Novel Mutants of EF-Tu

Abstract

The process of translation in bacteria is familiar in its broad outlines. However precise knowledge of many of its reactions, their kinetics and the strategies underlying them is still lacking. During the elongation cycle, EF-Tu, in a ternary complex with GTP and aminoacyl-tRNA, plays a role in mediating the interaction between the aminoacyl-tRNA and the ribosome. The elongation cycle in bacteria such as E.coli operates at a rate of up to 20 amino acids per second and with an error frequency in the range of 10–^3- 10–⁴ per codon. How EF-Tu contributes to this level of efficiency is unclear. We are studying wild-type and mutant derivatives of EF-Tu in an attempt to define in more detail the elongation cycle and the role of ternary complex in translation. The cycle of elongation may be formally considered to comprise two stages; the selection of a correct aminoacyl-tRNA on a codon-programmed ribosome, and the selection of a new codon involving movement of the selected tRNA relative to the ribosome. The selection of the correct aminoacyl-tRNA species involves a reversible initial selection (I) followed by hydrolysis of GTP on the ternary complex. The low accuracy of this initial selection is increased by one or more kinetic proofreading steps (F) resulting either in the successful formation of a peptide bond or the ejection of the aminoacyl-tRNA from the ribosome (Thompson and Stone, 1977; Ruusala et al, 1982).