PHOSPHORYLATION OF ELONGATION-FACTOR TU PREVENTS TERNARY COMPLEX-FORMATION

The elongation factor Tu (EF-Tu) is a member of the GTP/GDP-binding pr oteins and interacts with various partners during the elongation cycle of protein biosynthesis thereby mediating the correct binding of amin o-acylated transfer RNA (aa-tRNA) to the acceptor site (A-site) of the ribosome. After GTP hydrolysis EF-Tu is released in its GDP-bound sta te. lit vivo, EF-Tu is posttranslationally modified by phosphorylation . Here we report that the phosphorylation of EF-Tu by a ribosome assoc iated kinase activity is drastically enhanced by EF-Ts. The antibiotic kirromycin, known to block EF-Tu function, inhibits the modification. This effect is specific, since kirromycin-resistant mutants do become phosphorylated in the presence of the antibiotic. On the other hand, phosphorylated wild-type EF-Tu does not bind kirromycin. Most interest ingly, the phosphorylation of EF-Tu abolishes its ability to bind aa-t RNA. In the GTP conformation the site of modification is located at th e interface between domains 1 and 3 and is involved in a strong interd omain hydrogen bond. Introduction of a charged phosphate group at this position will change the interaction between the domains, leading to an opening of the molecule reminiscent of the GDP conformation. A mode l for the function of EF-Tu phosphorylation in protein biosynthesis is presented.