Kinetic studies of Escherichia coli elongation factor Tu-guanosine 5'-triphosphate-aminoacyl-tRNA complexes.
A new method for measuring the dissociation rate of the Escherichia coli elongation factor Tu-GTP--aminoacyl-tRNA complex has been developed and applied to the determination of the dissociation rates of ternary complexes formed between E. coli EF-Tu-GTP and a set of E. coli aminoacyl-tRNAs. The set of aminoacyl-tRNAs includes at least one tRNA coding for each of the 20 amino acids as well as purified isoacceptor tRNA species for arginine, glycine, leucine, lysine, and tyrosine. The results reveal that the dissociation rates vary for each ternary complex. Tu-GTP-Gln-tRNA dissociates the slowest and Tu-GTP-Val-tRNA the fastest of all noninitiator ternary complexes at 4 degrees C, pH 7.4. The equilibrium dissociation constant for Tu-GTP-Thr-tRNA has been determined to be 1.3 (0.4) X 10(-9) M under identical reaction conditions, and the absolute value of the equilibrium dissociation constant has been calculated for 28 ternary complexes from the relative equilibrium dissociation constant ratios previously measured [Louie, A., Ribeiro, N. S., Reid, B. R., & Jurnak, F. (1984) J. Biol. Chem. 259, 5010-5016]. The association rate of each ternary complex has been estimated from the ratio of the dissociation rate relative to the equilibrium dissociation constant. Tu-GTP-His-tRNA associates the fastest and Tu-GTP-Leu-tRNA1Leu the slowest. By inclusion of Tu-GTP-Met-tRNAfMet in the studies, evidence has been obtained that suggests that the initiator ternary complex does not function in the elongation cycle because the dissociation rate of the complex is very fast.
D000596:Amino Acids / Q000378:metabolism; D000604:Amino Acyl-tRNA Synthetases / Q000378:metabolism*; D002250:Carbon Radioisotopes; D004926:Escherichia coli / Q000378:metabolism*; D006160:Guanosine Triphosphate / Q000378:metabolism*; D007700:Kinetics; D010444:Peptide Elongation Factor Tu / Q000378:metabolism*; D012346:RNA, Transfer, Amino Acyl / Q000378:metabolism*
D000596:Amino Acids; D002250:Carbon Radioisotopes; D012346:RNA, Transfer, Amino Acyl; D006160:Guanosine Triphosphate; D010444:Peptide Elongation Factor Tu; D000604:Amino Acyl-tRNA Synthetases