Sandbox Reserved 1096: Difference between revisions

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==Structural Features==

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

In terms of primary structure, the '''''apoenzyme''''' described under PDB's [https://www.rcsb.org/structure/4N20 4N20] contains 4 key catalytic residues that are essential for citrullination in the active site (<scene name='82/829349/C647/1'>C647</scene>, D351, H471 and D473). The cysteine residue is involved in nucleophilic attacks that promote deimination of Arginine residues whereas the other residues are mainly responsible for stabilization of substrates entering the active site<ref name="ART1" />. Other key residues include D125, D127, E131 and E354 which may play an important role in Calcium binding for sites Ca1 and Ca2 given that side-chains are around 5Å away from Calcium cations. Considering both Aspartate and Glutamate are negatively charged at pH 7, electrostatic interactions may exist between these residues and Ca2+ in binding sites. In the other hand, the structure of '''''apoPAD2''''' shows some common secondary motifs such as superimposed anti-parallel beta sheets that generate beta sandiwiches. Closer to the C-terminal, both alpha and beta secondary structures may be observed with 17 alpha helixes and multiple parallel and anti-parallel beta sheets that make up an alpha/beta propeller<ref name="ART1" />. Tertiary structure of '''''apoPAD2''''' monomer is well described by [https://pubs.acs.org/doi/10.1021/cb500933j Slade et al.] as being composed of three distinct domains: 2 immunoglobin-like domains (IgG1 and IgG2) at positions (1-115)(116-295) respectively and a C-terminal catalytic domain at (296-665) that contains the active site. Both <scene name='82/829349/Igg_domains/1'>immunoglobin-like domains</scene> comprise typical beta-sandwiches and the catalytic domain generates an alpha/beta propeller. Finally, with respect to the quaternary structure, '''''PAD2 Apoenzyme''''' cristallises as a stable dimer of identical subunits (monomers). The biological assembly of the enzyme was thus observed to be an association of two monomers whose structural highlights were described above.

In terms of primary structure, the '''''apoenzyme''''' described under PDB's [https://www.rcsb.org/structure/4N20 4N20] contains 4 key catalytic residues that are essential for citrullination in the active site (<scene name='82/829349/C647/1'>C647</scene>, D351, H471 and D473). The cysteine residue is involved in nucleophilic attacks that promote deimination of Arginine residues whereas the other residues are mainly responsible for stabilization of substrates entering the active site<ref name="ART1" />. Other key residues include D125, D127, E131 and E354 which may play an important role in Calcium binding for sites <scene name='82/829349/Ca1/1'>Ca1</scene> and Ca2 given that side-chains are around 5Å away from Calcium cations. Considering both Aspartate and Glutamate are negatively charged at pH 7, electrostatic interactions may exist between these residues and Ca2+ in binding sites. In the other hand, the structure of '''''apoPAD2''''' shows some common secondary motifs such as superimposed anti-parallel beta sheets that generate beta sandiwiches. Closer to the C-terminal, both alpha and beta secondary structures may be observed with 17 alpha helixes and multiple parallel and anti-parallel beta sheets that make up an alpha/beta propeller<ref name="ART1" />. Tertiary structure of '''''apoPAD2''''' monomer is well described by [https://pubs.acs.org/doi/10.1021/cb500933j Slade et al.] as being composed of three distinct domains: 2 immunoglobin-like domains (IgG1 and IgG2) at positions (1-115)(116-295) respectively and a C-terminal catalytic domain at (296-665) that contains the active site. Both <scene name='82/829349/Igg_domains/1'>immunoglobin-like domains</scene> comprise typical beta-sandwiches and the catalytic domain generates an alpha/beta propeller. Finally, with respect to the quaternary structure, '''''PAD2 Apoenzyme''''' cristallises as a stable dimer of identical subunits (monomers). The biological assembly of the enzyme was thus observed to be an association of two monomers whose structural highlights were described above.

==='''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 in the holoenzyme.

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 '''''(<scene name='82/829349/Ca1/1'>Ca1</scene> to Ca6)''''', Ca2-5 are unoccupied in apoPAD2 but there is an electron density on <scene name='82/829349/Ca1/1'>Ca1</scene> and Ca6 so those are occupied by calcium in the holoenzyme.

The structure of the '''''PAD2 Ca2+ complex''''' in 10mM of CaCl2 ([[4n2b]]) differs from '''''apoPAD2'''''. Folding and 3D structure remain unchanged except C3-5 are occupied by Calcium cations when Ca2+ increases<ref name="ART1" />. The '''''apoPAD2''''' enzyme resolved structure shows 2 high-affinity Calcium binding sites that remain consistent. At 10mM Ca3-5 sites are occupied by Calcium cations and, even though there is still only one unoccupied site, the structure is not catalytically competent. This may be explained by the <scene name='82/829349/C647/1'>nucleophile active site C647</scene> that is still 12 angstrom away from the catalytic center at 10mM. Yet, the 3 other key catalytic residues: D351, H471 and 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+ represents an intermediate form between the 2 structures. This intermediate structure is stabilized by hydrogen bonds between R347 and Q350 in the active site. These bonds may also inhibit the movement of <scene name='82/829349/C647/1'>C647</scene> into the substrate binding pocket<ref name="ART1" /> .

