Abstract
This chapter discusses the methods of preparation of D -Serine Dehydrase ( Escherichia coli ). The dehydration of D -serine, catalyzed by D -serine dehydrase, continues until all of the amino acid is converted to pyruvate and ammonia. The most convenient and broadly applicable assay is based on the colorimetric determination of pyruvate. Pyruvate can also be determined by measuring the rate of Nicotinamide Adenine Dinucleotide - Hydrogen (NADH) oxidation in the presence of lactate dehydrogenase. Another sensitive assay of the enzyme is based on the determination of the ammonia produced, but this procedure is more time consuming than the pyruvate determination. One unit of dehydrase is the amount of enzyme that catalyzes the formation of 1 micromole of pyruvate (or ammonia)/minute. Specific activity is expressed as units per milligram of protein. The spectrophotometric assay cannot be used with impure enzyme preparations. The interfering enzyme is an NADH oxidase that is eventually separated from D -serine dehydrase by calcium phosphate gel-cellulose chromatography.
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