Neither arginine nor histidine can carry out the function of lysine-295 in the ATP-binding site of p60src.
All 15 protein kinases whose amino acid sequence is known contain a lysine residue at a position homologous to that of lysine-295 in p60src, the transforming protein of Rous sarcoma virus. The ATP analog p-fluorosulfonyl 5'-benzoyl adenosine inactivates both p60src and the catalytic subunit of the cyclic AMP-dependent protein kinase by modification of this lysine. We used oligonucleotide-directed mutagenesis to examine the possible functions of this residue. Lysine-295 in p60src was replaced with a glutamic acid, an arginine, or a histidine residue, and mutant p60src proteins were characterized in chicken cells infected by mutant viruses. None of these three mutant p60src proteins had tyrosine protein kinase activity in vitro, and none induced morphological transformation of infected cells. Since neither a histidine nor an arginine residue can replace the function of lysine-295, we suggest that it carries out the specialized function of proton transfer in the phosphotransferase reaction. All three mutant viruses underwent reversion to wild type during passage in tissue culture. Because the rate with which this occurred differed significantly among the mutants, reversion appears to have resulted from errors in transcription, rather than from recombination with the cellular src gene.
D000595:Amino Acid Sequence; D000818:Animals; D001120:Arginine; D001358:Avian Sarcoma Viruses / Q000201:enzymology / Q000235:genetics*; D001483:Base Sequence; D002642:Chick Embryo; D005347:Fibroblasts / Q000201:enzymology; D006639:Histidine; D008239:Lysine; D009154:Mutation; D015688:Oncogene Protein pp60(v-src); D011494:Protein Kinases / Q000235:genetics* / Q000378:metabolism; D012194:RNA-Directed DNA Polymerase / Q000378:metabolism; D012191:Retroviridae Proteins / Q000235:genetics* / Q000378:metabolism
D012191:Retroviridae Proteins; D006639:Histidine; D001120:Arginine; D011494:Protein Kinases; D015688:Oncogene Protein pp60(v-src); D012194:RNA-Directed DNA Polymerase; D008239:Lysine