can lead to hemorrhagic colitis (bloody stool) or hemolytic uremic syndrome (HUS). HUS is characterized as acute renal failure, thrombocytopenia, and microangiopathic anemia<ref>Kaper, James B. and Alison D. O’Brien. “Overview and Historical Perspectives.” Microbiology spectrum 2.2 (2014): 10.1128/microbiolspec.EHEC–0028–2014. PMC. Web</ref>.

As stated previously, stx enters cells through its binding to GB3, but curiously many intestinal cells do not express GB3. Some researchers hypothesize it enters the cells through macropinocyosis, but when macropinocytotic function is removed from cells, stx still manages to enter through an as yet undefined mechanism.

The genes necessary to produce the toxin are interestingly not native to ''S. dysenteriae''. The genes for stx are carried by prophages: bacteria that have had viral DNA inserted into their genome. During certain conditions, the phage genes are expressed leading to the production of more bacteriophages as well as the shiga toxin. When the bacteria undergo lysis, the bacteriophages and stx are released. It may seem as if there is no benefit for ''S. dysenteriae'' to carry the viral DNA, but studies have shown that while it is in the lysogenic cycle S. dysenteriae has increased acid tolerance and motility<ref>Krüger, Alejandra, and Paula María Alejandra Lucchesi. "Shiga toxins and stx-phages: highly diverse entities." Microbiology (2014): mic-0</ref>.

</StructureSection>
== References ==
<references/>

Sandbox Reserved 982

2015-05-01T16:57:50Z

Joseph P. Hoffmann:

{{Telford_CHEM321}}
==Shiga Toxin==
<StructureSection load='1R4Q' size='340' side='right' caption='Crystal Structure for Shiga Toxin (1R4Q)' scene=''>

== Structural Highlights ==
Shiga toxin (stx) has multiple classifications. The shiga toxin produced by ''Shigella dysenteriae'' is classified as stx, while shiga toxin produced by Escherichia coli can be classified as either stx1 or stx2. All three classifications very slightly in structure but have identical functions. The A sub-unit of stx1 differs from stx by only one residue: a serine in position 45 as opposed to threonine<ref name="Fraser">Fraser, M. E., Fujinaga, M., Cherney, M. M., Melton-Celsa, A. R., Twiddy, E. M., O’Brien, A. D., & James, M. N. G. (2004). Structure of shiga toxin type 2 (Stx2) from Escherichia coli O157:H7. The Journal of Biological Chemistry, 279(26), 27511–27517. doi:10.1074/jbc.M401939200</ref>. Stx is a 70 kDa AB5 protein, meaning that it is composed of an A sub-unit bound to a B pentamer. A noncovalent interaction causes the association between the A and B sub-units, as the carboxy terminal tail of the A sub-unit is surrounded by the B pentamer. <scene name='68/687332/Sub-unit_a/2'>The A sub-unit</scene> is 293 amino acids long with its active site being <scene name='68/687332/Active_site_for_stx/1'>glutamic acid 167 (1R4Q)</scene>. Though the active site is a single residue, studies have shown that the first 239 residues are essential for the enzymatic activity of the A sub-unit. Residues 240-251 are essential for the A sub-unit’s translocation from the endoplasmic reticulum of an infected cell to the cytosol<ref name="Sandvig">Sandvig, K., & van Deurs, B. (2000). Entry of ricin and Shiga toxin into cells: molecular mechanisms and medical perspectives. The EMBO Journal, 19(22), 5943–5950. doi:10.1093/emboj/19.22.5943</ref>. The A sub-unit can be broken down into A1 and A2 units. This break down occurs when a trypsin sensitive region, residues 248-251, is cleaved. This cleavage results in the only remaining link being a <scene name='68/687332/Disulfide_bridge/1'>disulfide bridge (1R4Q)</scene> between cysteine 242 and cysteine 261. If the disulfide bridge is reduced, the A1 and A2 fragments can completely separate. Interestingly, the disulfide bridge blocks the active site of the A sub-unit, so the A sub-unit is not enzymatically active unless it has been cleaved<ref name ="Fraser">Fraser, M. E., Fujinaga, M., Cherney, M. M., Melton-Celsa, A. R., Twiddy, E. M., O’Brien, A. D., & James, M. N. G. (2004). Structure of shiga toxin type 2 (Stx2) from Escherichia coli O157:H7. The Journal of Biological Chemistry, 279(26), 27511–27517. doi:10.1074/jbc.M401939200</ref>.
<scene name='68/687332/B_sub-unit/1'>The B sub-unit (2XSC)</scene> is a symmetrical pentamer that resembles a star and can be divided into five identical monomers. Each of the monomers is 69 amino acids in length. The purpose of the B sub-unit is to bind to globotriaosylceramide (GB3) which is a glycosphingolipid that is found on the lipid rafts of endothelial cells. Each monomer of the B sub-unit has three binding sites for GB3, binding to the <scene name='68/687332/Gb3_bound_to_sub-unit_b/1'> carbohydrate portion of GB3 (1CQF)</scene>. The affinity between the B sub-unit and GB3 is incredibly low, and is actually one of the lowest recorded affinities for carbohydrate-protein interaction<ref>Jacobson, J. M., Yin, J., Kitov, P. I., Mulvey, G., Griener, T. P., James, M. N. G., … Bundle, D. R. (2014). The crystal structure of Shiga toxin type 2 with bound disaccharide guides the design of a heterobifunctional toxin inhibitor. Journal of Biological Chemistry, 289(2), 885–894. doi:10.1074/jbc.M113.518886</ref>. Some studies have shown that the level of saturation and length of the fatty acid on GB3 affects the strength of its interaction with the B sub-unit. Since the B sub-unit of stx has fifteen binding sites though, it has an incredibly high avidity<ref name="Melton">Melton-Celsa, Angela R. “Shiga Toxin (Stx) Classification, Structure, and Function.” Microbiology spectrum 2.2 (2014): 10.1128/microbiolspec.EHEC–0024–2013. PMC. Web</ref>.
== Function ==
The pathway of stx entering a cell begins with the B sub-unit’s binding to GB3. Once this occurs, the A sub-unit disconnects from the B sub-unit and enters the cell through endocytosis. Using retrograde transport the A sub-unit passes through the Golgi apparatus and the rough endoplasmic reticulum<ref name="Melton">Melton-Celsa, Angela R. “Shiga Toxin (Stx) Classification, Structure, and Function.” Microbiology spectrum 2.2 (2014): 10.1128/microbiolspec.EHEC–0024–2013. PMC. Web</ref> In the rough endoplasmic reticulum, the A sub-unit is split into two parts called A1 and A2 through the cleavage of trypsin sensitive residues and the reduction of a disulfide bridge. A2 is degraded, but A1 freely enters the cytosol<ref name="Sandvig">Sandvig, K., & van Deurs, B. (2000). Entry of ricin and Shiga toxin into cells: molecular mechanisms and medical perspectives. The EMBO Journal, 19(22), 5943–5950. doi:10.1093/emboj/19.22.5943</ref>. Once in the cytosol, A1 acts as an N-glycosidase, which is an enzyme that hydrolyzes bonds that link sugars. With this enzymatic activity, A1 removes an adenine from the alpha-sarcin loop in the 28S RNA of the 60S ribosomal sub-unit<ref name="Melton">Melton-Celsa, Angela R. “Shiga Toxin (Stx) Classification, Structure, and Function.” Microbiology spectrum 2.2 (2014): 10.1128/microbiolspec.EHEC–0024–2013. PMC. Web</ref>. The removal of the adenine prevents elongation factors from associating with the ribosomal sub-unit. Without elongation factors, the ribosome can no longer synthesize proteins, leading to cell death<ref name="Melton">Melton-Cesla, A. (2012). Shiga toxin classification structure and function. Changes, 29(2), 997–1003</ref>.
== Disease and Pathogenesis ==
The shiga toxin is produced by ''Shigella dysenteriae'' and certain strains of ''Escherichia coli''. ''S. dysenteriae'' is a gram negative bacterium that causes disease in its host by entering into intestinal epithelial cells. It has a very low infectious dose of only ten to one hundred cells, and is contracted through the consumption of contaminated water or food. One it begins replicating in the intestines, its proliferation and secretion of stx erodes the intestinal lining and leads to dysentery<ref>Zaidi, Mussaret Bano, and Teresa Estrada-García. "Shigella: A Highly Virulent and Elusive pathogen." Current Tropical Medicine Reports 1.2 (2014): 81-87</ref>. Those with dysentery can simply have diarrhea as a symptom, but if the infection progresses, it can lead to hemorrhagic colitis (bloody stool) or hemolytic uremic syndrome (HUS). HUS is characterized as acute renal failure, thrombocytopenia, and microangiopathic anemia<ref>Kaper, James B. and Alison D. O’Brien. “Overview and Historical Perspectives.” Microbiology spectrum 2.2 (2014): 10.1128/microbiolspec.EHEC–0028–2014. PMC. Web</ref>. As stated previously, stx enters cells through its binding to GB3, but curiously many intestinal cells do not express GB3. Some researchers hypothesize it enters the cells through macropinocyosis, but when macropinocytotic function is removed from cells, stx still manages to enter through an undefined mechanism.
The genes necessary to produce the toxin are interestingly not native to ''S. dysenteriae''. The genes for stx are carried by prophages: bacteria that have had viral DNA inserted into their genome. During certain conditions, the phage genes are expressed leading to the production of more bacteriophages as well as the shiga toxin. When the bacteria undergo lysis, the bacteriophages and stx are released. It may seem as if there is no benefit for ''S. dysenteriae'' to carry the viral DNA, but studies have shown that while it is in the lysogenic cycle S. dysenteriae has increased acid tolerance and motility<ref>Krüger, Alejandra, and Paula María Alejandra Lucchesi. "Shiga toxins and stx-phages: highly diverse entities." Microbiology (2014): mic-0</ref>.

</StructureSection>
== References ==
<references/>

Sandbox Reserved 982

2015-05-01T16:56:37Z

Joseph P. Hoffmann:

{{Telford_CHEM321}}
==Shiga Toxin==
<StructureSection load='1R4Q' size='340' side='right' caption='Crystal Structure for Shiga Toxin (1R4Q)' scene=''>

