Biosynthesis of chlorophyll precursors in green algae purification and characterization of l-glutamate: 4,5-dioxovaleric acid aminotransferase from Chlorella fusca

Abstract

The green alga Chlorella fusca contains at least three enzymes which catalyze the transamination of 4,5-dioxovaleric acid to 5-aminolevulinic acid with several amino acids as amino donors. One of these enzymes which prefers l-glutamate as an amino donor was 60-fold enriched by column chromatography on calcium phosphate, DEAE-cellulose, Sepharose CL-4B and DEAE-Sepharose and there by purified to apparent homogeneity. The transaminase is quite stable, especially in the presence of 4,5-dioxovaleric acid and pyridoxamine phosphate, requires pyridoxal phosphate and has maximal activity at 37–40°C and pH 7.3. The enzyme has an apparent molecular weight of about 60 000 and contains 0.5 mol pyridoxal phosphate per mol. The K m values for 4,5-dioxovaleric acid and l-glutamate are 2.0 and 0.48 mM, respectively. Besides 4,5-dioxovaleric acid, glyoxylic acid is also transaminated, but the enzyme can only be activated by a combination of 4,5-dioxovaleric acid and pyridoxamine phosphate.