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Pure Appl. Chem. Vol. 72, No. 3, pp. 373-384 (2000)

Use of inhibitors to study reactions catalyzed by enzymes requiring pyridoxal phosphate as coenzyme*

Benjamin Adams, B. Svante Axelsson, Kenneth J. M. Beresford, Nicola J. Church, Philip A. Spencer, Sheena M. Whyte, and Douglas W. Young**

Sussex Centre for Biomolecular Design and Drug Development, University of Sussex, Falmer, Brighton, BN1 9QJ, UK

Abstract: The stereochemistry of a variety of pyridoxal phosphate-mediated enzymic reactions has been studied using enzyme inhibitors that are stereospecifically labeled in the b-position with deuterium. A versatile synthesis has been developed to prepare a wide variety of stereospecifically labeled D- and L-amino acids and inhibitors. Investigation of the "turnover" of b-chloro-D-alanine and D- and L-serine-O-sulfate by D-amino acid aminotransferase and L-aspartate aminotransferase respectively has shown that reaction within the active site of the former enzyme occurs with retention of stereochemistry. Although L-aspartate aminotransferase is an enzyme of the a-family, when it was incubated with b-chloro-L-alanine in the presence of 2-mercaptoethanol, b-substitution occurred. This was shown to involve retention of stereochemistry, an outcome typical of reactions catalyzed by enzymes of the b-family that have little or no homology with enzymes of the a-family. Formation of the "Schnackerz intermediate" has been studied as has the D-amino acid oxidase catalyzed reaction of the naturally occurring inhibitor D-propargylglycine.

*Lecture presented at the 5^th International IUPAC Symposium on Bioorganic Chemistry (ISBOC-5), Pune, India, 30 January - 4 February, 2000.
**Corresponding author

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