== Structural Highlights ==
Shiga toxin (stx) has multiple classifications. The shiga toxin produced by ''Shigella dysenteriae'' is classified as stx, while shiga toxin produced by Escherichia coli can be classified as either stx1 or stx2. All three classifications very slightly in structure but have identical functions. The A sub-unit of stx1 differs from stx by only one residue: a serine in position 45 as opposed to threonine<ref name="Fraser">Fraser, M. E., Fujinaga, M., Cherney, M. M., Melton-Celsa, A. R., Twiddy, E. M., O’Brien, A. D., & James, M. N. G. (2004). Structure of shiga toxin type 2 (Stx2) from Escherichia coli O157:H7. The Journal of Biological Chemistry, 279(26), 27511–27517. doi:10.1074/jbc.M401939200</ref>. Stx is a 70 kDa AB5 protein, meaning that it is composed of an A sub-unit bound to a B pentamer. A noncovalent interaction causes the association between the A and B sub-units, as the carboxy terminal tail of the A sub-unit is surrounded by the B pentamer. <scene name='68/687332/Sub-unit_a/2'>The A sub-unit</scene> is 293 amino acids long with its active site being <scene name='68/687332/Active_site_for_stx/1'>glutamic acid 167 (1R4Q)</scene>. Though the active site is a single residue, studies have shown that the first 239 residues are essential for the enzymatic activity of the A sub-unit. Residues 240-251 are essential for the A sub-unit’s translocation from the endoplasmic reticulum of an infected cell to the cytosol<ref name="Sandvig">Sandvig, K., & van Deurs, B. (2000). Entry of ricin and Shiga toxin into cells: molecular mechanisms and medical perspectives. The EMBO Journal, 19(22), 5943–5950. doi:10.1093/emboj/19.22.5943</ref>. The A sub-unit can be broken down into A1 and A2 units. This break down occurs when a trypsin sensitive region, residues 248-251, is cleaved. This cleavage results in the only remaining link being a <scene name='68/687332/Disulfide_bridge/1'>disulfide bridge (1R4Q)</scene> between cysteine 242 and cysteine 261. If the disulfide bridge is reduced, the A1 and A2 fragments can completely separate. Interestingly, the disulfide bridge blocks the active site of the A sub-unit, so the A sub-unit is not enzymatically active unless it has been cleaved<ref name ="Fraser">Fraser, M. E., Fujinaga, M., Cherney, M. M., Melton-Celsa, A. R., Twiddy, E. M., O’Brien, A. D., & James, M. N. G. (2004). Structure of shiga toxin type 2 (Stx2) from Escherichia coli O157:H7. The Journal of Biological Chemistry, 279(26), 27511–27517. doi:10.1074/jbc.M401939200</ref>.
<scene name='68/687332/B_sub-unit/1'>The B sub-unit (2XSC)</scene> is a symmetrical pentamer that resembles a star and can be divided into five identical monomers. Each of the monomers is 69 amino acids in length. The purpose of the B sub-unit is to bind to globotriaosylceramide (GB3) which is a glycosphingolipid that is found on the lipid rafts of endothelial cells. Each monomer of the B sub-unit has three binding sites for GB3, binding to the <scene name='68/687332/Gb3_bound_to_sub-unit_b/1'> carbohydrate portion of GB3 (1CQF)</scene>. The affinity between the B sub-unit and GB3 is incredibly low, and is actually one of the lowest recorded affinities for carbohydrate-protein interaction<ref>Jacobson, J. M., Yin, J., Kitov, P. I., Mulvey, G., Griener, T. P., James, M. N. G., … Bundle, D. R. (2014). The crystal structure of Shiga toxin type 2 with bound disaccharide guides the design of a heterobifunctional toxin inhibitor. Journal of Biological Chemistry, 289(2), 885–894. doi:10.1074/jbc.M113.518886</ref>. Some studies have shown that the level of saturation and length of the fatty acid on GB3 affects the strength of its interaction with the B sub-unit. Since the B sub-unit of stx has fifteen binding sites though, it has an incredibly high avidity<ref name="Melton">Melton-Celsa, Angela R. “Shiga Toxin (Stx) Classification, Structure, and Function.” Microbiology spectrum 2.2 (2014): 10.1128/microbiolspec.EHEC–0024–2013. PMC. Web</ref>.
== Function ==
The pathway of stx entering a cell begins with the B sub-unit’s binding to GB3. Once this occurs, the A sub-unit disconnects from the B sub-unit and enters the cell through endocytosis. Using retrograde transport the A sub-unit passes through the Golgi apparatus and the rough endoplasmic reticulum<ref name="Melton">Melton-Celsa, Angela R. “Shiga Toxin (Stx) Classification, Structure, and Function.” Microbiology spectrum 2.2 (2014): 10.1128/microbiolspec.EHEC–0024–2013. PMC. Web</ref> In the rough endoplasmic reticulum, the A sub-unit is split into two parts called A1 and A2 through the cleavage of trypsin sensitive residues and the reduction of a disulfide bridge. A2 is degraded, but A1 freely enters the cytosol<ref name="Sandvig">Sandvig, K., & van Deurs, B. (2000). Entry of ricin and Shiga toxin into cells: molecular mechanisms and medical perspectives. The EMBO Journal, 19(22), 5943–5950. doi:10.1093/emboj/19.22.5943</ref>. Once in the cytosol, A1 acts as an N-glycosidase, which is an enzyme that hydrolyzes bonds that link sugars. With this enzymatic activity, A1 removes an adenine from the alpha-sarcin loop in the 28S RNA of the 60S ribosomal sub-unit<ref name="Melton">Melton-Celsa, Angela R. “Shiga Toxin (Stx) Classification, Structure, and Function.” Microbiology spectrum 2.2 (2014): 10.1128/microbiolspec.EHEC–0024–2013. PMC. Web</ref>. The removal of the adenine prevents elongation factors from associating with the ribosomal sub-unit. Without elongation factors, the ribosome can no longer synthesize proteins, leading to cell death<ref name="Melton">Melton-Cesla, A. (2012). Shiga toxin classification structure and function. Changes, 29(2), 997–1003</ref>.
== Disease and Pathogenesis ==
The shiga toxin is produced by ''Shigella dysenteriae'' and certain strains of ''Escherichia coli''. ''S. dysenteriae'' is a gram negative bacterium that causes disease in its host by entering into intestinal epithelial cells. It has a very low infectious dose of only ten to one hundred cells, and is contracted through the consumption of contaminated water or food. One it begins replicating in the intestines, its proliferation and secretion of stx erodes the intestinal lining and leads to dysentery<ref>Zaidi, Mussaret Bano, and Teresa Estrada-García. "Shigella: A Highly Virulent and Elusive pathogen." Current Tropical Medicine Reports 1.2 (2014): 81-87</ref>. Those with dysentery can simply have diarrhea as a symptom, but if the infection progresses, it can lead to hemorrhagic colitis (bloody stool) or hemolytic uremic syndrome (HUS). HUS is characterized as acute renal failure, thrombocytopenia, and microangiopathic anemia<ref>Kaper, James B. and Alison D. O’Brien. “Overview and Historical Perspectives.” Microbiology spectrum 2.2 (2014): 10.1128/microbiolspec.EHEC–0028–2014. PMC. Web</ref>. As stated previously, stx enters cells through its binding to GB3, but curiously many intestinal cells do not express GB3. Some researchers hypothesize it enters the cells through macropinocyosis, but when macropinocytotic function is removed from cells, stx still manages to enter through an undefined mechanism.
The genes necessary to produce the toxin are interestingly not native to ''S. dysenteriae''. The genes for stx are carried by prophages: bacteria that have had viral DNA inserted into their genome. During certain conditions, the phage genes are expressed leading to the production of more bacteriophages as well as the shiga toxin. When the bacteria undergo lysis, the bacteriophages and stx are released. It may seem as if there is no benefit for ''S. dysenteriae'' to carry the viral DNA, but studies have shown that while it is in the lysogenic cycle S. dysenteriae has increased acid tolerance and motility<ref>Krüger, Alejandra, and Paula María Alejandra Lucchesi. "Shiga toxins and stx-phages: highly diverse entities." Microbiology (2014): mic-0</ref>.

This is a sample scene created with SAT to <scene name='68/687332/Scene_1/1'>TextToBeDisplayed</scene> by Group, and another to make <scene name="/12/3456/Sample/2">a transparent representation</scene> of the protein. You can make your own scenes on SAT starting from scratch or loading and editing one of these sample scenes.

</StructureSection>
== References ==
<references/>

Sandbox Reserved 982

2015-05-01T16:42:47Z

Joseph P. Hoffmann:

{{Telford_CHEM321}}
==Shiga Toxin==
<StructureSection load='1R4Q' size='340' side='right' caption='Crystal Structure for Shiga Toxin (1R4Q)' scene=''>

== Structural Highlights ==
Shiga toxin (stx) has multiple classifications. The shiga toxin produced by Shigella dysenteriae is classified as stx, while shiga toxin produced by Escherichia coli can be classified as either stx1 or stx2. All three classifications very slightly in structure but have identical functions. The A subunit of stx1 differs from stx by only one residue: a serine in position 45 as opposed to threonine<ref name="Fraser">Fraser, M. E., Fujinaga, M., Cherney, M. M., Melton-Celsa, A. R., Twiddy, E. M., O’Brien, A. D., & James, M. N. G. (2004). Structure of shiga toxin type 2 (Stx2) from Escherichia coli O157:H7. The Journal of Biological Chemistry, 279(26), 27511–27517. doi:10.1074/jbc.M401939200</ref>. Stx is a 70 kDa AB5 protein, meaning that it is composed of an A subunit bound to a B subunit pentamer. A noncovalent interaction causes the association between the A and B subunits, as the carboxy terminal tail of the A subunit is surrounded by the B pentamer. <scene name='68/687332/Sub-unit_a/2'>The A subunit</scene> is 293 amino acids long with its active site being <scene name='68/687332/Active_site_for_stx/1'>glutamic acid 167 (1R4Q)</scene>. Though the active site is a single residue, studies have shown that the first 239 residues are essential for the enzymatic activity of the A subunit. Residues 240-251 are essential for the A subunit’s translocation from the endoplasmic reticulum of an infected cell to the cytosol<ref name="Sandvig">Sandvig, K., & van Deurs, B. (2000). Entry of ricin and Shiga toxin into cells: molecular mechanisms and medical perspectives. The EMBO Journal, 19(22), 5943–5950. doi:10.1093/emboj/19.22.5943</ref>. The A subunit can be broken down into A1 and A2 units. This break down occurs when a trypsin sensitive region, residues 248-251, is cleaved. This cleavage results in the only remaining link being a <scene name='68/687332/Disulfide_bridge/1'>disulfide bridge (1R4Q)</scene> between cysteine 242 and cysteine 261. If the disulfide bridge is reduced, the A1 and A2 subunits can completely separate. Interestingly, the disulfide bridge blocks the active site of the A subunit, so the A subunit is not enzymatically active unless it has been cleaved<ref name ="Fraser">Fraser, M. E., Fujinaga, M., Cherney, M. M., Melton-Celsa, A. R., Twiddy, E. M., O’Brien, A. D., & James, M. N. G. (2004). Structure of shiga toxin type 2 (Stx2) from Escherichia coli O157:H7. The Journal of Biological Chemistry, 279(26), 27511–27517. doi:10.1074/jbc.M401939200</ref>.
<scene name='68/687332/B_subunit/1'>The B subunit (2XSC)</scene> is a symmetrical pentamer that resembles a star and can be divided into five identical subunits. Each of the subunits is 69 amino acids in length. The purpose of the B subunit is to bind to globotriaosylceramide (GB3) which is a glycosphingolipid that is found on the lipid rafts of endothelial cells. Each monomer of the B subunit has three binding sites for GB3, binding to the <scene name='68/687332/Gb3_bound_to_subunit_b/1'> carbohydrate portion of GB3 (1CQF)</scene>. The affinity between the B subunit and GB3 is incredibly low, and is actually one of the lowest recorded affinities for carbohydrate-protein interaction<ref>Jacobson, J. M., Yin, J., Kitov, P. I., Mulvey, G., Griener, T. P., James, M. N. G., … Bundle, D. R. (2014). The crystal structure of Shiga toxin type 2 with bound disaccharide guides the design of a heterobifunctional toxin inhibitor. Journal of Biological Chemistry, 289(2), 885–894. doi:10.1074/jbc.M113.518886</ref>. Some studies have shown that the level of saturation and length of the fatty acid on GB3 affects the strength of its interaction with the B subunit. Since the B subunit of stx has fifteen binding sites though, it has an incredibly high avidity<ref name="Melton">Melton-Celsa, Angela R. “Shiga Toxin (Stx) Classification, Structure, and Function.” Microbiology spectrum 2.2 (2014): 10.1128/microbiolspec.EHEC–0024–2013. PMC. Web</ref>.
== Function ==
The pathway of stx entering a cell begins with the B subunit’s binding to GB3. Once this occurs, the A subunit disconnects from the B subunit and enters the cell through endocytosis. Using retrograde transport the A subunit passes through the Golgi apparatus and the rough endoplasmic reticulum<ref name="Melton">Melton-Celsa, Angela R. “Shiga Toxin (Stx) Classification, Structure, and Function.” Microbiology spectrum 2.2 (2014): 10.1128/microbiolspec.EHEC–0024–2013. PMC. Web</ref> In the rough endoplasmic reticulum, the A subunit is split into two parts called A1 and A2 through the cleavage of trypsin sensitive residues and the reduction of a disulfide bridge. A2 is degraded, but A1 freely enters the cytosol<ref name="Sandvig">Sandvig, K., & van Deurs, B. (2000). Entry of ricin and Shiga toxin into cells: molecular mechanisms and medical perspectives. The EMBO Journal, 19(22), 5943–5950. doi:10.1093/emboj/19.22.5943</ref>. Once in the cytosol, A1 acts as an N-glycosidase, which is an enzyme that hydrolyzes bonds that link sugars. With this enzymatic activity, A1 removes an adenine from the alpha-sarcin loop in the 28S RNA of the 60S ribosomal subunit<ref name="Melton">Melton-Celsa, Angela R. “Shiga Toxin (Stx) Classification, Structure, and Function.” Microbiology spectrum 2.2 (2014): 10.1128/microbiolspec.EHEC–0024–2013. PMC. Web</ref>. The removal of the adenine prevents elongation factors from associating with the ribosomal subunit. Without elongation factors, the ribosome can no longer synthesize proteins, leading to cell death<ref name="Melton">Melton-Cesla, A. (2012). Shiga toxin classification structure and function. Changes, 29(2), 997–1003</ref>.
== Disease and It's Pathogenesis ==
The shiga toxin is produced by Shigella dysenteriae and certain strains of Escherichia coli. S. dysenteriae is a gram negative bacterium that causes disease in its host by entering into intestinal epithelial cells. It has a very low infectious dose of only ten to one hundred cells, and is contracted through the consumption of contaminated water or food. One it begins replicating in the intestines, its proliferation and secretion of stx erodes the intestinal lining and leads to dysentery<ref>Zaidi, Mussaret Bano, and Teresa Estrada-García. "Shigella: A Highly Virulent and Elusive pathogen." Current Tropical Medicine Reports 1.2 (2014): 81-87</ref>. Those with dysentery can simply have diarrhea as a symptom, but if the infection progresses, it can lead to hemorrhagic colitis (bloody stool) or hemolytic uremic syndrome (HUS). HUS is characterized as acute renal failure, thrombocytopenia, and microangiopathic anemia<ref>Kaper, James B. and Alison D. O’Brien. “Overview and Historical Perspectives.” Microbiology spectrum 2.2 (2014): 10.1128/microbiolspec.EHEC–0028–2014. PMC. Web</ref>. As stated previously, stx enters cells through its binding to GB3, but curiously many intestinal cells do not express GB3. Some figure it enters the cells through macropinocyosis, but when macropinocytotic function is removed from cells, stx still manages to enter through an undefined mechanism.
The genes necessary to produce the toxin are interestingly not native to S. dysenteriae. The genes for stx are carried by prophages: bacteria that have had viral DNA inserted into their genome. During certain conditions, the phage genes are expressed leading to the production of more bacteriophages as well as the shiga toxin. When the bacteria undergo lysis, the bacteriophages and stx are released. It may seem as if there is no benefit for S. dysenteriae to carry the viral DNA, but studies have shown that while it is in the lysogenic cycle S. dysenteriae has increased acid tolerance and motility<ref>Krüger, Alejandra, and Paula María Alejandra Lucchesi. "Shiga toxins and stx-phages: highly diverse entities." Microbiology (2014): mic-0</ref>.

