Sandbox Reserved 1096: Difference between revisions

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==='''Primary, secondary and tertiary structure'''===

==='''Calcium binding sites and active site'''===

The structure of the '''''apoenzyme''''' '''''apoPAD2''''' shows a stable head to tail dimer.The monomer is formed by 2 immunoglobulin-like domains and a C-term catalytic domain calcium binding site. There are six different calcium binding sites '''''C1 to C6''''' C2-5 are unoccupied in apoPAD2 but there is an electron density on C1 and C6 so those are occupied by calcium~~<ref>PMID:25621824<ref/>~~.

The structure of the '''''apoenzyme''''' '''''apoPAD2''''' shows a stable head to tail dimer.The monomer is formed by 2 immunoglobulin-like domains and a C-term catalytic domain calcium binding site. There are six different calcium binding sites '''''C1 to C6''''' C2-5 are unoccupied in apoPAD2 but there is an electron density on C1 and C6 so those are occupied by calcium.

The structure of the '''''PAD2.Ca2+ complex''''' in 10mM of CaCl2 differs from the apoPAD2 one's. There is the same folding except C3-5 are occupied. Even though just one site is unoccupied, the structure is not catalytically competent, it is explained by the active site nucleophile C647, indeed, it is just 12 angstrom away from the catalytic center. Yet, the 3 other key catalytic residues, D351, H471, D473 are properly positioned to promote catalysis and since they have the same conformation in both the apoenzyme and the holoenzyme, the complex structure PAD2+/Ca2+ represent an intermediaire between the 2 structures. This intermediaire structure is stabilized thanks to hydrogen bonds between R347 and Q350 who are in the active site. It also inhibits the movement of C647 who is in the substrate binding pocket~~<ref>PMID:25621824<ref/>~~.

The structure of the '''''PAD2.Ca2+ complex''''' in 10mM of CaCl2 differs from the apoPAD2 one's. There is the same folding except C3-5 are occupied. Even though just one site is unoccupied, the structure is not catalytically competent, it is explained by the active site nucleophile C647, indeed, it is just 12 angstrom away from the catalytic center. Yet, the 3 other key catalytic residues, D351, H471, D473 are properly positioned to promote catalysis and since they have the same conformation in both the apoenzyme and the holoenzyme, the complex structure PAD2+/Ca2+ represent an intermediaire between the 2 structures. This intermediaire structure is stabilized thanks to hydrogen bonds between R347 and Q350 who are in the active site. It also inhibits the movement of C647 who is in the substrate binding pocket.

To obtain the structure of the holoenzyme '''''PAD2''''', the scientists had engineer a double mutant, because the F221 and F222 binds in a hydrophobic pocket, this prevents the calcium-binding at Ca2.

So they studied the structure of the PAD2 F221/222A.Ca2+ mutant. It shows an important electron density at all 6 calcium binding sites. All sites are now binding with calcium. Moreover, The active site cysteine, C647 points toward the catalytic center. this conformation is competent for catalysis. It is not the only effect of calcium binding to Ca2 site, it also causes R347 to move out of the active site while W348 moves in to form one wall of the substrate-binding pocket~~<ref>PMID:25621824<ref/>~~.

So they studied the structure of the PAD2 F221/222A.Ca2+ mutant. It shows an important electron density at all 6 calcium binding sites. All sites are now binding with calcium. Moreover, The active site cysteine, C647 points toward the catalytic center. this conformation is competent for catalysis. It is not the only effect of calcium binding to Ca2 site, it also causes R347 to move out of the active site while W348 moves in to form one wall of the substrate-binding pocket.

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==='''PAD2 and ER Target-gene Expression in Breast Cancer'''===

PAD2 functions as an [https://en.wikipedia.org/wiki/Estrogen_receptor Estrogen Receptor] (ER) coactivator in [https://en.wikipedia.org/wiki/Breast_cancer Breast cancer] cells, using the citrullination of histone tail arginine residues at ER binding sites.~~<ref>PMID:25621824<ref/>~~ This makes it an attractive therapeutic target, yet, the mechanisms that regulates it are for a major part unknown. Indeed, it is used as a component of ER-related gene expression that is positively correlated with [https://proteopedia.org/wiki/index.php/Herceptin_-_Mechanism_of_Action HER2] protein levels in breast cancer cell lines, and in primary HER2+ breast tumors<ref>PMID:25621824<ref/>. This shows that PAD2 activity plays a role in breast cancer progression. The inhibition of PAD2 decreases ER target-gene expression. Moreover, when in presence of CL-amidine, which is PAD inhibitor, the tumor burden in a breast cancer~~<ref>PMID:25621824<ref/>~~.

PAD2 functions as an [https://en.wikipedia.org/wiki/Estrogen_receptor Estrogen Receptor] (ER) coactivator in [https://en.wikipedia.org/wiki/Breast_cancer Breast cancer] cells, using the citrullination of histone tail arginine residues at ER binding sites. This makes it an attractive therapeutic target, yet, the mechanisms that regulates it are for a major part unknown. Indeed, it is used as a component of ER-related gene expression that is positively correlated with [https://proteopedia.org/wiki/index.php/Herceptin_-_Mechanism_of_Action HER2] protein levels in breast cancer cell lines, and in primary HER2+ breast tumors<ref>PMID:25621824<ref/>. This shows that PAD2 activity plays a role in breast cancer progression. The inhibition of PAD2 decreases ER target-gene expression. Moreover, when in presence of CL-amidine, which is PAD inhibitor, the tumor burden in a breast cancer.

