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

Abstract

A particulate d -amino acid dehydrogenase has been partially purified from cell free extracts of Pseudomonas aeruginosa grown on dl -valine as the source of carbon and energy. A standard assay was developed which utilized 2,6-dichlorophenol-indophenol as the electron acceptor. The p H optimum for enzyme activity ranged from 6.0 to 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 found to be 1.3 × 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 hydrosulfite exhibited adsorption bands typical of the α, β, and γ 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 carbon and energy, but not when glucose, glycerol, or succinate was the energy source.