This is a sample scene created with SAT to <scene name='68/687332/Scene_1/1'>TextToBeDisplayed</scene> by Group, and another to make <scene name="/12/3456/Sample/2">a transparent representation</scene> of the protein. You can make your own scenes on SAT starting from scratch or loading and editing one of these sample scenes.

</StructureSection>
== References ==
<references/>

Sandbox Reserved 982

2015-05-01T16:27:46Z

Joseph P. Hoffmann:

{{Telford_CHEM321}}
==Shiga Toxin==
<StructureSection load='1R4Q' size='340' side='right' caption='Crystal Structure for Shiga Toxin (1R4Q)' scene=''>

== Structural Highlights ==
Shiga toxin (stx) has multiple classifications. The shiga toxin produced by Shigella dysenteriae is classified as stx, while shiga toxin produced by Escherichia coli can be classified as either stx1 or stx2. All three classifications very slightly in structure but have identical functions. The A subunit of stx1 differs from stx by only one residue: a serine in position 45 as opposed to threonine<ref name="Fraser">Fraser, M. E., Fujinaga, M., Cherney, M. M., Melton-Celsa, A. R., Twiddy, E. M., O’Brien, A. D., & James, M. N. G. (2004). Structure of shiga toxin type 2 (Stx2) from Escherichia coli O157:H7. The Journal of Biological Chemistry, 279(26), 27511–27517. doi:10.1074/jbc.M401939200</ref>. Stx is a 70 kDa AB5 protein, meaning that it is composed of an A subunit bound to a B subunit pentamer. A noncovalent interaction causes the association between the A and B subunits, as the carboxy terminal tail of the A subunit is surrounded by the B pentamer. <scene name='68/687332/Sub-unit_a/2'>The A subunit</scene> is 293 amino acids long with its active site being glutamic acid 167. Though the active site is a single residue, studies have shown that the first 239 residues are essential for the enzymatic activity of the A subunit. Residues 240-251 are essential for the A subunit’s translocation from the endoplasmic reticulum of an infected cell to the cytosol<ref name="Sandvig">Sandvig, K., & van Deurs, B. (2000). Entry of ricin and Shiga toxin into cells: molecular mechanisms and medical perspectives. The EMBO Journal, 19(22), 5943–5950. doi:10.1093/emboj/19.22.5943</ref>. The A subunit can be broken down into A1 and A2 units. This break down occurs when a trypsin sensitive region, residues 248-251, is cleaved. This cleavage results in the only remaining link being a <scene name='68/687332/Disulfide_bridge/1'>disulfide bridge (1R4Q)</scene> between cysteine 242 and cysteine 261. If the disulfide bridge is reduced, the A1 and A2 subunits can completely separate. Interestingly, the disulfide bridge blocks the active site of the A subunit, so the A subunit is not enzymatically active unless it has been cleaved<ref name ="Fraser">Fraser, M. E., Fujinaga, M., Cherney, M. M., Melton-Celsa, A. R., Twiddy, E. M., O’Brien, A. D., & James, M. N. G. (2004). Structure of shiga toxin type 2 (Stx2) from Escherichia coli O157:H7. The Journal of Biological Chemistry, 279(26), 27511–27517. doi:10.1074/jbc.M401939200</ref>.
<scene name='68/687332/B_subunit/1'>The B subunit (2XSC)</scene> is a symmetrical pentamer that resembles a star and can be divided into five identical subunits. Each of the subunits is 69 amino acids in length. The purpose of the B subunit is to bind to globotriaosylceramide (GB3) which is a glycosphingolipid that is found on the lipid rafts of endothelial cells. Each monomer of the B subunit has three binding sites for GB3, binding to the <scene name='68/687332/Gb3_bound_to_subunit_b/1'> carbohydrate portion of GB3 (1CQF)</scene>. The affinity between the B subunit and GB3 is incredibly low, and is actually one of the lowest recorded affinities for carbohydrate-protein interaction<ref>Jacobson, J. M., Yin, J., Kitov, P. I., Mulvey, G., Griener, T. P., James, M. N. G., … Bundle, D. R. (2014). The crystal structure of Shiga toxin type 2 with bound disaccharide guides the design of a heterobifunctional toxin inhibitor. Journal of Biological Chemistry, 289(2), 885–894. doi:10.1074/jbc.M113.518886</ref>. Some studies have shown that the level of saturation and length of the fatty acid on GB3 affects the strength of its interaction with the B subunit. Since the B subunit of stx has fifteen binding sites though, it has an incredibly high avidity<ref name="Melton">Melton-Celsa, Angela R. “Shiga Toxin (Stx) Classification, Structure, and Function.” Microbiology spectrum 2.2 (2014): 10.1128/microbiolspec.EHEC–0024–2013. PMC. Web</ref>.
== Function ==
The pathway of stx entering a cell begins with the B subunit’s binding to GB3. Once this occurs, the A subunit disconnects from the B subunit and enters the cell through endocytosis. Using retrograde transport the A subunit passes through the Golgi apparatus and the rough endoplasmic reticulum<ref name="Melton">Melton-Celsa, Angela R. “Shiga Toxin (Stx) Classification, Structure, and Function.” Microbiology spectrum 2.2 (2014): 10.1128/microbiolspec.EHEC–0024–2013. PMC. Web</ref> In the rough endoplasmic reticulum, the A subunit is split into two parts called A1 and A2 through the cleavage of trypsin sensitive residues and the reduction of a disulfide bridge. A2 is degraded, but A1 freely enters the cytosol<ref name="Sandvig">Sandvig, K., & van Deurs, B. (2000). Entry of ricin and Shiga toxin into cells: molecular mechanisms and medical perspectives. The EMBO Journal, 19(22), 5943–5950. doi:10.1093/emboj/19.22.5943</ref>. Once in the cytosol, A1 acts as an N-glycosidase, which is an enzyme that hydrolyzes bonds that link sugars. With this enzymatic activity, A1 removes an adenine from the alpha-sarcin loop in the 28S RNA of the 60S ribosomal subunit<ref name="Melton">Melton-Celsa, Angela R. “Shiga Toxin (Stx) Classification, Structure, and Function.” Microbiology spectrum 2.2 (2014): 10.1128/microbiolspec.EHEC–0024–2013. PMC. Web</ref>. The removal of the adenine prevents elongation factors from associating with the ribosomal subunit. Without elongation factors, the ribosome can no longer synthesize proteins, leading to cell death<ref name="Melton">Melton-Cesla, A. (2012). Shiga toxin classification structure and function. Changes, 29(2), 997–1003</ref>.
== Disease and It's Pathogenesis ==
The shiga toxin is produced by Shigella dysenteriae and certain strains of Escherichia coli. S. dysenteriae is a gram negative bacterium that causes disease in its host by entering into intestinal epithelial cells. It has a very low infectious dose of only ten to one hundred cells, and is contracted through the consumption of contaminated water or food. One it begins replicating in the intestines, its proliferation and secretion of stx erodes the intestinal lining and leads to dysentery<ref>Zaidi, Mussaret Bano, and Teresa Estrada-García. "Shigella: A Highly Virulent and Elusive pathogen." Current Tropical Medicine Reports 1.2 (2014): 81-87</ref>. Those with dysentery can simply have diarrhea as a symptom, but if the infection progresses, it can lead to hemorrhagic colitis (bloody stool) or hemolytic uremic syndrome (HUS). HUS is characterized as acute renal failure, thrombocytopenia, and microangiopathic anemia<ref>Kaper, James B. and Alison D. O’Brien. “Overview and Historical Perspectives.” Microbiology spectrum 2.2 (2014): 10.1128/microbiolspec.EHEC–0028–2014. PMC. Web</ref>. As stated previously, stx enters cells through its binding to GB3, but curiously many intestinal cells do not express GB3. Some figure it enters the cells through macropinocyosis, but when macropinocytotic function is removed from cells, stx still manages to enter through an undefined mechanism.
The genes necessary to produce the toxin are interestingly not native to S. dysenteriae. The genes for stx are carried by prophages: bacteria that have had viral DNA inserted into their genome. During certain conditions, the phage genes are expressed leading to the production of more bacteriophages as well as the shiga toxin. When the bacteria undergo lysis, the bacteriophages and stx are released. It may seem as if there is no benefit for S. dysenteriae to carry the viral DNA, but studies have shown that while it is in the lysogenic cycle S. dysenteriae has increased acid tolerance and motility<ref>Krüger, Alejandra, and Paula María Alejandra Lucchesi. "Shiga toxins and stx-phages: highly diverse entities." Microbiology (2014): mic-0</ref>.

This is a sample scene created with SAT to <scene name='68/687332/Scene_1/1'>TextToBeDisplayed</scene> by Group, and another to make <scene name="/12/3456/Sample/2">a transparent representation</scene> of the protein. You can make your own scenes on SAT starting from scratch or loading and editing one of these sample scenes.

</StructureSection>
== References ==
<references/>

Sandbox Reserved 982

2015-05-01T16:07:39Z

Joseph P. Hoffmann:

{{Telford_CHEM321}}
==Shiga Toxin==
<StructureSection load='1R4Q' size='340' side='right' caption='Crystal Structure for Shiga Toxin (1R4Q)' scene=''>

