NMR identification of an acyl-adenylate intermediate in the aryl-aldehyde oxidoreductase catalyzed reaction.
A new one-pot synthesis was designed to prepare benzoyl-AMP under anhydrous conditions in N,N-dimethylformamide. Reaction of benzoic acid with N,N'-carbonyldiimidazole and subsequently with 5'-adenosyl monophosphate gave the mixed anhydride in 76% isolated yield. The structure of benzoyl-AMP was confirmed by mass spectroscopy and 1H-, 31P-, and 13C-NMR. The purity of the preparation was greater than 98% as indicated by 31P- and 13C-NMR. Purified aryl-aldehyde oxidoreductase was incubated in NMR tubes together with either carboxy-13C-benzoyl-AMP or carboxy-13C-benzoic acid to demonstrate that benzoyl-AMP is an active intermediate during the enzymatic reduction of benzoic acid to benzaldehyde.
Li
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Division of Medicinal and Natural Products Chemistry, and Center for Biocatalysis and Bioprocessing, College of Pharmacy, University of Iowa, Iowa City, Iowa 52242, USA.
Rosazza
J P
JP
D000249:Adenosine Monophosphate / Q000031:analogs & derivatives* / Q000138:chemical synthesis / Q000378:metabolism; D000445:Aldehyde Oxidoreductases / Q000378:metabolism*; D001565:Benzoates / Q000138:chemical synthesis / Q000378:metabolism*; D019817:Benzoic Acid; D002384:Catalysis; D002855:Chromatography, Thin Layer; D007202:Indicators and Reagents; D009682:Magnetic Resonance Spectroscopy; D009615:Nocardia / Q000201:enzymology; D010084:Oxidation-Reduction
D001565:Benzoates; D007202:Indicators and Reagents; C116594:benzoyl-adenylate; D000249:Adenosine Monophosphate; D019817:Benzoic Acid; D000445:Aldehyde Oxidoreductases; C116588:aryl-aldehyde oxidoreductase