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