RRC ID 53749
Author Nemeria NS, Shome B, DeColli AA, Heflin K, Begley TP, Meyers CF, Jordan F.
Title Competence of Thiamin Diphosphate-Dependent Enzymes with 2'-Methoxythiamin Diphosphate Derived from Bacimethrin, a Naturally Occurring Thiamin Anti-vitamin.
Journal Biochemistry
Abstract Bacimethrin (4-amino-5-hydroxymethyl-2-methoxypyrimidine), a natural product isolated from some bacteria, has been implicated as an inhibitor of bacterial and yeast growth, as well as in inhibition of thiamin biosynthesis. Given that thiamin biosynthetic enzymes could convert bacimethrin to 2'-methoxythiamin diphosphate (MeOThDP), it is important to evaluate the effect of this coenzyme analogue on thiamin diphosphate (ThDP)-dependent enzymes. The potential functions of MeOThDP were explored on five ThDP-dependent enzymes: the human and Escherichia coli pyruvate dehydrogenase complexes (PDHc-h and PDHc-ec, respectively), the E. coli 1-deoxy-D-xylulose 5-phosphate synthase (DXPS), and the human and E. coli 2-oxoglutarate dehydrogenase complexes (OGDHc-h and OGDHc-ec, respectively). Using several mechanistic tools (fluorescence, circular dichroism, kinetics, and mass spectrometry), it was demonstrated that MeOThDP binds in the active centers of ThDP-dependent enzymes, however, with a binding mode different from that of ThDP. While modest activities resulted from addition of MeOThDP to E. coli PDHc (6-11%) and DXPS (9-14%), suggesting that MeOThDP-derived covalent intermediates are converted to the corresponding products (albeit with rates slower than that with ThDP), remarkably strong activity (up to 75%) resulted upon addition of the coenzyme analogue to PDHc-h. With PDHc-ec and PDHc-h, the coenzyme analogue could support all reactions, including communication between components in the complex. No functional substitution of MeOThDP for ThDP was in evidence with either OGDH-h or OGDH-ec, shown to be due to tight binding of ThDP.
Volume 55(7)
Pages 1135-48
Published 2016-2-23
DOI 10.1021/acs.biochem.5b01300
PMID 26813608
PMC PMC4852132
MeSH Amino Acid Substitution Apoenzymes / chemistry Apoenzymes / genetics Apoenzymes / metabolism Binding, Competitive Biocatalysis Catalytic Domain Escherichia coli Proteins / chemistry Escherichia coli Proteins / genetics Escherichia coli Proteins / metabolism* Humans Ketoglutarate Dehydrogenase Complex / chemistry Ketoglutarate Dehydrogenase Complex / genetics Ketoglutarate Dehydrogenase Complex / metabolism Models, Molecular* Mutation Protein Subunits / chemistry Protein Subunits / genetics Protein Subunits / metabolism Pyrimidines / chemistry Pyruvate Dehydrogenase Complex / chemistry Pyruvate Dehydrogenase Complex / genetics Pyruvate Dehydrogenase Complex / metabolism* Recombinant Proteins / chemistry Recombinant Proteins / metabolism Substrate Specificity Thiamine Pyrophosphate / analogs & derivatives* Thiamine Pyrophosphate / metabolism* Transferases / chemistry Transferases / metabolism*
IF 2.865
Times Cited 8
Prokaryotes E. coli