The crystal structure of ribosomal chaperone trigger factor from Vibrio cholerae

Abstract

Trigger factor is a molecular chaperone that is present in all species of eubacteria. It binds to the ribosomal 50S subunit near the translation exit tunnel and is thought to be the first protein to interact with nascent polypeptides emerging from the ribosome. The chaperone has a peptidyl-prolyl cis-trans isomerase (PPIase) activity that catalyzes the rate-limiting proline isomerization in the protein-folding process. We have determined the crystal structure of nearly full-length trigger factor from Vibrio cholerae by x-ray crystallography at 2.5-Å resolution. The structure is composed of two trigger-factor molecules related by a noncrystallographic twofold symmetry axis. The monomer has an elongated shape and is folded into three domains: an N-terminal domain I that binds to the ribosome, a central domain II that contains PPIase activity, and a C-terminal domain III. The active site of the PPIase domain is occupied by a loop from domain III, suggesting that the PPIase activity of the protein could be regulated. The dimer interface is formed between domains I and III and contains residues of mixed properties. Further implications about dimerization, ribosome binding, and other functions of trigger factor are discussed.

Structure of trigger factor binding domain in biologically homologous complex with eubacterial ribosome reveals its chaperone action

“…In general, the TFa bound structure accords with the previously determined tertiary folds of TF N-terminal domain (14,15,21). The signature motif ( 41 GFRXGXXP 48 ) is part of a loop, L1, surrounded by three ␣-helices (A1, A2, and A3), of which A2 and A3 are connected sequentially and appear together like a long broken ␣ helix.…”

“…In comparison with the unbound state (14,15,21), the binding region of TFa on the ribosome displays a rigid, well defined conformation, resembling that observed in the H50S-EcTFa complex. Helix A1 and helices A2 and A3, which in some structures appear as a single helix kinked at Gly-57 (Gly-59Ec) but not broken, are drawn 6 Å apart in the ribosomebound structure in contrast to their being ''wrapped on one another'' in unbound TFa (14,15,21). Also, the ␤-sheet is not tightly packed with helix A3, as observed in all structures of unbound TFa.…”

“…Farther from the ribosome, the electron density map for TFa is less well defined. Yet, the two ␤ turns and the N-terminal strand of the ␤-structure are unambiguously determined, enabling straightforward tracing of the ␣ carbons of the entire ␤ region, based on the previously published structures of this domain (14,21).…”

“…The crystal structure of the entire TFa, in complex with the large ribosomal subunit from eubacteria, reveals a significant conformational difference compared with its unbound structure, suggesting, in accord with previous biochemical results (14), that TFa adopts an altered structure upon its association with the ribosome. The previously reported native structures of TF and its N-terminal domain (14,15,21) display a substantial variability in the conformation of binding loop L1. Moreover, the 2.5-Å structure of unbound TF from Vibrio cholerae (21) does not show all of this loop's side chains, indicating its flexibility when not associated with the ribosome.…”

“…The previously reported native structures of TF and its N-terminal domain (14,15,21) display a substantial variability in the conformation of binding loop L1. Moreover, the 2.5-Å structure of unbound TF from Vibrio cholerae (21) does not show all of this loop's side chains, indicating its flexibility when not associated with the ribosome. In contrast, the electron density for this region in our difference Fourier maps and the resulting low B factors ensures rigidity of loop L1 in the bound state.…”