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@@ Line 9: / Line 9: @@
 ==Structural Features==
 ==='''Primary, secondary and tertiary structure'''===
-In terms of primary structure, the '''''apoenzyme''''' described under PDB's [https://www.rcsb.org/structure/4N20 4N20] contains 4 key catalytic residues that are essential for citrullination in the active site (<scene name='82/829349/C647/1'>C647</scene>, D351, H471 and D473). The cysteine residue is involved in nucleophilic attacks that promote deimination of Arginine residues whereas the other residues are mainly responsible for stabilization of substrates entering the active site<ref name="ART1" />. Other key residues include D125, D127, E131 and E354 which may play an important role in Calcium binding for sites Ca1 and Ca2 given that side-chains are around 5Å away from Calcium cations. Considering both Aspartate and Glutamate are negatively charged at pH 7, electrostatic interactions may exist between these residues and Ca2+ in binding sites. In the other hand, the structure of '''''apoPAD2''''' shows some common secondary motifs such as superimposed anti-parallel beta sheets that generate beta sandiwiches. Closer to the C-terminal, both alpha and beta secondary structures may be observed with 17 alpha helixes and multiple parallel and anti-parallel beta sheets that make up an alpha/beta propeller<ref name="ART1" />. Tertiary structure of '''''apoPAD2''''' monomer is well described by [https://pubs.acs.org/doi/10.1021/cb500933j Slade et al.] as being composed of three distinct domains: 2 immunoglobin-like domains (IgG1 and IgG2) at positions (1-115)(116-295) respectively and a C-terminal catalytic domain at (296-665) that contains the active site. Both <scene name='82/829349/Igg_domains/1'>immunoglobin-like domains</scene> comprise typical beta-sandwiches and the catalytic domain generates an alpha/beta propeller. Finally, with respect to the quaternary structure, '''''PAD2 Apoenzyme''''' cristallises as a stable dimer of identical subunits (monomers). The biological assembly of the enzyme was thus observed to be an association of two monomers whose structural highlights were described above.
+In terms of primary structure, the '''''apoenzyme''''' described under PDB's [https://www.rcsb.org/structure/4N20 4N20] contains 4 key catalytic residues that are essential for citrullination in the active site (<scene name='82/829349/C647/1'>C647</scene>, D351, H471 and D473). The cysteine residue is involved in nucleophilic attacks that promote deimination of Arginine residues whereas the other residues are mainly responsible for stabilization of substrates entering the active site<ref name="ART1" />. Other key residues include D125, D127, E131 and E354 which may play an important role in Calcium binding for sites <scene name='82/829349/Ca1/1'>Ca1</scene> and Ca2 given that side-chains are around 5Å away from Calcium cations. Considering both Aspartate and Glutamate are negatively charged at pH 7, electrostatic interactions may exist between these residues and Ca2+ in binding sites. In the other hand, the structure of '''''apoPAD2''''' shows some common secondary motifs such as superimposed anti-parallel beta sheets that generate beta sandiwiches. Closer to the C-terminal, both alpha and beta secondary structures may be observed with 17 alpha helixes and multiple parallel and anti-parallel beta sheets that make up an alpha/beta propeller<ref name="ART1" />. Tertiary structure of '''''apoPAD2''''' monomer is well described by [https://pubs.acs.org/doi/10.1021/cb500933j Slade et al.] as being composed of three distinct domains: 2 immunoglobin-like domains (IgG1 and IgG2) at positions (1-115)(116-295) respectively and a C-terminal catalytic domain at (296-665) that contains the active site. Both <scene name='82/829349/Igg_domains/1'>immunoglobin-like domains</scene> comprise typical beta-sandwiches and the catalytic domain generates an alpha/beta propeller. Finally, with respect to the quaternary structure, '''''PAD2 Apoenzyme''''' cristallises as a stable dimer of identical subunits (monomers). The biological assembly of the enzyme was thus observed to be an association of two monomers whose structural highlights were described above.
 ==='''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 in the holoenzyme.
+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 '''''(<scene name='82/829349/Ca1/1'>Ca1</scene> to Ca6)''''', Ca2-5 are unoccupied in apoPAD2 but there is an electron density on <scene name='82/829349/Ca1/1'>Ca1</scene> and Ca6 so those are occupied by calcium in the holoenzyme.
 The structure of the '''''PAD2 Ca2+ complex''''' in 10mM of CaCl2 ([[4n2b]]) differs from '''''apoPAD2'''''. Folding and 3D structure remain unchanged except C3-5 are occupied by Calcium cations when Ca2+ increases<ref name="ART1" />. The '''''apoPAD2''''' enzyme resolved structure shows 2 high-affinity Calcium binding sites that remain consistent. At 10mM Ca3-5 sites are occupied by Calcium cations and, even though there is still only one unoccupied site, the structure is not catalytically competent. This may be explained by the <scene name='82/829349/C647/1'>nucleophile active site C647</scene> that is still 12 angstrom away from the catalytic center at 10mM. Yet, the 3 other key catalytic residues: D351, H471 and 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+ represents an intermediate form between the 2 structures. This intermediate structure is stabilized by hydrogen bonds between R347 and Q350 in the active site. These bonds may also inhibit the movement of <scene name='82/829349/C647/1'>C647</scene> into the substrate binding pocket<ref name="ART1" /> .