Journal: PLoS ONE
Article Title: Kinetic Characterization and Allosteric Inhibition of the Yersinia pestis 1-Deoxy-D-Xylulose 5-Phosphate Reductoisomerase (MEP Synthase)
doi: 10.1371/journal.pone.0106243
Figure Lengend Snippet: A) The MVA pathway is utilized by humans and other eukaryotes, archaebacteria, and certain eubacteria to produce IPP and DMAPP, the building blocks of isoprenoids. The pathway is initiated by the enzymatic condensation of 3 molecules of acetyl-CoA (1) to form 3-hydroxy-3-methylglutaryl CoA (HMG-CoA) (3), which is then reduced to MVA by HMG-CoA reductase (4) Subsequent phosphorylation and decarboxylation yield IPP (7) which is converted to DMAPP (8) by an isomerase . B) The MEP pathway is used by higher plants, the plastids of algae, apicomplexan protozoa, and many eubacteria, including numerous human pathogens. Pyruvate (9) is condensed with glyceraldehyde 3-phosphate (10) to yield 1-deoxy-D-xylulose 5-phosphate (DXP; (11)) , a branch point intermediate with a role in E. coli vitamin B1 and B6 biosynthesis as well as isoprene biosynthesis. In the first committed step of the E. coli MEP pathway, 1-Deoxy-D-xylulose 5-phosphate reductoisomerase (also called MEP synthase, Dxr or IspC) catalyzes the reduction and rearrangement of 11 to yield MEP (12) . CDP-ME synthase then converts MEP into 4-(cytidine 5′-diphospho)-2-C-methyl-D-erythritol (CDP-ME; (13)). CDP-ME kinase phosphorylates CDP-ME, which is subsequently cyclized (coupled with the loss of CMP) by cMEPP synthase to yield 2-C-methyl-D-erythritol 2,4-cyclodiphosphate (15) . A reductive ring opening of 15 produces 1-hydroxy-2-methyl-2-butenyl diphosphate (HMBPP; (16)) , which is then reduced to both IPP and DMAPP in a ∼5:1 ratio . C) The reaction catalyzed by MEP synthase. The intermediate 2-C-methyl-D-erythrose 4-phosphate (18), produced by isomerization via cleavage of the bond between C3 and C4 and formation of a new bond between C2 and C4 , is subsequently reduced to yield MEP (12).
Article Snippet: To determine the apparent K M for 1 deoxy-D-xylulose 5-phosphate (DXP), 120 µL assay solutions contained 100 mM Tris pH 7.8, 25 mM MgCl 2 , 150 µM NADPH, 0.89 µM MEP synthase, and variable concentrations of DXP (Echelon Biosciences, Salt Lake City, UT).
Techniques: Produced