== Structural Highlights ==
Shiga toxin (stx) has multiple classifications. The shiga toxin produced by Shigella dysenteriae is classified as stx, while shiga toxin produced by Escherichia coli can be classified as either stx1 or stx2. All three classifications very slightly in structure but have identical functions. The A subunit of stx1 differs from stx by only one residue: a serine in position 45 as opposed to threonine<ref name="Fraser">Fraser, M. E., Fujinaga, M., Cherney, M. M., Melton-Celsa, A. R., Twiddy, E. M., O’Brien, A. D., & James, M. N. G. (2004). Structure of shiga toxin type 2 (Stx2) from Escherichia coli O157:H7. The Journal of Biological Chemistry, 279(26), 27511–27517. doi:10.1074/jbc.M401939200</ref>. Stx is a 70 kDa AB5 protein, meaning that it is composed of an A subunit bound to a B subunit pentamer. A noncovalent interaction causes the association between the A and B subunits, as the carboxy terminal tail of the A subunit is surrounded by the B pentamer. <scene name='68/687332/Sub-unit_a/2'>The A subunit</scene> is 293 amino acids long with its active site being glutamic acid 167. Though the active site is a single residue, studies have shown that the first 239 residues are essential for the enzymatic activity of the A subunit. Residues 240-251 are essential for the A subunit’s translocation from the endoplasmic reticulum of an infected cell to the cytosol<ref name="Sandvig">Sandvig, K., & van Deurs, B. (2000). Entry of ricin and Shiga toxin into cells: molecular mechanisms and medical perspectives. The EMBO Journal, 19(22), 5943–5950. doi:10.1093/emboj/19.22.5943</ref>. The A subunit can be broken down into A1 and A2 units. This break down occurs when a trypsin sensitive region, residues 248-251, is cleaved. This cleavage results in the only remaining link being a disulfide bridge between cysteine 241 and cysteine 260. If the disulfide bridge is reduced, the A1 and A2 subunits can completely separate. Interestingly, the disulfide bridge blocks the active site of the A subunit, so the A subunit is not enzymatically active unless it has been cleaved<ref name ="Fraser">Fraser, M. E., Fujinaga, M., Cherney, M. M., Melton-Celsa, A. R., Twiddy, E. M., O’Brien, A. D., & James, M. N. G. (2004). Structure of shiga toxin type 2 (Stx2) from Escherichia coli O157:H7. The Journal of Biological Chemistry, 279(26), 27511–27517. doi:10.1074/jbc.M401939200</ref>.
<scene name='68/687332/B_subunit/1'>The B subunit (2XSC)</scene> is a symmetrical pentamer that resembles a star and can be divided into five identical subunits. Each of the subunits is 69 amino acids in length. The purpose of the B subunit is to bind to globotriaosylceramide (GB3) which is a glycosphingolipid that is found on the lipid rafts of endothelial cells. Each monomer of the B subunit has three binding sites for GB3, binding to the <scene name='68/687332/Gb3_bound_to_subunit_b/1'> carbohydrate portion of GB3 (1CQF)</scene>. The affinity between the B subunit and GB3 is incredibly low, and is actually one of the lowest recorded affinities for carbohydrate-protein interaction<ref>Jacobson, J. M., Yin, J., Kitov, P. I., Mulvey, G., Griener, T. P., James, M. N. G., … Bundle, D. R. (2014). The crystal structure of Shiga toxin type 2 with bound disaccharide guides the design of a heterobifunctional toxin inhibitor. Journal of Biological Chemistry, 289(2), 885–894. doi:10.1074/jbc.M113.518886</ref>. Some studies have shown that the level of saturation and length of the fatty acid on GB3 affects the strength of its interaction with the B subunit. Since the B subunit of stx has fifteen binding sites though, it has an incredibly high avidity<ref name="Melton">Melton-Celsa, Angela R. “Shiga Toxin (Stx) Classification, Structure, and Function.” Microbiology spectrum 2.2 (2014): 10.1128/microbiolspec.EHEC–0024–2013. PMC. Web</ref>.
== Function ==
The pathway of stx entering a cell begins with the B subunit’s binding to GB3. Once this occurs, the A subunit disconnects from the B subunit and enters the cell through endocytosis. Using retrograde transport the A subunit passes through the Golgi apparatus and the rough endoplasmic reticulum<ref name="Melton">Melton-Celsa, Angela R. “Shiga Toxin (Stx) Classification, Structure, and Function.” Microbiology spectrum 2.2 (2014): 10.1128/microbiolspec.EHEC–0024–2013. PMC. Web</ref> In the rough endoplasmic reticulum, the A subunit is split into two parts called A1 and A2 through the cleavage of trypsin sensitive residues and the reduction of a disulfide bridge. A2 is degraded, but A1 freely enters the cytosol<ref name="Sandvig">Sandvig, K., & van Deurs, B. (2000). Entry of ricin and Shiga toxin into cells: molecular mechanisms and medical perspectives. The EMBO Journal, 19(22), 5943–5950. doi:10.1093/emboj/19.22.5943</ref>. Once in the cytosol, A1 acts as an N-glycosidase, which is an enzyme that hydrolyzes bonds that link sugars. With this enzymatic activity, A1 removes an adenine from the alpha-sarcin loop in the 28S RNA of the 60S ribosomal subunit<ref name="Melton">Melton-Celsa, Angela R. “Shiga Toxin (Stx) Classification, Structure, and Function.” Microbiology spectrum 2.2 (2014): 10.1128/microbiolspec.EHEC–0024–2013. PMC. Web</ref>. The removal of the adenine prevents elongation factors from associating with the ribosomal subunit. Without elongation factors, the ribosome can no longer synthesize proteins, leading to cell death<ref name="Melton">Melton-Cesla, A. (2012). Shiga toxin classification structure and function. Changes, 29(2), 997–1003</ref>.
== Disease and It's Pathogenesis ==
The shiga toxin is produced by Shigella dysenteriae and certain strains of Escherichia coli. S. dysenteriae is a gram negative bacterium that causes disease in its host by entering into intestinal epithelial cells. It has a very low infectious dose of only ten to one hundred cells, and is contracted through the consumption of contaminated water or food. One it begins replicating in the intestines, its proliferation and secretion of stx erodes the intestinal lining and leads to dysentery<ref>Zaidi, Mussaret Bano, and Teresa Estrada-García. "Shigella: A Highly Virulent and Elusive pathogen." Current Tropical Medicine Reports 1.2 (2014): 81-87</ref>. Those with dysentery can simply have diarrhea as a symptom, but if the infection progresses, it can lead to hemorrhagic colitis (bloody stool) or hemolytic uremic syndrome (HUS). HUS is characterized as acute renal failure, thrombocytopenia, and microangiopathic anemia<ref>Kaper, James B. and Alison D. O’Brien. “Overview and Historical Perspectives.” Microbiology spectrum 2.2 (2014): 10.1128/microbiolspec.EHEC–0028–2014. PMC. Web</ref>. As stated previously, stx enters cells through its binding to GB3, but curiously many intestinal cells do not express GB3. Some figure it enters the cells through macropinocyosis, but when macropinocytotic function is removed from cells, stx still manages to enter through an undefined mechanism.
The genes necessary to produce the toxin are interestingly not native to S. dysenteriae. The genes for stx are carried by prophages: bacteria that have had viral DNA inserted into their genome. During certain conditions, the phage genes are expressed leading to the production of more bacteriophages as well as the shiga toxin. When the bacteria undergo lysis, the bacteriophages and stx are released. It may seem as if there is no benefit for S. dysenteriae to carry the viral DNA, but studies have shown that while it is in the lysogenic cycle S. dysenteriae has increased acid tolerance and motility<ref>Krüger, Alejandra, and Paula María Alejandra Lucchesi. "Shiga toxins and stx-phages: highly diverse entities." Microbiology (2014): mic-0</ref>.

This is a sample scene created with SAT to <scene name='68/687332/Scene_1/1'>TextToBeDisplayed</scene> by Group, and another to make <scene name="/12/3456/Sample/2">a transparent representation</scene> of the protein. You can make your own scenes on SAT starting from scratch or loading and editing one of these sample scenes.

</StructureSection>
== References ==
<references/>

Sandbox Reserved 982

2015-05-01T15:52:33Z

Joseph P. Hoffmann:

{{Telford_CHEM321}}
==Shiga Toxin==
<StructureSection load='1R4Q' size='340' side='right' caption='Crystal Structure for Shiga Toxin (1R4Q)' scene=''>

== Structural Highlights ==
Shiga toxin (stx) has multiple classifications. The shiga toxin produced by Shigella dysenteriae is classified as stx, while shiga toxin produced by Escherichia coli can be classified as either stx1 or stx2. All three classifications very slightly in structure but have identical functions. The A subunit of stx1 differs from stx by only one residue: a serine in position 45 as opposed to threonine<ref name="Fraser">Fraser, M. E., Fujinaga, M., Cherney, M. M., Melton-Celsa, A. R., Twiddy, E. M., O’Brien, A. D., & James, M. N. G. (2004). Structure of shiga toxin type 2 (Stx2) from Escherichia coli O157:H7. The Journal of Biological Chemistry, 279(26), 27511–27517. doi:10.1074/jbc.M401939200</ref>. Stx is a 70 kDa AB5 protein, meaning that it is composed of an A subunit bound to a B subunit pentamer. A noncovalent interaction causes the association between the A and B subunits, as the carboxy terminal tail of the A subunit is surrounded by the B pentamer. The A subunit is 293 amino acids long with its active site being glutamic acid 167. Though the active site is a single residue, studies have shown that the first 239 residues are essential for the enzymatic activity of the A subunit. Residues 240-251 are essential for the A subunit’s translocation from the endoplasmic reticulum of an infected cell to the cytosol<ref name="Sandvig">Sandvig, K., & van Deurs, B. (2000). Entry of ricin and Shiga toxin into cells: molecular mechanisms and medical perspectives. The EMBO Journal, 19(22), 5943–5950. doi:10.1093/emboj/19.22.5943</ref>. The A subunit can be broken down into A1 and A2 units. This break down occurs when a trypsin sensitive region, residues 248-251, is cleaved. This cleavage results in the only remaining link being a disulfide bridge between cysteine 241 and cysteine 260. If the disulfide bridge is reduced, the A1 and A2 subunits can completely separate. Interestingly, the disulfide bridge blocks the active site of the A subunit, so the A subunit is not enzymatically active unless it has been cleaved<ref name ="Fraser">Fraser, M. E., Fujinaga, M., Cherney, M. M., Melton-Celsa, A. R., Twiddy, E. M., O’Brien, A. D., & James, M. N. G. (2004). Structure of shiga toxin type 2 (Stx2) from Escherichia coli O157:H7. The Journal of Biological Chemistry, 279(26), 27511–27517. doi:10.1074/jbc.M401939200</ref>.
<scene name='68/687332/B_subunit/1'>The B subunit (2XSC)</scene> is a symmetrical pentamer that resembles a star and can be divided into five identical subunits. Each of the subunits is 69 amino acids in length. The purpose of the B subunit is to bind to globotriaosylceramide (GB3) which is a glycosphingolipid that is found on the lipid rafts of endothelial cells. Each monomer of the B subunit has three binding sites for GB3, binding to the <scene name='68/687332/Gb3_bound_to_subunit_b/1'> carbohydrate portion of GB3 (1CQF)</scene>. The affinity between the B subunit and GB3 is incredibly low, and is actually one of the lowest recorded affinities for carbohydrate-protein interaction<ref>Jacobson, J. M., Yin, J., Kitov, P. I., Mulvey, G., Griener, T. P., James, M. N. G., … Bundle, D. R. (2014). The crystal structure of Shiga toxin type 2 with bound disaccharide guides the design of a heterobifunctional toxin inhibitor. Journal of Biological Chemistry, 289(2), 885–894. doi:10.1074/jbc.M113.518886</ref>. Some studies have shown that the level of saturation and length of the fatty acid on GB3 affects the strength of its interaction with the B subunit. Since the B subunit of stx has fifteen binding sites though, it has an incredibly high avidity<ref name="Melton">Melton-Celsa, Angela R. “Shiga Toxin (Stx) Classification, Structure, and Function.” Microbiology spectrum 2.2 (2014): 10.1128/microbiolspec.EHEC–0024–2013. PMC. Web</ref>.
== Function ==
The pathway of stx entering a cell begins with the B subunit’s binding to GB3. Once this occurs, the A subunit disconnects from the B subunit and enters the cell through endocytosis. Using retrograde transport the A subunit passes through the Golgi apparatus and the rough endoplasmic reticulum<ref name="Melton">Melton-Celsa, Angela R. “Shiga Toxin (Stx) Classification, Structure, and Function.” Microbiology spectrum 2.2 (2014): 10.1128/microbiolspec.EHEC–0024–2013. PMC. Web</ref> In the rough endoplasmic reticulum, the A subunit is split into two parts called A1 and A2 through the cleavage of trypsin sensitive residues and the reduction of a disulfide bridge. A2 is degraded, but A1 freely enters the cytosol<ref name="Sandvig">Sandvig, K., & van Deurs, B. (2000). Entry of ricin and Shiga toxin into cells: molecular mechanisms and medical perspectives. The EMBO Journal, 19(22), 5943–5950. doi:10.1093/emboj/19.22.5943</ref>. Once in the cytosol, A1 acts as an N-glycosidase, which is an enzyme that hydrolyzes bonds that link sugars. With this enzymatic activity, A1 removes an adenine from the alpha-sarcin loop in the 28S RNA of the 60S ribosomal subunit<ref name="Melton">Melton-Celsa, Angela R. “Shiga Toxin (Stx) Classification, Structure, and Function.” Microbiology spectrum 2.2 (2014): 10.1128/microbiolspec.EHEC–0024–2013. PMC. Web</ref>. The removal of the adenine prevents elongation factors from associating with the ribosomal subunit. Without elongation factors, the ribosome can no longer synthesize proteins, leading to cell death<ref name="Melton">Melton-Cesla, A. (2012). Shiga toxin classification structure and function. Changes, 29(2), 997–1003</ref>.
== Disease and It's Pathogenesis ==
The shiga toxin is produced by Shigella dysenteriae and certain strains of Escherichia coli. S. dysenteriae is a gram negative bacterium that causes disease in its host by entering into intestinal epithelial cells. It has a very low infectious dose of only ten to one hundred cells, and is contracted through the consumption of contaminated water or food. One it begins replicating in the intestines, its proliferation and secretion of stx erodes the intestinal lining and leads to dysentery<ref>Zaidi, Mussaret Bano, and Teresa Estrada-García. "Shigella: A Highly Virulent and Elusive pathogen." Current Tropical Medicine Reports 1.2 (2014): 81-87</ref>. Those with dysentery can simply have diarrhea as a symptom, but if the infection progresses, it can lead to hemorrhagic colitis (bloody stool) or hemolytic uremic syndrome (HUS). HUS is characterized as acute renal failure, thrombocytopenia, and microangiopathic anemia<ref>Kaper, James B. and Alison D. O’Brien. “Overview and Historical Perspectives.” Microbiology spectrum 2.2 (2014): 10.1128/microbiolspec.EHEC–0028–2014. PMC. Web</ref>. As stated previously, stx enters cells through its binding to GB3, but curiously many intestinal cells do not express GB3. Some figure it enters the cells through macropinocyosis, but when macropinocytotic function is removed from cells, stx still manages to enter through an undefined mechanism.
The genes necessary to produce the toxin are interestingly not native to S. dysenteriae. The genes for stx are carried by prophages: bacteria that have had viral DNA inserted into their genome. During certain conditions, the phage genes are expressed leading to the production of more bacteriophages as well as the shiga toxin. When the bacteria undergo lysis, the bacteriophages and stx are released. It may seem as if there is no benefit for S. dysenteriae to carry the viral DNA, but studies have shown that while it is in the lysogenic cycle S. dysenteriae has increased acid tolerance and motility<ref>Krüger, Alejandra, and Paula María Alejandra Lucchesi. "Shiga toxins and stx-phages: highly diverse entities." Microbiology (2014): mic-0</ref>.

This is a sample scene created with SAT to <scene name='68/687332/Scene_1/1'>TextToBeDisplayed</scene> by Group, and another to make <scene name="/12/3456/Sample/2">a transparent representation</scene> of the protein. You can make your own scenes on SAT starting from scratch or loading and editing one of these sample scenes.

