Oxidation of d -Amino Acids by a Particulate Enzyme from Pseudomonas aeruginosa

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Abstract
A particulate D-amino acid dehydrogenase was partially purified from cell free extracts of P. aeruginosa grown on DL-valine as the source of C and energy. A standard assay was developed which utilized 2,6-dichlorophenolindophenol as the electron acceptor. The pH optimum for enzyme activity ranged from 6. 0-8. 0 depending on the amino-acid assayed. The enzyme was most active with monoamino-monocarboxylic amino-acids and histidine. The Michaelis constant for D-phenylalanine was 1. 3 x 10-3 M D-phenylalanine. Constants could not be calculated for the other amino-acids oxidized because anomalous plots of V as a function of V/S were obtained. Spectra of enzyme preparations reduced with D-valine or sodium hydrosulf ite exhibited adsorption bands typical of the [alpha], [beta], and [gamma] bands of cytochromes as well as bleaching in the flavin region of the spectrum. When DL-valine was added to a medium with glycerol as the energy source, D-amino-acid dehydrogenase was detected after the addition of valine and was produced at a rate directly proportional to the synthesis of total protein. The enzyme was formed when D-valine, L-valine, or DL-alanine was the source of C and energy, but not when glucose, glycerol, or succinate was the energy source.