Purification and Properties of Dissociable Chorismate Mutase from Brevibacterium flavum
1980 年 88 巻 1 号 p. 167-176
詳細
1980 年 88 巻 1 号 p. 167-176
Component B of chorismate mutase of Brevibacterium flavum, the first enzyme specific for phenylalanine and tyrosine biosynthesis, was purified to near homogeneity. The molecular weights of component B and its subunit were estimated to be 25, 000 and 13, 500, respectively. Component A (previously purified) or B alone did not show any chorismate mutase activity but together they showed activity. The enzyme activity was not proportional to the amount of the enzyme. The optimum pH of the reaction was 8.0. Double-reciprocal plots of the reaction rate against chorismate concentration curved upwards. S0.5 and Hill coefficient values were estimated to be 5.5mM and 3.1, respectively. The chorismate mutase component A and 3-deoxy-D-arabino-heptulosonate 7-phosphate synthetase activities of component A were labile and were stabilized by tryptophan, dithiothreitol or cobalt ions. Phenylalanine and tyrosine inhibited the enzyme activity partially and competitively. The simultaneous presence of phenylalanine and tyrosine caused cumulative inhibition at saturated concentrations. The concentrations of phenylalanine, tyrosine, and phenylalanine plus tyrosine (each) giving 50% inhibition under the standard conditions were 0.0041, 0.095, and 0.0023mM, respectively. Tryptophan activated the enzyme about 6-fold. The concentration giving the half-maximum activation was 0.0023mM. Furthermore, tryptophan overcame the inhibition caused by phenylalanine and tyrosine. The tryptophan activation affected only S0.5, not the maximum velocity or the Hill coefficient. β-2-Thienylalanine, m-fluorophenylalanine, α-methylphenylalanine, and phenylalanine hydroxamate inhibited the enzyme in the same way as phenylalanine, while tyrosine hydroxamate and α-methyltyrosine inhibited it in the same way as tyrosine. 4-Methyltryptophan, 5-fluorotryptophan, 6-fluorotryptophan, tryptophan hydroxamate, and α-methyltryptophan activated the enzyme in the same way as tryptophan.
