Single turnover kinetic studies of guanosine triphosphate hydrolysis and peptide formation in the elongation factor Tu-dependent binding of aminoacyl-tRNA to Escherichia coli ribosomes.
The rates of GTP hydrolysis and peptide formation during the reaction of Phe-tRNA . elongation factor Tu . GTP complex with acetyl-Phe-tRNA polyuridylate-programmed ribosomes have been measured. The GTPase reaction is second-order up to reactant concentrations of 0.2 microM and has a rate constant of 5 X 10(6) M-1 s-1 at 5 degrees C and 5 mM Mg2+, pH 7.2. The formation of peptide shows a lag phase and has a rate constant of 0.4 S-1 under these conditions. The results of a series of experiments between 5 degrees C and 25 degrees C show that GTP hydrolysis and peptide formation have Arrhenius activation energies of 13.1 and 15.3 kcal mol-1, respectively. The results indicate that these reactions proceed in vitro at rates comparable to those observed for protein biosynthesis in vivo, and that peptide bond formation occurs after GTP hydrolysis.
D001426:Bacterial Proteins / Q000378:metabolism*; D004926:Escherichia coli / Q000378:metabolism*; D006160:Guanosine Triphosphate / Q000378:metabolism*; D006868:Hydrolysis; D007700:Kinetics; D010441:Peptide Chain Elongation, Translational; D010444:Peptide Elongation Factor Tu; D011072:Poly U; D012346:RNA, Transfer, Amino Acyl / Q000378:metabolism*; D012270:Ribosomes / Q000378:metabolism*
D001426:Bacterial Proteins; D012346:RNA, Transfer, Amino Acyl; D011072:Poly U; D006160:Guanosine Triphosphate; D010444:Peptide Elongation Factor Tu