Anacystis nidulans mutants resistant to aromatic amino acid analogues.
Three classes of mutants of Anacystis nidulans were selected on the basis of resistance to fluorophenylalanine and 2-amino-3-phenylbutanoic acid. The most frequent type exhibited DAHP synthetase (7-phospho-2-keto-3-deoxy-D-arabino-heptonate-D-erythrose-4-phosphate-lyase [pyruvate phosphorylating], EC 4.1.2.15) activity identical to that of the parental strain. The second type was characterized by extremely low levels of the activity. The third type had a DAHP synthetase showing decreased sensitivity to inhibition by L-tyrosine. The enzyme was purified 140-fold from wild-type and feedback-insensitive strains, and the kinetics of the reaction was examined. The activity of the wild-type enzyme was inhibited 75% in the presence of 2.0 X 10-3 M tyrosine, and the altered enzyme was inhibited 10%. The following apparent constants were obtained from kinetic studies with partially purified wild-type enzyme: S0.5 for D-erythrose-4-phophate equal to 7.1 X 10-4 M; S0.5 for phosphoenolpyruvate equal to 1.4 X 10-4 M. Inhibition by tyrosine was mixed with respect to binding of both D-erythrose-4-phosphate and phosphoenolpyruvate. In addition, tyrosine promoted cooperative interactions in the binding of phosphoenolpyruvate. For the altered enzyme the following apparent constants were obtained: S0.5 for D-erythrose-4-phosphate equal to 7.1 X 10-4 M; S0.5 for phosphoenolpyruvate equal to 2.9 X 10-4 M. Inhibition by tyrosine was mixed with respect to D-erythrose-4-phosphate and competitive with respect to phosphoenolpyruvate. Tyrosine did not promote cooperative effects in the binding of phosphoenolpyruvate to the altered enzyme.
D000612:4-Aminobutyrate Transaminase / Q000494:pharmacology; D000446:Aldehyde-Lyases / Q000302:isolation & purification / Q000378:metabolism; D000613:Aminobutyrates; D000645:Ammonium Sulfate; D002474:Cell-Free System; D005591:Chemical Fractionation; D002850:Chromatography, Gel; D000458:Cyanobacteria / Q000187:drug effects* / Q000201:enzymology; D004352:Drug Resistance, Microbial; D005246:Feedback; D006539:Heptoses; D006882:Hydroxyapatites; D007700:Kinetics; D009154:Mutation; D010649:Phenylalanine / Q000031:analogs & derivatives*; D010654:Phenylbutyrates / Q000494:pharmacology*; D010728:Phosphoenolpyruvate / Q000378:metabolism; D013403:Sugar Phosphates / Q000378:metabolism; D013780:Tetroses; D014443:Tyrosine / Q000494:pharmacology; D010135:p-Fluorophenylalanine / Q000494:pharmacology
D000613:Aminobutyrates; D006539:Heptoses; D006882:Hydroxyapatites; D010654:Phenylbutyrates; D013403:Sugar Phosphates; D013780:Tetroses; C010535:2-amino-3-phenylbutanoic acid; D014443:Tyrosine; D010649:Phenylalanine; D010135:p-Fluorophenylalanine; D010728:Phosphoenolpyruvate; D000612:4-Aminobutyrate Transaminase; D000446:Aldehyde-Lyases; D000645:Ammonium Sulfate