==Publication Abstract from ACS Publications <ref name="ART1" />==

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@@ Line 9: / Line 9: @@
 ==='''Primary, secondary and tertiary structure'''===
 ==='''Calcium binding sites and active site'''===
-The structure of the '''''apoenzyme''''' '''''apoPAD2''''' shows a stable head to tail dimer.The monomer is formed by 2 immunoglobulin-like domains and a C-term catalytic domain calcium binding site. There are six different calcium binding sites '''''C1 to C6''''' C2-5 are unoccupied in apoPAD2 but there is an electron density on C1 and C6 so those are occupied by calcium<ref>PMID:25621824<ref/>.
+The structure of the '''''apoenzyme''''' '''''apoPAD2''''' shows a stable head to tail dimer.The monomer is formed by 2 immunoglobulin-like domains and a C-term catalytic domain calcium binding site. There are six different calcium binding sites '''''C1 to C6''''' C2-5 are unoccupied in apoPAD2 but there is an electron density on C1 and C6 so those are occupied by calcium.
-The structure of the '''''PAD2.Ca2+ complex''''' in 10mM of CaCl2 differs from the apoPAD2 one's. There is the same folding except C3-5 are occupied. Even though just one site is unoccupied, the structure is not catalytically competent, it is explained by the active site nucleophile C647, indeed, it is just 12 angstrom away from the catalytic center. Yet, the 3 other key catalytic residues, D351, H471, D473 are properly positioned to promote catalysis and since they have the same conformation in both the apoenzyme and the holoenzyme, the complex structure PAD2+/Ca2+ represent an intermediaire between the 2 structures. This intermediaire structure is stabilized thanks to hydrogen bonds between R347 and Q350 who are in the active site. It also inhibits the movement of C647 who is in the substrate binding pocket<ref>PMID:25621824<ref/>.
+The structure of the '''''PAD2.Ca2+ complex''''' in 10mM of CaCl2 differs from the apoPAD2 one's. There is the same folding except C3-5 are occupied. Even though just one site is unoccupied, the structure is not catalytically competent, it is explained by the active site nucleophile C647, indeed, it is just 12 angstrom away from the catalytic center. Yet, the 3 other key catalytic residues, D351, H471, D473 are properly positioned to promote catalysis and since they have the same conformation in both the apoenzyme and the holoenzyme, the complex structure PAD2+/Ca2+ represent an intermediaire between the 2 structures. This intermediaire structure is stabilized thanks to hydrogen bonds between R347 and Q350 who are in the active site. It also inhibits the movement of C647 who is in the substrate binding pocket.
 To obtain the structure of the holoenzyme '''''PAD2''''', the scientists had engineer a double mutant, because the F221 and F222 binds in a hydrophobic pocket, this prevents the calcium-binding at Ca2.
-So they studied the structure of the PAD2 F221/222A.Ca2+ mutant. It shows an important electron density at all 6 calcium binding sites. All sites are now binding with calcium. Moreover, The active site cysteine, C647 points toward the catalytic center. this conformation is competent for catalysis. It is not the only effect of calcium binding to Ca2 site, it also causes R347 to move out of the active site while W348 moves in to form one wall of the substrate-binding pocket<ref>PMID:25621824<ref/>.
+So they studied the structure of the PAD2 F221/222A.Ca2+ mutant. It shows an important electron density at all 6 calcium binding sites. All sites are now binding with calcium. Moreover, The active site cysteine, C647 points toward the catalytic center. this conformation is competent for catalysis. It is not the only effect of calcium binding to Ca2 site, it also causes R347 to move out of the active site while W348 moves in to form one wall of the substrate-binding pocket.
@@ Line 28: / Line 28: @@
 ==='''PAD2 and ER Target-gene Expression in Breast Cancer'''===
-PAD2 functions as an [https://en.wikipedia.org/wiki/Estrogen_receptor Estrogen Receptor] (ER) coactivator in [https://en.wikipedia.org/wiki/Breast_cancer Breast cancer] cells, using the citrullination of histone tail arginine residues at ER binding sites.<ref>PMID:25621824<ref/> This makes it an attractive therapeutic target, yet, the mechanisms that regulates it are for a major part unknown. Indeed, it is used as a component of ER-related gene expression that is positively correlated with [https://proteopedia.org/wiki/index.php/Herceptin_-_Mechanism_of_Action HER2] protein levels in breast cancer cell lines, and in primary HER2+ breast tumors<ref>PMID:25621824<ref/>. This shows that PAD2 activity plays a role in breast cancer progression. The inhibition of PAD2 decreases ER target-gene expression. Moreover, when in presence of CL-amidine, which is PAD inhibitor, the tumor burden in a breast cancer<ref>PMID:25621824<ref/>.
+PAD2 functions as an [https://en.wikipedia.org/wiki/Estrogen_receptor Estrogen Receptor] (ER) coactivator in [https://en.wikipedia.org/wiki/Breast_cancer Breast cancer] cells, using the citrullination of histone tail arginine residues at ER binding sites. This makes it an attractive therapeutic target, yet, the mechanisms that regulates it are for a major part unknown. Indeed, it is used as a component of ER-related gene expression that is positively correlated with [https://proteopedia.org/wiki/index.php/Herceptin_-_Mechanism_of_Action HER2] protein levels in breast cancer cell lines, and in primary HER2+ breast tumors<ref>PMID:25621824<ref/>. This shows that PAD2 activity plays a role in breast cancer progression. The inhibition of PAD2 decreases ER target-gene expression. Moreover, when in presence of CL-amidine, which is PAD inhibitor, the tumor burden in a breast cancer.
 ==Publication Abstract from ACS Publications <ref name="ART1" />==