Altered alpha subunits in phenylalanyl-tRNA synthetases from p-fluorophenylalanine-resistant strains of Escherichis coli.
Three different phenylalanyl-tRNA synthetases have been purified to near homogeneity, one from a wild-type strain of Escherichia coli and the others from two independently isolated p-fluorophenyalanine-resistant strains. The mutant enzymes were not able to use p-fluorophenylalanine as a substrate for activation and attachment to tRNA. They proved to be indistinguishable from the wild-type enzyme by several electrophoretic and immunological criteria. The alpha and beta subunits of all three enzymes have been prepared by a method described in this paper. The isolated subunits per se did not reveal any significant enzyme activity, but combined they were able to form active phenylalanyl tRNA synthetase after a defined reconstitution process. Mixed reconstitution experiments between wild-type and mutant subunits indicate that the mutant alpha subunit is responsible for p-fluorophenylalanine resistance and therefore seems to carry the phenylalanine-binding site or to participate in its formation.
D000604:Amino Acyl-tRNA Synthetases / Q000378:metabolism*; D004352:Drug Resistance, Microbial; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000201:enzymology*; D010446:Peptide Fragments / Q000378:metabolism; D010649:Phenylalanine / Q000031:analogs & derivatives*; D010652:Phenylalanine-tRNA Ligase / Q000276:immunology / Q000302:isolation & purification / Q000378:metabolism*; D010135:p-Fluorophenylalanine / Q000378:metabolism*
D010446:Peptide Fragments; D010649:Phenylalanine; D010135:p-Fluorophenylalanine; D000604:Amino Acyl-tRNA Synthetases; D010652:Phenylalanine-tRNA Ligase