4-Aminobutyrate:2-oxoglutarate aminotransferase of Streptomyces griseus: purification and properties.
4-Aminobutyrate: 2-oxoglutarate aminotransferase of Streptomyces griseus was purified to homogeneity on disc electrophoresis. The relative molecular mass of the enzyme was found to be 100 000 +/- 10 000 by a gel filtration method. The enzyme consists of two subunits identical in molecular mass (Mr 50 000 +/- 1000). The transaminase is composed of 486 amino acids/subunit containing 10 and 12 residues of half-cystine and methionine respectively. The NH2-terminal amino acid sequence of the enzyme was determined to be Thr-Ala-Phe-Pro-Gln. The enzyme exhibits absorption maxima at 278 nm, 340 nm and 415 nm with a molar absorption coefficient of 104 000, 11 400 and 7280 M-1 cm-1 respectively. The pyridoxal 5'-phosphate content was calculated to be 2 mol/mol enzyme. The enzyme has a maximum activity in the pH range of 7.5-8.5 and at 50 degrees C. The enzyme is stable at pH 6.0-10.0 and at temperatures up to 50 degrees C. Pyridoxal 5'-phosphate protects the enzyme from thermal inactivation. The enzyme catalyzes the transamination of omega-amino acids with 2-oxoglutarate; 4-aminobutyrate is the best amino donor. The Michaelis constants are 3.3 mM for 4-aminobutyrate and 8.3 mM for 2-oxoglutarate. Low activity was observed with beta-alanine. In addition to omega-amino acids the enzyme catalyzes transamination with ornithine and lysine; in both cases the D isomer is preferred. Carbonyl reagents and sulfhydryl reagents inhibit the enzyme activity. Chelating agents, non-substrate L and D-2-amino acids, and metal ions except cupric ion showed no effect on the enzyme activity.
D000612:4-Aminobutyrate Transaminase / Q000302:isolation & purification*; D000595:Amino Acid Sequence; D000596:Amino Acids / Q000032:analysis; D055598:Chemical Phenomena; D002621:Chemistry; D006863:Hydrogen-Ion Concentration; D013053:Spectrophotometry; D013305:Streptomyces griseus / Q000201:enzymology*; D013379:Substrate Specificity; D013696:Temperature
D000596:Amino Acids; D000612:4-Aminobutyrate Transaminase