Receptor site for the 5 '-phosphate of elongator tRNAs governs substrate selection by peptidyl-tRNA hydrolase

Eubacterial peptidyl-tRNA hydrolase (PTH) recycles all N-blocked aminoacyl- tRNA molecules but initiator formyl-methionyl-tRNA(f)(Met), the acceptor he lix of which is characterized by a 1-72 mismatch. Positive selection by PTH of noninitiator tRNA molecules with a full 1-72 base pair is abolished, ho wever, upon the removal of the S'-phosphate. The tRNA 5'-phosphate plays th erefore the role of a relay between the enzyme and the status of the 1-72 b ase pair. In this study, the receptor site for the S-phosphate of elongator peptidyl-tRNAs and the position at the surface of PTH of the 3'-end of com plexed peptidyl-tRNA are identified by site-directed mutagenesis experiment s. The former site comprehends two cationic side chains (K105 and R133) whi ch are likely to clamp the phosphate. The second corresponds to a four aspa ragine cluster (N10, N21, N68, and N114). By using these two positional con straints, the acceptor arm of elongation factor Tu-bound Phe-tRNA(Phe) coul d be docked to PTH. Contacts involve the accepter and T Psi C stems. By com paring the obtained 3D model to that of EF-Tu:Phe-tRNA(Phe) crystalline com plex in which the 5'-phosphate of the ligand also lies between a K and an R side chain, we propose that, in both systems, the capacity of the 5'-phosp hate of a tRNA to reach or not a receptor site is the main identity element governing generic selection of elongator tRNAs. On the other hand, while t he 1-72 mismatch acts as an antideterminant for PTH or EF-Tu recognition, i t behaves as a positive determinant for the formylation of initiator Met-tR NA(f)(Met).