Structure-specific tRNA-binding protein from the extreme thermophile Aquifex aeolicus

Abstract

The genome of the bacterium Aquifex aeolicus encodes a polypeptide which is related to a small portion of a sequence found in one prokaryotic and two eukaryotic tRNA synthetases. It also is related to a portion of Arc1p, a tRNA-binding protein believed to be important for nuclear trafficking of tRNAs. Here we cloned, expressed and purified the 111 amino acid polypeptide (designated Trbp111) and showed by ultracentrifugation analysis that it is a stable dimer in solution. The protein was also crystallized in a monoclinic lattice. X-ray diffraction analysis at 2.8 Å resolution revealed a prominent non-crystallographic 2-fold axis, consistent with the presence of a symmetric homodimeric structure. Band-shift analysis with polyacrylamide gels showed that the dimer binds tRNAs, but not RNA duplexes, RNA hairpins, single-stranded RNA nor 5S rRNA. Complex formation with respect to tRNA is non-specific, with a single tRNA bound per dimer. Thus, Trbp111 is a structure-specific tRNA-binding protein. These results and other considerations raise the possibility that Trbp111 is a tRNA-specific chaperone which stabilizes the native L-shaped fold in the extreme thermophile and which has been incorporated into much larger tRNA-binding proteins of higher organisms.

Association between Archaeal Prolyl- and Leucyl-tRNA Synthetases Enhances tRNAPro Aminoacylation

“…Another auxiliary factor of the mammalian complex, p43, which is located in the core of the particle (4,35), has also been implicated in complex assembly. p43 orthologs have been identified in yeast (Arc1p) and bacteria (Trbp111) (12,14). This group of proteins preferentially binds tRNA and facilitates tRNA binding to synthetases.…”

“…The association with Arc1p was shown to increase the catalytic efficiency of the two synthetases and enhances nuclear export of tRNA. The bacterial homologue of Arc1p, Trbp111, was first found in the extreme thermophile Aquifex aeolicus and was shown to promote tRNA binding by aaRSs (14,15). Factors unrelated to the translation machinery have also been found to associate with aaRSs.…”

Aminoacyl-tRNA synthetase complexes: beyond translation

“…Trbp111, a homolog of Arc1p, is present in the extreme thermophile Aquifex aeolicus (Morales et al, 1999) and binds to single tRNA molecules as a dimer (Swairjo et al, 2000). However, it is not yet known whether this protein forms any specific complex with ARSs.…”

“…For example, the C-terminus of E. coli MRS (Morales et al, 1999), and the N-terminus of the E. coli FRS β-subunit (Simos et al, 1996) (Fig. 2) share structural similarity with the nonenzymatic factors in the mammalian ARS complex, and thus they probably function similarly to their trans-acting counterparts (Valenzuela and Schulman, 1986;Kim et al, 1993;Mosyak et al, 1995;Goldgur et al, 1997).…”

Swapping of Three‐Dimensional Domains as a Molecular Mechanism of Dimerization of Aminoacyl‐tRNA Synthetases

“…Removal of the last one from a bacterial enzyme has no significant influence on the activity of truncated MetRS monomer (18,19). A structural counterpart of the C-terminal fragment of bacterial MetRS is the dimeric protein Trbp 111 that has tRNA binding capacity and carries it to components of translational machinery (20,21). On the other hand, MetRS of Oryza sativa (rice), with significant homology in amino acid sequence (58% of similarity to the bacterial enzyme in the active site region), behaves as a monomer at low concentration (1 µM) but at high concentration can be a dimer (22).…”