2gvw
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(New page: 200px<br /><applet load="2gvw" size="450" color="white" frame="true" align="right" spinBox="true" caption="2gvw, resolution 1.86Å" /> '''Structure of diisopr...)
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Revision as of 09:18, 21 November 2007
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Structure of diisopropyl fluorophosphatase (DFPase) holoenzyme (RT)
Overview
A wide range of organophosphorus nerve agents, including Soman, Sarin, and, Tabun is efficiently hydrolyzed by the phosphotriesterase enzyme, diisopropyl fluorophosphatase (DFPase) from Loligo vulgaris. To date, the, lack of available inhibitors of DFPase has limited studies on its, mechanism. The de novo design, synthesis, and characterization of, substrate analogues acting as competitive inhibitors of DFPase are, reported. The 1.73 A crystal structure of, O,O-dicyclopentylphosphoroamidate (DcPPA) bound to DFPase shows a direct, coordination of the phosphoryl oxygen by the catalytic calcium ion. The, binding mode of this substrate analogue suggests a crucial role for, electrostatics in the orientation of the ligand in the active site. This, interpretation is further supported by the crystal structures of double, mutants D229N/N120D and D229N/N175D, designed to reorient the, electrostatic environment around the catalytic calcium. The structures, show no differences in their calcium coordinating environment, although, they are enzymatically inactive. Additional double mutants E21Q/N120D and, E21Q/N175D are also inactive. On the basis of these crystal structures and, kinetic and mutagenesis data as well as isotope labeling we propose a new, mechanism for DFPase activity. Calcium coordinating residue D229, in, concert with direct substrate activation by the metal ion, renders the, phosphorus atom of the substrate susceptible for attack of water, through, generation of a phosphoenzyme intermediate. Our proposed mechanism may be, applicable to the structurally related enzyme paraoxonase (PON), a, component of high-density lipoprotein (HDL).
About this Structure
2GVW is a Single protein structure of sequence from Loligo vulgaris with CA as ligand. Active as Diisopropyl-fluorophosphatase, with EC number 3.1.8.2 Full crystallographic information is available from OCA.
Reference
Binding of a designed substrate analogue to diisopropyl fluorophosphatase: implications for the phosphotriesterase mechanism., Blum MM, Lohr F, Richardt A, Ruterjans H, Chen JC, J Am Chem Soc. 2006 Oct 4;128(39):12750-7. PMID:17002369
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