TY - JOUR
T1 - Competence of Thiamin Diphosphate-Dependent Enzymes with 2′-Methoxythiamin Diphosphate Derived from Bacimethrin, a Naturally Occurring Thiamin Anti-vitamin
AU - Nemeria, Natalia S.
AU - Shome, Brateen
AU - Decolli, Alicia A.
AU - Heflin, Kathryn
AU - Begley, Tadhg P.
AU - Meyers, Caren Freel
AU - Jordan, Frank
N1 - Funding Information:
This work was supported, in whole or in part, by National Institutes of Health (NIH) Grant R15GM116077 and National Science Foundation Grant CHE-1402675 (to F.J.), NIH Grant GM084998 (to C.F.M. and F.J.), NIH Grant T32M008763 (to A.A.D.), NIH Grant T32M08018901 (to K.H.), NIH Grant DK44083 and Robert A. Welch Foundation Grant A-0034 (to T.P.B.). The Rutgers group is grateful to Prof. Mulchand Patel for supplying the plasmids with the human PDHc components.
Publisher Copyright:
© 2016 American Chemical Society.
PY - 2016/3/1
Y1 - 2016/3/1
N2 - 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.
AB - 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.
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U2 - 10.1021/acs.biochem.5b01300
DO - 10.1021/acs.biochem.5b01300
M3 - Article
C2 - 26813608
AN - SCOPUS:84959440360
SN - 0006-2960
VL - 55
SP - 1135
EP - 1148
JO - Biochemistry
JF - Biochemistry
IS - 7
ER -