Sandbox Reserved 714: Difference between revisions
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The Human soluble Epoxide hydrolase is a protein of 555 residues. In vivo, it exists under the form of a homodimer, with a monomeric unit of 62,5 kDa. Each subunit has <scene name='Sandbox_Reserved_714/Catalytic_domains/5'>two catalytic domains</scene>, linked by a proline-rich section. | The Human soluble Epoxide hydrolase is a protein of 555 residues. In vivo, it exists under the form of a homodimer, with a monomeric unit of 62,5 kDa. Each subunit has <scene name='Sandbox_Reserved_714/Catalytic_domains/5'>two catalytic domains</scene>, linked by a proline-rich section. | ||
The secondary structure of this enzyme is made of beta-sheets and alpha-helices and a few 310-helices, which form the two pockets of the active sites, in the C-term domain and the N-term domain, in which the substrates (lipid-phosphates and ions for the N-term domain, and epoxides for the C-term domain) can bind. The 3D dimensions of the enzyme are the following : a = 92.55 Å, b = 92.55 Å, c = 244.64 Å. | The secondary structure of this enzyme is made of beta-sheets (16 strands) and alpha-helices and a few 310-helices (34 helices), which form the two pockets of the active sites, in the C-term domain and the N-term domain, in which the substrates (lipid-phosphates and ions for the N-term domain, and epoxides for the C-term domain) can bind. The 3D dimensions of the enzyme are the following : a = 92.55 Å, b = 92.55 Å, c = 244.64 Å. | ||
The quaternary structure is called a domain-swapped dimer. The C-ter domain adopts an α/β-hydrolase fold, corresponding to its catalytic activity. Remarkably, the N-term domain has an α/β fold which is similar to HAD (Haloacid Dehalogenase) family. This N-term domain has no dehalogenase activity but a phosphatase one. | |||
The N-terminal domain has specific features that facilitate the binding of a lipid substrate. There are <scene name='Sandbox_Reserved_714/Nter_cleft-tunnel-activesite/2'>three sites</scene> that ensure the proper positioning of the substrate. First, a hydrophobic cleft of about 25 Å long is situated near the N-term core so that one of the two ends of the aliphatic substrate is near the interface between the two domains N-term and C-term (proline-rich linker). Secondly, a hydrophobic tunnel (about 14 Å long) binds the aliphatic chain of the substrate and allows the second end to be in the active site. The active site is a negatively charged pocket of about 15 Å deep, and contains a Mg<sup>2+</sup> cation necessary to the catalytic function. | The N-terminal domain has specific features that facilitate the binding of a lipid substrate. There are <scene name='Sandbox_Reserved_714/Nter_cleft-tunnel-activesite/2'>three sites</scene> that ensure the proper positioning of the substrate. First, a hydrophobic cleft of about 25 Å long is situated near the N-term core so that one of the two ends of the aliphatic substrate is near the interface between the two domains N-term and C-term (proline-rich linker). Secondly, a hydrophobic tunnel (about 14 Å long) binds the aliphatic chain of the substrate and allows the second end to be in the active site. The active site is a negatively charged pocket of about 15 Å deep, and contains a Mg<sup>2+</sup> cation necessary to the catalytic function. | ||
The aminoacids which are involved in the active site of the N-term domain are the Asp9, Asp11 and Asp185 basic aminoacids which can bind with a | The aminoacids which are involved in the active site of the N-term domain are the Asp9, Asp11 and Asp185 basic aminoacids which can bind with a Mg<sup>2+</sup> ion each, which is necessary to permit the substrate binding. These residues are characteristic of a significant part of phosphatases and phosphonatases. There is also a modified lysine, the Lys43, which has an acetyl group on its N6. | ||
The aminoacids which are involved in the active site of the C-term domain can be divided in two groups : the binding aminoacids and the catalytic aminoacids. | The aminoacids which are involved in the active site of the C-term domain can be divided in two groups : the binding aminoacids and the catalytic aminoacids. | ||
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The His239, Tyr241, Arg249 and Glu298 are involved in the binding of the substrate, the Hexaethylene Glycol (P6G). | The His239, Tyr241, Arg249 and Glu298 are involved in the binding of the substrate, the Hexaethylene Glycol (P6G). | ||
The | The Asp335, Asp496, Tyr466 and His524 are the catalytic aminoacids of the active site. Asp335 can lead nucleophilic additions, Tyr466 is a proton donor and His524 is a proton acceptor. | ||
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=== C-terminal domain === | === C-terminal domain === | ||
The C-terminal domain is called Cytosolic epoxide hydrolase 2: it catalyzes the trans-addition of water to epoxides in order to product glycols<ref>PMID:15822179</ref>. | The C-terminal domain is called Cytosolic epoxide hydrolase 2: it catalyzes the trans-addition of water to epoxides in order to product glycols <ref name="EH">PMID:15822179</ref>. | ||
The corresponding reaction equation is the following: | The corresponding reaction equation is the following: | ||
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As many enzymes, the human soluble epoxide hydrolase can be inhibited by some inhibitors, causing a loss of activity for the enzyme. | As many enzymes, the human soluble epoxide hydrolase can be inhibited by some inhibitors, causing a loss of activity for the enzyme. | ||
The sEH can be inhibited by some metal ions such as Zn<sup>2+</sup>, Cu<sup>2+</sup>, Hg<sup>2+</sup>. It is a noncompetitive inhibition: the metal ion binding to the enzyme reduces its activity without affecting directly the substrate binding, so that the V<sub>max</sub> decreases but the K<sub>m</sub> remains unchanged. It may be that those metal ions replace the Mg<sup>2+</sup> in the N-term active site of the enzyme subunits, which can result in some conformation changes, but this is only an hypothesis. | The sEH can be inhibited by some metal ions such as Zn<sup>2+</sup>, Cu<sup>2+</sup>, Hg<sup>2+</sup> <ref name="EH" />. It is a noncompetitive inhibition: the metal ion binding to the enzyme reduces its activity without affecting directly the substrate binding, so that the V<sub>max</sub> decreases but the K<sub>m</sub> remains unchanged. It may be that those metal ions replace the Mg<sup>2+</sup> in the N-term active site of the enzyme subunits, which can result in some conformation changes, but this is only an hypothesis. | ||
However, there are also some chemical inhibitors that inhibit the sEH. They are 1-3 disubstitued ureas, carbamates and amides, which are stable inhibitors for the sEH. | However, there are also some chemical inhibitors that inhibit the sEH. They are 1-3 disubstitued ureas, carbamates and amides, which are stable inhibitors for the sEH. | ||