</StructureSection>
== References ==
<references/>

Sandbox Reserved 982

2015-05-01T15:51:19Z

Joseph P. Hoffmann:

{{Telford_CHEM321}}
==Shiga Toxin==
<StructureSection load='1R4Q' size='340' side='right' caption='Crystal Structure for Shiga Toxin (1R4Q)' scene=''>

== Structural Highlights ==
Shiga toxin (stx) has multiple classifications. The shiga toxin produced by Shigella dysenteriae is classified as stx, while shiga toxin produced by Escherichia coli can be classified as either stx1 or stx2. All three classifications very slightly in structure but have identical functions. The A subunit of stx1 differs from stx by only one residue: a serine in position 45 as opposed to threonine<ref name="Fraser">Fraser, M. E., Fujinaga, M., Cherney, M. M., Melton-Celsa, A. R., Twiddy, E. M., O’Brien, A. D., & James, M. N. G. (2004). Structure of shiga toxin type 2 (Stx2) from Escherichia coli O157:H7. The Journal of Biological Chemistry, 279(26), 27511–27517. doi:10.1074/jbc.M401939200</ref>. Stx is a 70 kDa AB5 protein, meaning that it is composed of an A subunit bound to a B subunit pentamer. A noncovalent interaction causes the association between the A and B subunits, as the carboxy terminal tail of the A subunit is surrounded by the B pentamer. The A subunit is 293 amino acids long with its active site being glutamic acid 167. Though the active site is a single residue, studies have shown that the first 239 residues are essential for the enzymatic activity of the A subunit. Residues 240-251 are essential for the A subunit’s translocation from the endoplasmic reticulum of an infected cell to the cytosol<ref name="Sandvig">Sandvig, K., & van Deurs, B. (2000). Entry of ricin and Shiga toxin into cells: molecular mechanisms and medical perspectives. The EMBO Journal, 19(22), 5943–5950. doi:10.1093/emboj/19.22.5943</ref>. The A subunit can be broken down into A1 and A2 units. This break down occurs when a trypsin sensitive region, residues 248-251, is cleaved. This cleavage results in the only remaining link being a disulfide bridge between cysteine 241 and cysteine 260. If the disulfide bridge is reduced, the A1 and A2 subunits can completely separate. Interestingly, the disulfide bridge blocks the active site of the A subunit, so the A subunit is not enzymatically active unless it has been cleaved<ref name ="Fraser">Fraser, M. E., Fujinaga, M., Cherney, M. M., Melton-Celsa, A. R., Twiddy, E. M., O’Brien, A. D., & James, M. N. G. (2004). Structure of shiga toxin type 2 (Stx2) from Escherichia coli O157:H7. The Journal of Biological Chemistry, 279(26), 27511–27517. doi:10.1074/jbc.M401939200</ref>.
<scene name='68/687332/B_subunit/1'>The B subunit</scene> is a symmetrical pentamer that resembles a star and can be divided into five identical subunits. Each of the subunits is 69 amino acids in length. The purpose of the B subunit is to bind to globotriaosylceramide (GB3) which is a glycosphingolipid that is found on the lipid rafts of endothelial cells. Each monomer of the B subunit has three binding sites for GB3, binding to the <scene name='68/687332/Gb3_bound_to_subunit_b/1'> carbohydrate portion of GB3 (1CQF)</scene>. The affinity between the B subunit and GB3 is incredibly low, and is actually one of the lowest recorded affinities for carbohydrate-protein interaction<ref>Jacobson, J. M., Yin, J., Kitov, P. I., Mulvey, G., Griener, T. P., James, M. N. G., … Bundle, D. R. (2014). The crystal structure of Shiga toxin type 2 with bound disaccharide guides the design of a heterobifunctional toxin inhibitor. Journal of Biological Chemistry, 289(2), 885–894. doi:10.1074/jbc.M113.518886</ref>. Some studies have shown that the level of saturation and length of the fatty acid on GB3 affects the strength of its interaction with the B subunit. Since the B subunit of stx has fifteen binding sites though, it has an incredibly high avidity<ref name="Melton">Melton-Celsa, Angela R. “Shiga Toxin (Stx) Classification, Structure, and Function.” Microbiology spectrum 2.2 (2014): 10.1128/microbiolspec.EHEC–0024–2013. PMC. Web</ref>.
== Function ==
The pathway of stx entering a cell begins with the B subunit’s binding to GB3. Once this occurs, the A subunit disconnects from the B subunit and enters the cell through endocytosis. Using retrograde transport the A subunit passes through the Golgi apparatus and the rough endoplasmic reticulum<ref name="Melton">Melton-Celsa, Angela R. “Shiga Toxin (Stx) Classification, Structure, and Function.” Microbiology spectrum 2.2 (2014): 10.1128/microbiolspec.EHEC–0024–2013. PMC. Web</ref> In the rough endoplasmic reticulum, the A subunit is split into two parts called A1 and A2 through the cleavage of trypsin sensitive residues and the reduction of a disulfide bridge. A2 is degraded, but A1 freely enters the cytosol<ref name="Sandvig">Sandvig, K., & van Deurs, B. (2000). Entry of ricin and Shiga toxin into cells: molecular mechanisms and medical perspectives. The EMBO Journal, 19(22), 5943–5950. doi:10.1093/emboj/19.22.5943</ref>. Once in the cytosol, A1 acts as an N-glycosidase, which is an enzyme that hydrolyzes bonds that link sugars. With this enzymatic activity, A1 removes an adenine from the alpha-sarcin loop in the 28S RNA of the 60S ribosomal subunit<ref name="Melton">Melton-Celsa, Angela R. “Shiga Toxin (Stx) Classification, Structure, and Function.” Microbiology spectrum 2.2 (2014): 10.1128/microbiolspec.EHEC–0024–2013. PMC. Web</ref>. The removal of the adenine prevents elongation factors from associating with the ribosomal subunit. Without elongation factors, the ribosome can no longer synthesize proteins, leading to cell death<ref name="Melton">Melton-Cesla, A. (2012). Shiga toxin classification structure and function. Changes, 29(2), 997–1003</ref>.
== Disease and It's Pathogenesis ==
The shiga toxin is produced by Shigella dysenteriae and certain strains of Escherichia coli. S. dysenteriae is a gram negative bacterium that causes disease in its host by entering into intestinal epithelial cells. It has a very low infectious dose of only ten to one hundred cells, and is contracted through the consumption of contaminated water or food. One it begins replicating in the intestines, its proliferation and secretion of stx erodes the intestinal lining and leads to dysentery<ref>Zaidi, Mussaret Bano, and Teresa Estrada-García. "Shigella: A Highly Virulent and Elusive pathogen." Current Tropical Medicine Reports 1.2 (2014): 81-87</ref>. Those with dysentery can simply have diarrhea as a symptom, but if the infection progresses, it can lead to hemorrhagic colitis (bloody stool) or hemolytic uremic syndrome (HUS). HUS is characterized as acute renal failure, thrombocytopenia, and microangiopathic anemia<ref>Kaper, James B. and Alison D. O’Brien. “Overview and Historical Perspectives.” Microbiology spectrum 2.2 (2014): 10.1128/microbiolspec.EHEC–0028–2014. PMC. Web</ref>. As stated previously, stx enters cells through its binding to GB3, but curiously many intestinal cells do not express GB3. Some figure it enters the cells through macropinocyosis, but when macropinocytotic function is removed from cells, stx still manages to enter through an undefined mechanism.
The genes necessary to produce the toxin are interestingly not native to S. dysenteriae. The genes for stx are carried by prophages: bacteria that have had viral DNA inserted into their genome. During certain conditions, the phage genes are expressed leading to the production of more bacteriophages as well as the shiga toxin. When the bacteria undergo lysis, the bacteriophages and stx are released. It may seem as if there is no benefit for S. dysenteriae to carry the viral DNA, but studies have shown that while it is in the lysogenic cycle S. dysenteriae has increased acid tolerance and motility<ref>Krüger, Alejandra, and Paula María Alejandra Lucchesi. "Shiga toxins and stx-phages: highly diverse entities." Microbiology (2014): mic-0</ref>.

This is a sample scene created with SAT to <scene name='68/687332/Scene_1/1'>TextToBeDisplayed</scene> by Group, and another to make <scene name="/12/3456/Sample/2">a transparent representation</scene> of the protein. You can make your own scenes on SAT starting from scratch or loading and editing one of these sample scenes.

</StructureSection>
== References ==
<references/>

Sandbox Reserved 982

2015-05-01T15:42:18Z

Joseph P. Hoffmann:

{{Telford_CHEM321}}
==Shiga Toxin==
<StructureSection load='1R4Q' size='340' side='right' caption='Crystal Structure for Shiga Toxin (1R4Q)' scene=''>

== Structural Highlights ==
Shiga toxin (stx) has multiple classifications. The shiga toxin produced by Shigella dysenteriae is classified as stx, while shiga toxin produced by Escherichia coli can be classified as either stx1 or stx2. All three classifications very slightly in structure but have identical functions. The A subunit of stx1 differs from stx by only one residue: a serine in position 45 as opposed to threonine<ref name="Fraser">Fraser, M. E., Fujinaga, M., Cherney, M. M., Melton-Celsa, A. R., Twiddy, E. M., O’Brien, A. D., & James, M. N. G. (2004). Structure of shiga toxin type 2 (Stx2) from Escherichia coli O157:H7. The Journal of Biological Chemistry, 279(26), 27511–27517. doi:10.1074/jbc.M401939200</ref>. Stx is a 70 kDa AB5 protein, meaning that it is composed of an A subunit bound to a B subunit pentamer. A noncovalent interaction causes the association between the A and B subunits, as the carboxy terminal tail of the A subunit is surrounded by the B pentamer. The A subunit is 293 amino acids long with its active site being glutamic acid 167. Though the active site is a single residue, studies have shown that the first 239 residues are essential for the enzymatic activity of the A subunit. Residues 240-251 are essential for the A subunit’s translocation from the endoplasmic reticulum of an infected cell to the cytosol<ref name="Sandvig">Sandvig, K., & van Deurs, B. (2000). Entry of ricin and Shiga toxin into cells: molecular mechanisms and medical perspectives. The EMBO Journal, 19(22), 5943–5950. doi:10.1093/emboj/19.22.5943</ref>. The A subunit can be broken down into A1 and A2 units. This break down occurs when a trypsin sensitive region, residues 248-251, is cleaved. This cleavage results in the only remaining link being a disulfide bridge between cysteine 241 and cysteine 260. If the disulfide bridge is reduced, the A1 and A2 subunits can completely separate. Interestingly, the disulfide bridge blocks the active site of the A subunit, so the A subunit is not enzymatically active unless it has been cleaved<ref name ="Fraser">Fraser, M. E., Fujinaga, M., Cherney, M. M., Melton-Celsa, A. R., Twiddy, E. M., O’Brien, A. D., & James, M. N. G. (2004). Structure of shiga toxin type 2 (Stx2) from Escherichia coli O157:H7. The Journal of Biological Chemistry, 279(26), 27511–27517. doi:10.1074/jbc.M401939200</ref>.
The B subunit is a symmetrical pentamer that resembles a star and can be divided into five identical subunits. Each of the subunits is 69 amino acids in length. The purpose of the B subunit is to bind to globotriaosylceramide (GB3) which is a glycosphingolipid that is found on the lipid rafts of endothelial cells. Each monomer of the B subunit has three binding sites for GB3, binding to the <scene name='68/687332/Gb3_bound_to_subunit_b/1'> carbohydrate portion of GB3 (1CQF)</scene>. The affinity between the B subunit and GB3 is incredibly low, and is actually one of the lowest recorded affinities for carbohydrate-protein interaction<ref>Jacobson, J. M., Yin, J., Kitov, P. I., Mulvey, G., Griener, T. P., James, M. N. G., … Bundle, D. R. (2014). The crystal structure of Shiga toxin type 2 with bound disaccharide guides the design of a heterobifunctional toxin inhibitor. Journal of Biological Chemistry, 289(2), 885–894. doi:10.1074/jbc.M113.518886</ref>. Some studies have shown that the level of saturation and length of the fatty acid on GB3 affects the strength of its interaction with the B subunit. Since the B subunit of stx has fifteen binding sites though, it has an incredibly high avidity<ref name="Melton">Melton-Celsa, Angela R. “Shiga Toxin (Stx) Classification, Structure, and Function.” Microbiology spectrum 2.2 (2014): 10.1128/microbiolspec.EHEC–0024–2013. PMC. Web</ref>.
== Function ==
The pathway of stx entering a cell begins with the B subunit’s binding to GB3. Once this occurs, the A subunit disconnects from the B subunit and enters the cell through endocytosis. Using retrograde transport the A subunit passes through the Golgi apparatus and the rough endoplasmic reticulum<ref name="Melton">Melton-Celsa, Angela R. “Shiga Toxin (Stx) Classification, Structure, and Function.” Microbiology spectrum 2.2 (2014): 10.1128/microbiolspec.EHEC–0024–2013. PMC. Web</ref> In the rough endoplasmic reticulum, the A subunit is split into two parts called A1 and A2 through the cleavage of trypsin sensitive residues and the reduction of a disulfide bridge. A2 is degraded, but A1 freely enters the cytosol<ref name="Sandvig">Sandvig, K., & van Deurs, B. (2000). Entry of ricin and Shiga toxin into cells: molecular mechanisms and medical perspectives. The EMBO Journal, 19(22), 5943–5950. doi:10.1093/emboj/19.22.5943</ref>. Once in the cytosol, A1 acts as an N-glycosidase, which is an enzyme that hydrolyzes bonds that link sugars. With this enzymatic activity, A1 removes an adenine from the alpha-sarcin loop in the 28S RNA of the 60S ribosomal subunit<ref name="Melton">Melton-Celsa, Angela R. “Shiga Toxin (Stx) Classification, Structure, and Function.” Microbiology spectrum 2.2 (2014): 10.1128/microbiolspec.EHEC–0024–2013. PMC. Web</ref>. The removal of the adenine prevents elongation factors from associating with the ribosomal subunit. Without elongation factors, the ribosome can no longer synthesize proteins, leading to cell death<ref name="Melton">Melton-Cesla, A. (2012). Shiga toxin classification structure and function. Changes, 29(2), 997–1003</ref>.
== Disease and It's Pathogenesis ==
The shiga toxin is produced by Shigella dysenteriae and certain strains of Escherichia coli. S. dysenteriae is a gram negative bacterium that causes disease in its host by entering into intestinal epithelial cells. It has a very low infectious dose of only ten to one hundred cells, and is contracted through the consumption of contaminated water or food. One it begins replicating in the intestines, its proliferation and secretion of stx erodes the intestinal lining and leads to dysentery<ref>Zaidi, Mussaret Bano, and Teresa Estrada-García. "Shigella: A Highly Virulent and Elusive pathogen." Current Tropical Medicine Reports 1.2 (2014): 81-87</ref>. Those with dysentery can simply have diarrhea as a symptom, but if the infection progresses, it can lead to hemorrhagic colitis (bloody stool) or hemolytic uremic syndrome (HUS). HUS is characterized as acute renal failure, thrombocytopenia, and microangiopathic anemia<ref>Kaper, James B. and Alison D. O’Brien. “Overview and Historical Perspectives.” Microbiology spectrum 2.2 (2014): 10.1128/microbiolspec.EHEC–0028–2014. PMC. Web</ref>. As stated previously, stx enters cells through its binding to GB3, but curiously many intestinal cells do not express GB3. Some figure it enters the cells through macropinocyosis, but when macropinocytotic function is removed from cells, stx still manages to enter through an undefined mechanism.
The genes necessary to produce the toxin are interestingly not native to S. dysenteriae. The genes for stx are carried by prophages: bacteria that have had viral DNA inserted into their genome. During certain conditions, the phage genes are expressed leading to the production of more bacteriophages as well as the shiga toxin. When the bacteria undergo lysis, the bacteriophages and stx are released. It may seem as if there is no benefit for S. dysenteriae to carry the viral DNA, but studies have shown that while it is in the lysogenic cycle S. dysenteriae has increased acid tolerance and motility<ref>Krüger, Alejandra, and Paula María Alejandra Lucchesi. "Shiga toxins and stx-phages: highly diverse entities." Microbiology (2014): mic-0</ref>.

