Stereo-specificity for pro-(R) hydrogen of NAD(P)H during enzyme-catalyzed hydride transfer to CL-20

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DOIResolve DOI: http://doi.org/10.1016/j.bbrc.2005.09.166
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TypeArticle
Journal titleBiochemical and Biophysical Research Communications
Volume337
Issue4
Pages10801083; # of pages: 4
AbstractA dehydrogenase from Clostridium sp. EDB2 and a diaphorase from Clostridium kluyveri were reacted with CL-20 to gain insights into the enzyme-catalyzed hydride transfer to CL-20, and the enzyme's stereo-specificity for either pro-R or pro-S hydrogens of NAD(P)H. Both enzymes biotransformed CL-20 at rates of 18.5 and 24 nmol/h/mg protein, using NADH and NADPH as hydride-source, respectively, to produce a N-denitrohydrogenated product with a molecular weight of 393 Da. In enzyme kinetics studies using reduced deuterated pyridine nucleotides, we found a kinetic deuterium isotopic effect of 2-fold on CL-20 biotransformation rate using dehydrogenase enzyme against (R)NADD as a hydride-source compared to either (S)NADD or NADH. Whereas, in case of diaphorase, the kinetic deuterium isotopic effect of about 1.5-fold was observed on CL-20 biotransformation rate using (R)NADPD as hydride-source. In a comparative study with LC-MS, using deuterated and non-deuterated NAD(P)H, we found a positive mass-shift of 1 Da in the N-denitrohydrogenated product suggesting the involvement of a deuteride (D-) transfer from NAD(P)D. The present study thus revealed that both dehydrogenase and diaphorase enzymes from the two Clostridium species catalyzed a hydride transfer to CL-20 and showed stereo-specificity for pro-R hydrogen of NAD(P)H.
Publication date
LanguageEnglish
AffiliationNRC Biotechnology Research Institute; National Research Council Canada
Peer reviewedNo
NRC number47245
NPARC number3540086
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Record identifier59a735d0-38a3-4713-944b-777d3b26c9f0
Record created2009-03-01
Record modified2016-05-09
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