Essential arginine residues in tryptophanase from Escherichia coli.
Tryptophanase from Escherichia coli B/1t7-A is inactivated by the arginine-specific reagent, phenylglyoxal, in potassium phosphate buffer at pH 7.8 AND 25 degrees. Apo- and holoenzyme are inactivated at the same rate, and inactivation of both is correlated with modification of 2 arginine residues/tryptophanase monomer. Substrate analogs having a carboxyl group protect the holoenzyme against both inactivation and arginine modification but have no effect on the inactivation or modification of the apoenzyme. Phenylglyoxal-modified apotryptophanase retains the capacity to bind the coenzyme, pyridoxal-P, but the spectrum of this reconstituted species differs from that of native holotryptophanase. Neither this reconstituted species nor the phenyglyoxal-modified holoenzyme shows the 500 nm absorption characteristic of the native enzyme when substrates are added. These results demonstrate a requirement for specific arginine residues for substrate binding and are discussed in the context of the known conformational and spectal forms of tryptophanase with regard to a possible role for arginine residues in formation of a catalytically effective enzyme-pyridoxal-P complex.
D001120:Arginine; D001665:Binding Sites; D004926:Escherichia coli / Q000201:enzymology*; D007700:Kinetics; D008190:Lyases / Q000378:metabolism; D011732:Pyridoxal Phosphate / Q000494:pharmacology; D013053:Spectrophotometry; D013056:Spectrophotometry, Ultraviolet; D013379:Substrate Specificity; D014368:Tryptophanase / Q000378:metabolism
D011732:Pyridoxal Phosphate; D001120:Arginine; D008190:Lyases; D014368:Tryptophanase