This is a sample scene created with SAT to <scene name='68/687332/Scene_1/1'>TextToBeDisplayed</scene> by Group, and another to make <scene name="/12/3456/Sample/2">a transparent representation</scene> of the protein. You can make your own scenes on SAT starting from scratch or loading and editing one of these sample scenes.

</StructureSection>
== References ==
<references/>

Sandbox Reserved 982

2015-05-01T15:40:04Z

Joseph P. Hoffmann:

{{Telford_CHEM321}}
==Shiga Toxin==
<StructureSection load='1R4Q' size='340' side='right' caption='Crystal Structure for Shiga Toxin (1R4Q)' scene=''>

== Structural Highlights ==
Shiga toxin (stx) has multiple classifications. The shiga toxin produced by Shigella dysenteriae is classified as stx, while shiga toxin produced by Escherichia coli can be classified as either stx1 or stx2. All three classifications very slightly in structure but have identical functions. The A subunit of stx1 differs from stx by only one residue: a serine in position 45 as opposed to threonine<ref name="Fraser">Fraser, M. E., Fujinaga, M., Cherney, M. M., Melton-Celsa, A. R., Twiddy, E. M., O’Brien, A. D., & James, M. N. G. (2004). Structure of shiga toxin type 2 (Stx2) from Escherichia coli O157:H7. The Journal of Biological Chemistry, 279(26), 27511–27517. doi:10.1074/jbc.M401939200</ref>. Stx is a 70 kDa AB5 protein, meaning that it is composed of an A subunit bound to a B subunit pentamer. A noncovalent interaction causes the association between the A and B subunits, as the carboxy terminal tail of the A subunit is surrounded by the B pentamer. The A subunit is 293 amino acids long with its active site being glutamic acid 167. Though the active site is a single residue, studies have shown that the first 239 residues are essential for the enzymatic activity of the A subunit. Residues 240-251 are essential for the A subunit’s translocation from the endoplasmic reticulum of an infected cell to the cytosol<ref name="Sandvig">Sandvig, K., & van Deurs, B. (2000). Entry of ricin and Shiga toxin into cells: molecular mechanisms and medical perspectives. The EMBO Journal, 19(22), 5943–5950. doi:10.1093/emboj/19.22.5943</ref>. The A subunit can be broken down into A1 and A2 units. This break down occurs when a trypsin sensitive region, residues 248-251, is cleaved. This cleavage results in the only remaining link being a disulfide bridge between cysteine 241 and cysteine 260. If the disulfide bridge is reduced, the A1 and A2 subunits can completely separate. Interestingly, the disulfide bridge blocks the active site of the A subunit, so the A subunit is not enzymatically active unless it has been cleaved<ref name ="Fraser">Fraser, M. E., Fujinaga, M., Cherney, M. M., Melton-Celsa, A. R., Twiddy, E. M., O’Brien, A. D., & James, M. N. G. (2004). Structure of shiga toxin type 2 (Stx2) from Escherichia coli O157:H7. The Journal of Biological Chemistry, 279(26), 27511–27517. doi:10.1074/jbc.M401939200</ref>.
The B subunit is a symmetrical pentamer that resembles a star and can be divided into five identical subunits. Each of the subunits is 69 amino acids in length. The purpose of the B subunit is to bind to globotriaosylceramide (GB3) which is a glycosphingolipid that is found on the lipid rafts of endothelial cells. Each monomer of the B subunit has three binding sites for GB3, binding to the <scene name='68/687332/Gb3_bound_to_subunit_b/1'> carbohydrate portion of GB3</scene>. The affinity between the B subunit and GB3 is incredibly low, and is actually one of the lowest recorded affinities for carbohydrate-protein interaction<ref>Jacobson, J. M., Yin, J., Kitov, P. I., Mulvey, G., Griener, T. P., James, M. N. G., … Bundle, D. R. (2014). The crystal structure of Shiga toxin type 2 with bound disaccharide guides the design of a heterobifunctional toxin inhibitor. Journal of Biological Chemistry, 289(2), 885–894. doi:10.1074/jbc.M113.518886</ref>. Some studies have shown that the level of saturation and length of the fatty acid on GB3 affects the strength of its interaction with the B subunit. Since the B subunit of stx has fifteen binding sites though, it has an incredibly high avidity<ref name="Melton">Melton-Celsa, Angela R. “Shiga Toxin (Stx) Classification, Structure, and Function.” Microbiology spectrum 2.2 (2014): 10.1128/microbiolspec.EHEC–0024–2013. PMC. Web</ref>.
== Function ==
The pathway of stx entering a cell begins with the B subunit’s binding to GB3. Once this occurs, the A subunit disconnects from the B subunit and enters the cell through endocytosis. Using retrograde transport the A subunit passes through the Golgi apparatus and the rough endoplasmic reticulum<ref name="Melton">Melton-Celsa, Angela R. “Shiga Toxin (Stx) Classification, Structure, and Function.” Microbiology spectrum 2.2 (2014): 10.1128/microbiolspec.EHEC–0024–2013. PMC. Web</ref> In the rough endoplasmic reticulum, the A subunit is split into two parts called A1 and A2 through the cleavage of trypsin sensitive residues and the reduction of a disulfide bridge. A2 is degraded, but A1 freely enters the cytosol<ref name="Sandvig">Sandvig, K., & van Deurs, B. (2000). Entry of ricin and Shiga toxin into cells: molecular mechanisms and medical perspectives. The EMBO Journal, 19(22), 5943–5950. doi:10.1093/emboj/19.22.5943</ref>. Once in the cytosol, A1 acts as an N-glycosidase, which is an enzyme that hydrolyzes bonds that link sugars. With this enzymatic activity, A1 removes an adenine from the alpha-sarcin loop in the 28S RNA of the 60S ribosomal subunit<ref name="Melton">Melton-Celsa, Angela R. “Shiga Toxin (Stx) Classification, Structure, and Function.” Microbiology spectrum 2.2 (2014): 10.1128/microbiolspec.EHEC–0024–2013. PMC. Web</ref>. The removal of the adenine prevents elongation factors from associating with the ribosomal subunit. Without elongation factors, the ribosome can no longer synthesize proteins, leading to cell death<ref name="Melton">Melton-Cesla, A. (2012). Shiga toxin classification structure and function. Changes, 29(2), 997–1003</ref>.
== Disease and It's Pathogenesis ==
The shiga toxin is produced by Shigella dysenteriae and certain strains of Escherichia coli. S. dysenteriae is a gram negative bacterium that causes disease in its host by entering into intestinal epithelial cells. It has a very low infectious dose of only ten to one hundred cells, and is contracted through the consumption of contaminated water or food. One it begins replicating in the intestines, its proliferation and secretion of stx erodes the intestinal lining and leads to dysentery<ref>Zaidi, Mussaret Bano, and Teresa Estrada-García. "Shigella: A Highly Virulent and Elusive pathogen." Current Tropical Medicine Reports 1.2 (2014): 81-87</ref>. Those with dysentery can simply have diarrhea as a symptom, but if the infection progresses, it can lead to hemorrhagic colitis (bloody stool) or hemolytic uremic syndrome (HUS). HUS is characterized as acute renal failure, thrombocytopenia, and microangiopathic anemia<ref>Kaper, James B. and Alison D. O’Brien. “Overview and Historical Perspectives.” Microbiology spectrum 2.2 (2014): 10.1128/microbiolspec.EHEC–0028–2014. PMC. Web</ref>. As stated previously, stx enters cells through its binding to GB3, but curiously many intestinal cells do not express GB3. Some figure it enters the cells through macropinocyosis, but when macropinocytotic function is removed from cells, stx still manages to enter through an undefined mechanism.
The genes necessary to produce the toxin are interestingly not native to S. dysenteriae. The genes for stx are carried by prophages: bacteria that have had viral DNA inserted into their genome. During certain conditions, the phage genes are expressed leading to the production of more bacteriophages as well as the shiga toxin. When the bacteria undergo lysis, the bacteriophages and stx are released. It may seem as if there is no benefit for S. dysenteriae to carry the viral DNA, but studies have shown that while it is in the lysogenic cycle S. dysenteriae has increased acid tolerance and motility<ref>Krüger, Alejandra, and Paula María Alejandra Lucchesi. "Shiga toxins and stx-phages: highly diverse entities." Microbiology (2014): mic-0</ref>.

This is a sample scene created with SAT to <scene name='68/687332/Scene_1/1'>TextToBeDisplayed</scene> by Group, and another to make <scene name="/12/3456/Sample/2">a transparent representation</scene> of the protein. You can make your own scenes on SAT starting from scratch or loading and editing one of these sample scenes.

</StructureSection>
== References ==
<references/>

Sandbox Reserved 982

2015-05-01T06:08:14Z

Joseph P. Hoffmann:

{{Telford_CHEM321}}
==Shiga Toxin==
<StructureSection load='1R4Q' size='340' side='right' caption='Crystal Structure for Shiga Toxin (1R4Q)' scene=''>

