1yqn
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(New page: 200px<br /><applet load="1yqn" size="450" color="white" frame="true" align="right" spinBox="true" caption="1yqn, resolution 3.11Å" /> '''E. coli ispF double ...)
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Revision as of 04:52, 21 November 2007
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E. coli ispF double mutant
Overview
The essential enzyme 2C-methyl-D-erythritol-2,4-cyclodiphosphate (MECP), synthase, found in most eubacteria and the apicomplexan parasites, participates in isoprenoid-precursor biosynthesis and is a validated, target for the development of broad-spectrum antimicrobial drugs. The, structure and mechanism of the enzyme have been elucidated and the recent, exciting finding that the enzyme actually binds diphosphate-containing, isoprenoids at the interface formed by the three subunits that constitute, the active protein suggests the possibility of feedback regulation of MECP, synthase. To investigate such a possibility, a form of the enzyme was, sought that did not bind these ligands but which would retain the, quaternary structure necessary to create the active site. Two amino acids, Arg142 and Glu144, in Escherichia coli MECP synthase were identified as, contributing to ligand binding. Glu144 interacts directly with Arg142 and, positions the basic residue to form two hydrogen bonds with the terminal, phosphate group of the isoprenoid diphosphate ligand. This association, occurs at the trimer interface and three of these arginines interact with, the ligand phosphate group. A dual mutation was designed (Arg142 to, methionine and Glu144 to leucine) to disrupt the electrostatic attractions, between the enzyme and the phosphate group to investigate whether an, enzyme without isoprenoid diphosphate could be obtained. A low-resolution, crystal structure of the mutated MECP synthase Met142/Leu144 revealed that, geranyl diphosphate was retained despite the removal of six hydrogen bonds, normally formed with the enzyme. This indicates that these two hydrophilic, residues on the surface of the enzyme are not major determinants of, isoprenoid binding at the trimer interface but rather that hydrophobic, interactions between the hydrocarbon tail and the core of the enzyme, trimer dominate ligand binding.
About this Structure
1YQN is a Single protein structure of sequence from Escherichia coli with ZN, MN, CDP and GPP as ligands. Active as 2-C-methyl-D-erythritol 2,4-cyclodiphosphate synthase, with EC number 4.6.1.12 Full crystallographic information is available from OCA.
Reference
A double mutation of Escherichia coli2C-methyl-D-erythritol-2,4-cyclodiphosphate synthase disrupts six hydrogen bonds with, yet fails to prevent binding of, an isoprenoid diphosphate., Sgraja T, Kemp LE, Ramsden N, Hunter WN, Acta Crystallograph Sect F Struct Biol Cryst Commun. 2005 Jul 1;61(Pt, 7):625-9. Epub 2005 Jun 30. PMID:16511114
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