Rodnina, MarinaMarinaRodninaPape, TillmannTillmannPapeFricke, R.R.FrickeKuhn, L .L .KuhnWintermeyer, WolfgangWolfgangWintermeyer2017-09-072017-09-071996https://resolver.sub.uni-goettingen.de/purl?gro-2/2312The first step in the sequence of interactions between the ribosome and the complex of elongation factor Tu (EF-Tu), GTP, and aminoacyl-tRNA, which eventually leads to A site-bound aminoacyl-tRNA, is the codon-independent formation of an initial complex. We have characterized the initial binding and the resulting complex by time resolved (stopped-flow) and steady-state fluorescence measurements using several fluorescent tRNA derivatives. The complex is labile, with rate constants of 6 x 10(7) M(-1) s(-1) and 24 s(-1) (20 degrees C, 10 mM Mg2+) for binding and dissociation, respectively. Both thermodynamic and activation parameters of initial binding were determined, and five Mg2+ ions were estimated to participate in the interaction. While a cognate ternary complex proceeds from initial binding through codon recognition to rapid GTP hydrolysis, the rate constant of GTP hydrolysis in the non-cognate complex is 4 orders of magnitude lower, despite the rapid formation of the initial complex in both cases, Hence, the ribosome-induced GTP hydrolysis by EF-Tu is strongly affected by the presence of the tRNA. This suggests that codon-anticodon recognition, which takes place after the formation of the initial binding complex, provides a specific signal that triggers fast GTP hydrolysis by EF-Tu on the ribosome.journal_article8557669A1996TP889000123144663