== Structural Highlights ==
Shiga toxin (stx) has multiple classifications. The shiga toxin produced by Shigella dysenteriae is classified as stx, while shiga toxin produced by Escherichia coli can be classified as either stx1 or stx2. All three classifications very slightly in structure but have identical functions. The A subunit of stx1 differs from stx by only one residue: a serine in position 45 as opposed to threonine<ref name="Fraser">Fraser, M. E., Fujinaga, M., Cherney, M. M., Melton-Celsa, A. R., Twiddy, E. M., O’Brien, A. D., & James, M. N. G. (2004). Structure of shiga toxin type 2 (Stx2) from Escherichia coli O157:H7. The Journal of Biological Chemistry, 279(26), 27511–27517. doi:10.1074/jbc.M401939200</ref>. Stx is a 70 kDa AB5 protein, meaning that it is composed of an A subunit bound to a B subunit pentamer. A noncovalent interaction causes the association between the A and B subunits, as the carboxy terminal tail of the A subunit is surrounded by the B pentamer. The A subunit is 293 amino acids long with its active site being glutamic acid 167. Though the active site is a single residue, studies have shown that the first 239 residues are essential for the enzymatic activity of the A subunit. Residues 240-251 are essential for the A subunit’s translocation from the endoplasmic reticulum of an infected cell to the cytosol<ref name="Sandvig">Sandvig, K., & van Deurs, B. (2000). Entry of ricin and Shiga toxin into cells: molecular mechanisms and medical perspectives. The EMBO Journal, 19(22), 5943–5950. doi:10.1093/emboj/19.22.5943</ref>. The A subunit can be broken down into A1 and A2 units. This break down occurs when a trypsin sensitive region, residues 248-251, is cleaved. This cleavage results in the only remaining link being a disulfide bridge between cysteine 241 and cysteine 260. If the disulfide bridge is reduced, the A1 and A2 subunits can completely separate. Interestingly, the disulfide bridge blocks the active site of the A subunit, so the A subunit is not enzymatically active unless it has been cleaved<ref name ="Fraser">Fraser, M. E., Fujinaga, M., Cherney, M. M., Melton-Celsa, A. R., Twiddy, E. M., O’Brien, A. D., & James, M. N. G. (2004). Structure of shiga toxin type 2 (Stx2) from Escherichia coli O157:H7. The Journal of Biological Chemistry, 279(26), 27511–27517. doi:10.1074/jbc.M401939200</ref>.
The B subunit is a symmetrical pentamer that resembles a star and can be divided into five identical subunits. Each of the subunits is 69 amino acids in length. The purpose of the B subunit is to bind to globotriaosylceramide (GB3) which is a glycosphingolipid that is found on the lipid rafts of endothelial cells. Each monomer of the B subunit has three binding sites for GB3, binding to the carbohydrate portion of GB3. The affinity between the B subunit and GB3 is incredibly low, and is actually one of the lowest recorded affinities for carbohydrate-protein interaction<ref>Jacobson, J. M., Yin, J., Kitov, P. I., Mulvey, G., Griener, T. P., James, M. N. G., … Bundle, D. R. (2014). The crystal structure of Shiga toxin type 2 with bound disaccharide guides the design of a heterobifunctional toxin inhibitor. Journal of Biological Chemistry, 289(2), 885–894. doi:10.1074/jbc.M113.518886</ref>. Some studies have shown that the level of saturation and length of the fatty acid on GB3 affects the strength of its interaction with the B subunit. Since the B subunit of stx has fifteen binding sites though, it has an incredibly high avidity<ref name="Melton">Melton-Celsa, Angela R. “Shiga Toxin (Stx) Classification, Structure, and Function.” Microbiology spectrum 2.2 (2014): 10.1128/microbiolspec.EHEC–0024–2013. PMC. Web</ref>.
== Function ==
The pathway of stx entering a cell begins with the B subunit’s binding to GB3. Once this occurs, the A subunit disconnects from the B subunit and enters the cell through endocytosis. Using retrograde transport the A subunit passes through the Golgi apparatus and the rough endoplasmic reticulum<ref name="Melton">Melton-Celsa, Angela R. “Shiga Toxin (Stx) Classification, Structure, and Function.” Microbiology spectrum 2.2 (2014): 10.1128/microbiolspec.EHEC–0024–2013. PMC. Web</ref> In the rough endoplasmic reticulum, the A subunit is split into two parts called A1 and A2 through the cleavage of trypsin sensitive residues and the reduction of a disulfide bridge. A2 is degraded, but A1 freely enters the cytosol<ref name="Sandvig">Sandvig, K., & van Deurs, B. (2000). Entry of ricin and Shiga toxin into cells: molecular mechanisms and medical perspectives. The EMBO Journal, 19(22), 5943–5950. doi:10.1093/emboj/19.22.5943</ref>. Once in the cytosol, A1 acts as an N-glycosidase, which is an enzyme that hydrolyzes bonds that link sugars. With this enzymatic activity, A1 removes an adenine from the alpha-sarcin loop in the 28S RNA of the 60S ribosomal subunit<ref name="Melton">Melton-Celsa, Angela R. “Shiga Toxin (Stx) Classification, Structure, and Function.” Microbiology spectrum 2.2 (2014): 10.1128/microbiolspec.EHEC–0024–2013. PMC. Web</ref>. The removal of the adenine prevents elongation factors from associating with the ribosomal subunit. Without elongation factors, the ribosome can no longer synthesize proteins, leading to cell death<ref name="Melton">Melton-Cesla, A. (2012). Shiga toxin classification structure and function. Changes, 29(2), 997–1003</ref>.
== Disease and It's Pathogenesis ==
The shiga toxin is produced by Shigella dysenteriae and certain strains of Escherichia coli. S. dysenteriae is a gram negative bacterium that causes disease in its host by entering into intestinal epithelial cells. It has a very low infectious dose of only ten to one hundred cells, and is contracted through the consumption of contaminated water or food. One it begins replicating in the intestines, its proliferation and secretion of stx erodes the intestinal lining and leads to dysentery<ref>Zaidi, Mussaret Bano, and Teresa Estrada-García. "Shigella: A Highly Virulent and Elusive pathogen." Current Tropical Medicine Reports 1.2 (2014): 81-87</ref>. Those with dysentery can simply have diarrhea as a symptom, but if the infection progresses, it can lead to hemorrhagic colitis (bloody stool) or hemolytic uremic syndrome (HUS). HUS is characterized as acute renal failure, thrombocytopenia, and microangiopathic anemia<ref>Kaper, James B. and Alison D. O’Brien. “Overview and Historical Perspectives.” Microbiology spectrum 2.2 (2014): 10.1128/microbiolspec.EHEC–0028–2014. PMC. Web</ref>. As stated previously, stx enters cells through its binding to GB3, but curiously many intestinal cells do not express GB3. Some figure it enters the cells through macropinocyosis, but when macropinocytotic function is removed from cells, stx still manages to enter through an undefined mechanism.
The genes necessary to produce the toxin are interestingly not native to S. dysenteriae. The genes for stx are carried by prophages: bacteria that have had viral DNA inserted into their genome. During certain conditions, the phage genes are expressed leading to the production of more bacteriophages as well as the shiga toxin. When the bacteria undergo lysis, the bacteriophages and stx are released. It may seem as if there is no benefit for S. dysenteriae to carry the viral DNA, but studies have shown that while it is in the lysogenic cycle S. dysenteriae has increased acid tolerance and motility<ref>Krüger, Alejandra, and Paula María Alejandra Lucchesi. "Shiga toxins and stx-phages: highly diverse entities." Microbiology (2014): mic-0</ref>.

This is a sample scene created with SAT to <scene name='68/687332/Scene_1/1'>TextToBeDisplayed</scene> by Group, and another to make <scene name="/12/3456/Sample/2">a transparent representation</scene> of the protein. You can make your own scenes on SAT starting from scratch or loading and editing one of these sample scenes.

</StructureSection>
== References ==
<references/>

Sandbox Reserved 982

2015-05-01T05:41:35Z

Joseph P. Hoffmann:

{{Telford_CHEM321}}
==Shiga Toxin==
<StructureSection load='1R4Q' size='340' side='right' caption='Crystal Structure for Shiga Toxin (1R4Q)' scene=''>

== Structural Highlights ==
Shiga toxin (stx) has multiple classifications. The shiga toxin produced by Shigella dysenteriae is classified as stx, while shiga toxin produced by Escherichia coli can be classified as either stx1 or stx2. All three classifications very slightly in structure but have identical functions. The A subunit of stx1 differs from stx by only one residue: a serine in position 45 as opposed to threonine<ref>Fraser, M. E., Fujinaga, M., Cherney, M. M., Melton-Celsa, A. R., Twiddy, E. M., O’Brien, A. D., & James, M. N. G. (2004). Structure of shiga toxin type 2 (Stx2) from Escherichia coli O157:H7. The Journal of Biological Chemistry, 279(26), 27511–27517. doi:10.1074/jbc.M401939200</ref>. Stx is a 70 kDa AB5 protein, meaning that it is composed of an A subunit bound to a B subunit pentamer. A noncovalent interaction causes the association between the A and B subunits, as the carboxy terminal tail of the A subunit is surrounded by the B pentamer. The A subunit is 293 amino acids long with its active site being glutamic acid 167. Though the active site is a single residue, studies have shown that the first 239 residues are essential for the enzymatic activity of the A subunit. Residues 240-251 are essential for the A subunit’s translocation from the endoplasmic reticulum of an infected cell to the cytosol<ref>Sandvig, K., & van Deurs, B. (2000). Entry of ricin and Shiga toxin into cells: molecular mechanisms and medical perspectives. The EMBO Journal, 19(22), 5943–5950. doi:10.1093/emboj/19.22.5943</ref>. The A subunit can be broken down into A1 and A2 units. This break down occurs when a trypsin sensitive region, residues 248-251, is cleaved. This cleavage results in the only remaining link being a disulfide bridge between cysteine 241 and cysteine 260. If the disulfide bridge is reduced, the A1 and A2 subunits can completely separate. Interestingly, the disulfide bridge blocks the active site of the A subunit, so the A subunit is not enzymatically active unless it has been cleaved<ref>Fraser, M. E., Fujinaga, M., Cherney, M. M., Melton-Celsa, A. R., Twiddy, E. M., O’Brien, A. D., & James, M. N. G. (2004). Structure of shiga toxin type 2 (Stx2) from Escherichia coli O157:H7. The Journal of Biological Chemistry, 279(26), 27511–27517. doi:10.1074/jbc.M401939200</ref>.
The B subunit is a symmetrical pentamer that resembles a star and can be divided into five identical subunits. Each of the subunits is 69 amino acids in length. The purpose of the B subunit is to bind to globotriaosylceramide (GB3) which is a glycosphingolipid that is found on the lipid rafts of endothelial cells. Each monomer of the B subunit has three binding sites for GB3, binding to the carbohydrate portion of GB3. The affinity between the B subunit and GB3 is incredibly low, and is actually one of the lowest recorded affinities for carbohydrate-protein interaction. Some studies have shown that the level of saturation and length of the fatty acid on GB3 affects the strength of its interaction with the B subunit. Since the B subunit of stx has fifteen binding sites though, it has an incredibly high avidity<ref>Melton-Cesla, A. (2012). Shiga toxin classification structure and function. Changes, 29(2), 997–1003. doi:10.1016/j.biotechadv.2011.08.021.Secreted</ref>.
== Function ==
The pathway of stx entering a cell begins with the B subunit’s binding to GB3. Once this occurs, the A subunit disconnects from the B subunit and enters the cell through endocytosis. Using retrograde transport the A subunit passes through the Golgi apparatus and the rough endoplasmic reticulum. In the rough endoplasmic reticulum, the A subunit is split into two parts called A1 and A2 through the cleavage of trypsin sensitive residues and the reduction of a disulfide bridge. A2 is degraded, but A1 freely enters the cytosol (Sandvig 2000). Once in the cytosol, A1 acts as an N-glycosidase, which is an enzyme that hydrolyzes bonds that link sugars. With this enzymatic activity, A1 removes an adenine from the alpha-sarcin loop in the 28S RNA of the 60S ribosomal subunit (Melton-Celsa 2013). The removal of the adenine prevents elongation factors from associating with the ribosomal subunit. Without elongation factors, the ribosome can no longer synthesize proteins, leading to cell death.
== Disease and It's Pathogenesis ==
The shiga toxin is produced by Shigella dysenteriae and certain strains of Escherichia coli. S. dysenteriae is a gram negative bacterium that causes disease in its host by entering into intestinal epithelial cells (Zaidi 2014). It has a very low infectious dose of only ten to one hundred cells, and is contracted through the consumption of contaminated water or food. One it begins replicating in the intestines, its proliferation and secretion of stx erodes the intestinal lining and leads to dysentery. Those with dysentery can simply have diarrhea as a symptom, but if the infection progresses, it can lead to hemorrhagic colitis (bloody stool) or hemolytic uremic syndrome (HUS). HUS is characterized as acute renal failure, thrombocytopenia, and microangiopathic anemia (Kaper). As stated previously, stx enters cells through its binding to GB3, but curiously many intestinal cells do not express GB3. Some figure it enters the cells through macropinocyosis, but when macropinocytotic function is removed from cells, stx still manages to enter through an undefined mechanism.
The genes necessary to produce the toxin are interestingly not native to S. dysenteriae. The genes for stx are carried by prophages: bacteria that have had viral DNA inserted into their genome. During certain conditions, the phage genes are expressed leading to the production of more bacteriophages as well as the shiga toxin. When the bacteria undergo lysis, the bacteriophages and stx are released. It may seem as if there is no benefit for S. dysenteriae to carry the viral DNA, but studies have shown that while it is in the lysogenic cycle S. dysenteriae has increased acid tolerance and motility (Krüger 2014).

This is a sample scene created with SAT to <scene name='68/687332/Scene_1/1'>TextToBeDisplayed</scene> by Group, and another to make <scene name="/12/3456/Sample/2">a transparent representation</scene> of the protein. You can make your own scenes on SAT starting from scratch or loading and editing one of these sample scenes.

