Structure-specific tRNA-binding protein from the extreme thermophile Aquifex aeolicus.
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 A 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.
Morales
A J
AJ
Skaggs Institute for Chemical Biology and Departments of Molecular Biology and Chemistry, The Scripps Research Institute, 10550 N. Torrey Pines Rd, La Jolla, CA 92037, USA.
Swairjo
M A
MA
Schimmel
P
P
D000595:Amino Acid Sequence; D000604:Amino Acyl-tRNA Synthetases / Q000737:chemistry / Q000235:genetics; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D001483:Base Sequence; D001667:Binding, Competitive; D003001:Cloning, Molecular; D003460:Crystallization; D018360:Crystallography, X-Ray; D019281:Dimerization; D016946:Gram-Negative Aerobic Rods and Cocci / Q000235:genetics*; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010455:Peptides / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D011485:Protein Binding; D011487:Protein Conformation; D012343:RNA, Transfer / Q000235:genetics / Q000378:metabolism*; D012358:RNA, Transfer, Met / Q000235:genetics / Q000378:metabolism; D016601:RNA-Binding Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity
D001426:Bacterial Proteins; D010455:Peptides; D012358:RNA, Transfer, Met; D016601:RNA-Binding Proteins; C120618:Trbp111 protein, Aquifex aeolicus; D012343:RNA, Transfer; D000604:Amino Acyl-tRNA Synthetases