Sandbox Reserved 486: Difference between revisions
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<Structure load='1mdt' size='500' frame='true' align='right' caption='Diphtheria toxin structure' scene='Insert optional scene name here' /> | <Structure load='1mdt' size='500' frame='true' align='right' caption='Diphtheria toxin structure' scene='Insert optional scene name here' /> | ||
== History == | == History == | ||
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== Structure == | == Structure == | ||
Diphtheria toxin is a protein made of 535 amino acids | Diphtheria toxin is a protein made of 535 amino acids and is 58kDa in weight. The [http://en.wikipedia.org/wiki/Zymogen proenzyme](zymogen) must be cleaved into fragments A & B and reduced in order for the toxicity gene to be turn on. In each fragment there are two <scene name='Sandbox_Reserved_486/Disulfide_bonds/1'>Disulfide Bonds</scene>. There are also three domains C,T, and R that have different functions. The first crystal structure of the toxin was obtained in 1992 by x-ray crystallography. However, the structure was a dimer and generated by freezing the protein. The dimer is non-toxic but becomes a toxic monomer at neutral pH by dissociation. The <scene name='Sandbox_Reserved_486/Secondary_structure/1'>Secondary Structure</scene> of the molecule is made up of 24 helices, 31 beta sheets, and turns between each of them which adds up to be 535 residue protein. This makes for a mixture of <scene name='Sandbox_Reserved_486/Hydrophobic_hydrophilic/1'>hydrophilic and hydrophobic areas</scene> that allow for the seondary structure. The purple portion of the molecule is the polar ares while the gray was the hydrophobic areas. | ||
<scene name='Sandbox_Reserved_486/ | *<scene name='Sandbox_Reserved_486/Fragment_a/1'>Fragment A</scene> is about molecular mass of 21kDa and is the catalytically active portion of the protein. | ||
*<scene name='Sandbox_Reserved_486/Fragment_b/1'>Fragment B</scene> is about 37kDa and contains the receptor binding & translocation portions of the protein. | |||
The two fragments are then separated into three domains C, T, and R which correspond to three major functions of the toxin. | |||
<scene name='Sandbox_Reserved_486/ | *In fragment A contains domain C which contains the <scene name='Sandbox_Reserved_486/Active_site/1'>Active site</scene> and does the catalysis for the protein. It contains eight β strands and seven α helices. | ||
*The T domain is found in fragment B and is the translocation portion of the protein. The structure is made up of three layers of α helices. The outer layer contains 3 helices that are rich in polar residues that allow for the molecule to maintain the T domain as well as keeping the toxin soluble in neutral pH. | |||
<scene name='Sandbox_Reserved_486/ | *Another domain in fragment B is domain R which has the <scene name='Sandbox_Reserved_486/Receptor_binding_site/1'>Receptor binding site</scene> for the protein. The domain consist mostly of β sheets and changes conformation when interacting with cell membranes. In the R domain both Lsy516 and Phe530 play a role in recognizing receptors on cell surfaces. | ||
<scene name='Sandbox_Reserved_486/ | In the mechanism for Diphtheria toxin the R Domain binds to <scene name='Sandbox_Reserved_486/Ligands/2'>ligands</scene> which are NAD<sup>+</sup> which can sometimes be switched with ApUp. The molecule ApUp is bound to a cleft on the front side of the C domain. | ||
< | [[Image:Page41-2.jpg]] <ref>Parker, M. (1996). Protein toxin structure. (1 ed., pp. 25-43). Georgetown: R.G. Landes Company.</ref> | ||
== Mechanism == | |||
The mechanism begins by the R domain recognizing the target cells by the molecules on the cells surface. A prominent β-hairpin loop of the R domain binds to the surface and allows for a docking station. The ligand that is typically bound on the cell surface is pro-HB-EFG. Binding leads to receptor mediated endocytosis. The T domain translocates the C domain into the cytosol of the cell. The organism ''Corynebacterium diphtheriae'' first secretes the toxin the loop between the C and T domain has to be nicked in order for the toxicity to be turned on. There is one surface protease named [http://en.wikipedia.org/wiki/Furin furin] that is known to nick the area but it is unclear if any other protein helps the process. This process allows for the C domain to dissociate from the T domain after transported(translocated) into the cytosol. This is where the disulfide bonds become reduced. After this happens a drop in pH leads to the formation of the dimer to become an open monomer form. The reduction of the disulfide bonds is the rate determining step of the toxin entrying the cell. | |||
< | Once entried into the cytosol, the C domain does ADP-ribosylation of EF-2 at diphthamide, a posttranslationally modified histidine residue. This shuts down all protein synthesis and kills the cell. The K<sub>cat</sub>/K<sub>M</sub> of the reaction is about 10<sup>8</sup> min<sup>-1</sup> M<sup>-1</sup>. The figure below shows the reaction happening at the C domain. The C domain binds to the ligand NAD<sup>+</sup> to begin the reaction. The C domian does have an inhibitor and that is ApUp which is near in the C domain. | ||
[[Image:Toxin-1.jpg]] figure 3: Mechanism of inactivation of elongation factor 2 (EF2) by diphtheria toxin <ref>Stephen, J., & Pietrowski, R. (1981). Bacterial toxins. (1 ed., p. 11). Washington: Van Nostrand Reinhold Co. Ltd. </ref> | [[Image:Toxin-1.jpg]] figure 3: Mechanism of inactivation of elongation factor 2 (EF2) by diphtheria toxin <ref>Stephen, J., & Pietrowski, R. (1981). Bacterial toxins. (1 ed., p. 11). Washington: Van Nostrand Reinhold Co. Ltd. </ref> | ||
== Medical Implications & Possible Application == | == Medical Implications & Possible Application == | ||
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== References == | == References == | ||
* Parker, M. (1996). Protein toxin structure. (1 ed., pp. 25-43). Georgetown: R.G. Landes Company. | * Parker, M. (1996). Protein toxin structure. (1 ed., pp. 25-43). Georgetown: R.G. Landes Company. | ||
*Bennett, M., & Eisenberg, D. (2009, febuary 24). The refined structure of monomeric diphtheria toxin at 2.3 angstroms resolution. Retrieved from http://www.pdb.org/pdb/explore/explore.do? | |||
* Stephen, J., & Pietrowski, R. (1981). Bacterial toxins. (1 ed., p. 11). Washington: Van Nostrand Reinhold Co. Ltd. | * Stephen, J., & Pietrowski, R. (1981). Bacterial toxins. (1 ed., p. 11). Washington: Van Nostrand Reinhold Co. Ltd. | ||