</StructureSection>
== References ==
<references/>

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{{Telford_CHEM321}}
==Shiga Toxin==
<StructureSection load='1R4Q' size='340' side='right' caption='Crystal Structure for Shiga Toxin (1R4Q)' scene=''>

== Structural Highlights ==
Shiga toxin (stx) has multiple classifications. The shiga toxin produced by Shigella dysenteriae is classified as stx, while shiga toxin produced by Escherichia coli can be classified as either stx1 or stx2. All three classifications very slightly in structure but have identical functions. The A subunit of stx1 differs from stx by only one residue: a serine in position 45 as opposed to threonine<ref>Fraser, M. E., Fujinaga, M., Cherney, M. M., Melton-Celsa, A. R., Twiddy, E. M., O’Brien, A. D., & James, M. N. G. (2004). Structure of shiga toxin type 2 (Stx2) from Escherichia coli O157:H7. The Journal of Biological Chemistry, 279(26), 27511–27517. doi:10.1074/jbc.M401939200</ref>. Stx is a 70 kDa AB5 protein, meaning that it is composed of an A subunit bound to a B subunit pentamer. A noncovalent interaction causes the association between the A and B subunits, as the carboxy terminal tail of the A subunit is surrounded by the B pentamer. The A subunit is 293 amino acids long with its active site being glutamic acid 167. Though the active site is a single residue, studies have shown that the first 239 residues are essential for the enzymatic activity of the A subunit. Residues 240-251 are essential for the A subunit’s translocation from the endoplasmic reticulum of an infected cell to the cytosol. The A subunit can be broken down into A1 and A2 units. This break down occurs when a trypsin sensitive region, residues 248-251 is cleaved. This cleavage results in the only remaining link being a disulfide bridge between cysteine 241 and cysteine 260. If the disulfide bridge is reduced, the A1 and A2 subunits can completely separate. Interestingly, the disulfide bridge blocks the active site of the A subunit, so the A subunit is not enzymatically active unless it has been cleaved.
The B subunit is a symmetrical pentamer that resembles a star and can be divided into five identical subunits. Each of the subunits is 69 amino acids in length. The purpose of the B subunit is to bind to globotriaosylceramide (GB3) which is a glycosphingolipid that is found on the lipid rafts of endothelial cells. Each monomer of the B subunit has three binding sites for GB3, binding to the carbohydrate portion of GB3. The affinity between the B subunit and GB3 is incredibly low, and is actually one of the lowest recorded affinities for carbohydrate-protein interaction. Some studies have shown that the level of saturation and length of the fatty acid on GB3 affects the strength of its interaction with the B subunit. Since the B subunit of stx has fifteen binding sites though, it has an incredibly high avidity (Melton-Celsa 2013).
== Function ==
The pathway of stx entering a cell begins with the B subunit’s binding to GB3. Once this occurs, the A subunit disconnects from the B subunit and enters the cell through endocytosis. Using retrograde transport the A subunit passes through the Golgi apparatus and the rough endoplasmic reticulum. In the rough endoplasmic reticulum, the A subunit is split into two parts called A1 and A2 through the cleavage of trypsin sensitive residues and the reduction of a disulfide bridge. A2 is degraded, but A1 freely enters the cytosol (Sandvig 2000). Once in the cytosol, A1 acts as an N-glycosidase, which is an enzyme that hydrolyzes bonds that link sugars. With this enzymatic activity, A1 removes an adenine from the alpha-sarcin loop in the 28S RNA of the 60S ribosomal subunit (Melton-Celsa 2013). The removal of the adenine prevents elongation factors from associating with the ribosomal subunit. Without elongation factors, the ribosome can no longer synthesize proteins, leading to cell death.
== Disease and It's Pathogenesis ==
The shiga toxin is produced by Shigella dysenteriae and certain strains of Escherichia coli. S. dysenteriae is a gram negative bacterium that causes disease in its host by entering into intestinal epithelial cells (Zaidi 2014). It has a very low infectious dose of only ten to one hundred cells, and is contracted through the consumption of contaminated water or food. One it begins replicating in the intestines, its proliferation and secretion of stx erodes the intestinal lining and leads to dysentery. Those with dysentery can simply have diarrhea as a symptom, but if the infection progresses, it can lead to hemorrhagic colitis (bloody stool) or hemolytic uremic syndrome (HUS). HUS is characterized as acute renal failure, thrombocytopenia, and microangiopathic anemia (Kaper). As stated previously, stx enters cells through its binding to GB3, but curiously many intestinal cells do not express GB3. Some figure it enters the cells through macropinocyosis, but when macropinocytotic function is removed from cells, stx still manages to enter through an undefined mechanism.
The genes necessary to produce the toxin are interestingly not native to S. dysenteriae. The genes for stx are carried by prophages: bacteria that have had viral DNA inserted into their genome. During certain conditions, the phage genes are expressed leading to the production of more bacteriophages as well as the shiga toxin. When the bacteria undergo lysis, the bacteriophages and stx are released. It may seem as if there is no benefit for S. dysenteriae to carry the viral DNA, but studies have shown that while it is in the lysogenic cycle S. dysenteriae has increased acid tolerance and motility (Krüger 2014).

This is a sample scene created with SAT to <scene name='68/687332/Scene_1/1'>TextToBeDisplayed</scene> by Group, and another to make <scene name="/12/3456/Sample/2">a transparent representation</scene> of the protein. You can make your own scenes on SAT starting from scratch or loading and editing one of these sample scenes.

</StructureSection>
== References ==
<references/>

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{{Telford_CHEM321}}
==Shiga Toxin==
<StructureSection load='1R4Q' size='340' side='right' caption='Crystal Structure for Shiga Toxin (1R4Q)' scene=''>

== Structural Highlights ==
<scene name='68/687332/Shiga_toxin/5'>Crystal Structure of Shiga Toxin</scene>
Stx is composed of two main subunits labeled A and B. The A subunit bonds noncovalently to the B subunit. The A subunit is 293 amino acids long with its active site being glutamic acid 167. Studies have shown that the first 239 residues are essential for the enzymatic activity of the A subunit. The B subunit is a pentamer that resembles a star and can be divided into five identical subunits. Each of the subunits is 69 amino acids in length. The purpose of the B subunit is to bind to globotriaosylceramide (GB3) which is a glycosphingolipid that resides on the surface of some Eukaryotic cells. Each monomer of the B subunit has three binding sites for GB3 making the protein very potent (Melton-Celsa 2013).
== Function ==
The pathway of stx entering a cell begins with the B subunit’s binding to GB3. Once this occurs, the A subunit disconnects from the B subunit and enters the cell through endocytosis. Using retrograde transport the A subunit passes through the Golgi apparatus and the rough endoplasmic reticulum. In the rough endoplasmic reticulum, the A subunit is cleaved into two parts called A1 and A2. A2 is degraded, but A1 freely enters the cytosol (Sandvig 2000). Once in the cytosol, A1 acts as an N-glycosidase, which is an enzyme that hydrolyzes bonds that link sugars. With this enzymatic activity, A1 removes adenines from the 28S RNA of the 60S ribosomal subunit (Melton-Celsa 2013). This inhibits protein synthesis and ultimately leads to cell death.

== Disease and It's Pathogenesis ==

This is a sample scene created with SAT to <scene name='68/687332/Scene_1/1'>TextToBeDisplayed</scene> by Group, and another to make <scene name="/12/3456/Sample/2">a transparent representation</scene> of the protein. You can make your own scenes on SAT starting from scratch or loading and editing one of these sample scenes.

</StructureSection>
== References ==
<references/>

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Joseph P. Hoffmann:

{{Telford_CHEM321}}
==Shiga Toxin==
<StructureSection load='1R4Q' size='340' side='right' caption='Crystal Structure for Shiga Toxin (1R4Q)' scene=''>

== Structural Highlights ==
<scene name='68/687332/Shiga_toxin/5'>Crystal Structure of Shiga Toxin</scene>
Stx is composed of two main subunits labeled A and B. The A subunit bonds noncovalently to the B subunit. The A subunit is 293 amino acids long with its active site being glutamic acid 167. Studies have shown that the first 239 residues are essential for the enzymatic activity of the A subunit. The B subunit is a pentamer that resembles a star and can be divided into five identical subunits. Each of the subunits is 69 amino acids in length. The purpose of the B subunit is to bind to globotriaosylceramide (GB3) which is a glycosphingolipid that resides on the surface of some Eukaryotic cells. Each monomer of the B subunit has three binding sites for GB3 making the protein very potent (Melton-Celsa 2013).
== Function ==
The pathway of stx entering a cell begins with the B subunit’s binding to GB3. Once this occurs, the A subunit disconnects from the B subunit and enters the cell through endocytosis. Using retrograde transport the A subunit passes through the Golgi apparatus and the rough endoplasmic reticulum. In the rough endoplasmic reticulum, the A subunit is cleaved into two parts called A1 and A2. A2 is degraded, but A1 freely enters the cytosol (Sandvig 2000). Once in the cytosol, A1 acts as an N-glycosidase, which is an enzyme that hydrolyzes bonds that link sugars. With this enzymatic activity, A1 removes adenines from the 28S RNA of the 60S ribosomal subunit (Melton-Celsa 2013). This inhibits protein synthesis and ultimately leads to cell death.
== Function ==

== Disease and It's Pathogenesis ==

This is a sample scene created with SAT to <scene name='68/687332/Scene_1/1'>TextToBeDisplayed</scene> by Group, and another to make <scene name="/12/3456/Sample/2">a transparent representation</scene> of the protein. You can make your own scenes on SAT starting from scratch or loading and editing one of these sample scenes.

</StructureSection>
== References ==
<references/>

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Joseph P. Hoffmann:

{{Telford_CHEM321}}
==Your Heading Here (maybe something like 'Structure')==
<StructureSection load='1R4Q' size='340' side='right' caption='Crystal Structure for Shiga Toxin (1R4Q)' scene=''>

== Structure ==
<scene name='68/687332/Shiga_toxin/5'>Crystal Structure of Shiga Toxin</scene>
Stx is composed of two main subunits labeled A and B. The A subunit bonds noncovalently to the B subunit. The A subunit is 293 amino acids long with its active site being glutamic acid 167. Studies have shown that the first 239 residues are essential for the enzymatic activity of the A subunit. The B subunit is a pentamer that resembles a star and can be divided into five identical subunits. Each of the subunits is 69 amino acids in length. The purpose of the B subunit is to bind to globotriaosylceramide (GB3) which is a glycosphingolipid that resides on the surface of some Eukaryotic cells. Each monomer of the B subunit has three binding sites for GB3 making the protein very potent (Melton-Celsa 2013).
== Function ==
The pathway of stx entering a cell begins with the B subunit’s binding to GB3. Once this occurs, the A subunit disconnects from the B subunit and enters the cell through endocytosis. Using retrograde transport the A subunit passes through the Golgi apparatus and the rough endoplasmic reticulum. In the rough endoplasmic reticulum, the A subunit is cleaved into two parts called A1 and A2. A2 is degraded, but A1 freely enters the cytosol (Sandvig 2000). Once in the cytosol, A1 acts as an N-glycosidase, which is an enzyme that hydrolyzes bonds that link sugars. With this enzymatic activity, A1 removes adenines from the 28S RNA of the 60S ribosomal subunit (Melton-Celsa 2013). This inhibits protein synthesis and ultimately leads to cell death.
== Disease ==

== Relevance ==

== Structural highlights ==

This is a sample scene created with SAT to <scene name='68/687332/Scene_1/1'>TextToBeDisplayed</scene> by Group, and another to make <scene name="/12/3456/Sample/2">a transparent representation</scene> of the protein. You can make your own scenes on SAT starting from scratch or loading and editing one of these sample scenes.

</StructureSection>
== References ==
<references/>

Sandbox Reserved 982

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{{Telford_CHEM321}}
==Your Heading Here (maybe something like 'Structure')==
<StructureSection load='1stp' size='340' side='right' caption='Caption for this structure' scene=''>
This is a default text for your page ''''''. Click above on '''edit this page''' to modify. Be careful with the < and > signs.
You may include any references to papers as in: the use of JSmol in Proteopedia <ref>DOI 10.1002/ijch.201300024</ref> or to the article describing Jmol <ref>PMID:21638687</ref> to the rescue.

== Structure ==
<scene name='68/687332/Shiga_toxin/5'>Crystal Structure of Shiga Toxin</scene>
Stx is composed of two main subunits labeled A and B. The A subunit bonds noncovalently to the B subunit. The A subunit is 293 amino acids long with its active site being glutamic acid 167. Studies have shown that the first 239 residues are essential for the enzymatic activity of the A subunit. The B subunit is a pentamer that resembles a star and can be divided into five identical subunits. Each of the subunits is 69 amino acids in length. The purpose of the B subunit is to bind to globotriaosylceramide (GB3) which is a glycosphingolipid that resides on the surface of some Eukaryotic cells. Each monomer of the B subunit has three binding sites for GB3 making the protein very potent (Melton-Celsa 2013).
== Function ==
The pathway of stx entering a cell begins with the B subunit’s binding to GB3. Once this occurs, the A subunit disconnects from the B subunit and enters the cell through endocytosis. Using retrograde transport the A subunit passes through the Golgi apparatus and the rough endoplasmic reticulum. In the rough endoplasmic reticulum, the A subunit is cleaved into two parts called A1 and A2. A2 is degraded, but A1 freely enters the cytosol (Sandvig 2000). Once in the cytosol, A1 acts as an N-glycosidase, which is an enzyme that hydrolyzes bonds that link sugars. With this enzymatic activity, A1 removes adenines from the 28S RNA of the 60S ribosomal subunit (Melton-Celsa 2013). This inhibits protein synthesis and ultimately leads to cell death.
== Disease ==

== Relevance ==

== Structural highlights ==

This is a sample scene created with SAT to <scene name='68/687332/Scene_1/1'>TextToBeDisplayed</scene> by Group, and another to make <scene name="/12/3456/Sample/2">a transparent representation</scene> of the protein. You can make your own scenes on SAT starting from scratch or loading and editing one of these sample scenes.

</StructureSection>
== References ==
<references/>

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