Our system is currently under heavy load due to increased usage. We're actively working on upgrades to improve performance. Thank you for your patience.
Abstract:Background: Bacterial peptidyl-tRNA hydrolase is essential in recycling of ribosome-dissociated peptidyl-tRNAs. Results: Comparing minimalist substrates and using NMR mapping, the RNA-binding site of the hydrolase is characterized. Conclusion: Interaction between the hydrolase and tRNA involves features common to all elongator tRNAs. Significance: Knowledge of a bacterial peptidyl-tRNA hydrolase⅐substrate complex may drive the search for enzyme inhibitors.
“…The structural difference within the acceptor stem part probably arose from the deletion of the D arm, anticodon arm and variable loop. However, a previous report demonstrated that the catalytic efficiency of Pth for the N -acetyl-histidyl-CCA-acceptor-TΨC domain of tRNA His is comparable to that for the N -acetyl-histidyl-full-length tRNA His (41), indicating that the deletion does not affect the interaction with Pth. Figure 2.Overall structure of Pth in complex with the CCA-acceptor-TΨC domain of tRNA.…”
Peptidyl-tRNA hydrolase (Pth) cleaves the ester bond between the peptide and the tRNA of peptidyl-tRNA molecules, which are produced by aborted translation, to recycle tRNA for further rounds of protein synthesis. Pth is ubiquitous in nature, and its enzymatic activity is essential for bacterial viability. We have determined the crystal structure of Escherichia coli Pth in complex with the tRNA CCA-acceptor-TΨC domain, the enzyme-binding region of the tRNA moiety of the substrate, at 2.4 Å resolution. In combination with site-directed mutagenesis studies, the structure identified the amino acid residues involved in tRNA recognition. The structure also revealed that Pth interacts with the tRNA moiety through the backbone phosphates and riboses, and no base-specific interactions were observed, except for the interaction with the highly conserved base G53. This feature enables Pth to accept the diverse sequences of the elongator-tRNAs as substrate components. Furthermore, we propose an authentic Pth:peptidyl-tRNA complex model and a detailed mechanism for the hydrolysis reaction, based on the present crystal structure and the previous studies’ results.
“…The structural difference within the acceptor stem part probably arose from the deletion of the D arm, anticodon arm and variable loop. However, a previous report demonstrated that the catalytic efficiency of Pth for the N -acetyl-histidyl-CCA-acceptor-TΨC domain of tRNA His is comparable to that for the N -acetyl-histidyl-full-length tRNA His (41), indicating that the deletion does not affect the interaction with Pth. Figure 2.Overall structure of Pth in complex with the CCA-acceptor-TΨC domain of tRNA.…”
Peptidyl-tRNA hydrolase (Pth) cleaves the ester bond between the peptide and the tRNA of peptidyl-tRNA molecules, which are produced by aborted translation, to recycle tRNA for further rounds of protein synthesis. Pth is ubiquitous in nature, and its enzymatic activity is essential for bacterial viability. We have determined the crystal structure of Escherichia coli Pth in complex with the tRNA CCA-acceptor-TΨC domain, the enzyme-binding region of the tRNA moiety of the substrate, at 2.4 Å resolution. In combination with site-directed mutagenesis studies, the structure identified the amino acid residues involved in tRNA recognition. The structure also revealed that Pth interacts with the tRNA moiety through the backbone phosphates and riboses, and no base-specific interactions were observed, except for the interaction with the highly conserved base G53. This feature enables Pth to accept the diverse sequences of the elongator-tRNAs as substrate components. Furthermore, we propose an authentic Pth:peptidyl-tRNA complex model and a detailed mechanism for the hydrolysis reaction, based on the present crystal structure and the previous studies’ results.
“…X-ray characterization of Pth with bound bases, substrate mimics, and acceptor-TψC fragments of tRNA have also been reported (Ito et al 2012;Kaushik et al 2013;Singh et al 2014a,b). NMR-based chemical shift perturbation studies have been carried out for EcPth in the presence of a minimal substrate analog (Giorgi et al 2011b), and uncharged fragments of tRNA (Giorgi et al 2011a). In the case of Pth enzymes, the catalytic site resides in a crevice on the surface of the protein.…”
Bacterial peptidyl-tRNA hydrolase (Pth; EC 3.1.1.29) hydrolyzes the peptidyl-tRNAs accumulated in the cytoplasm and thereby prevents cell death by alleviating tRNA starvation. X-ray and NMR studies of Vibrio cholerae Pth (VcPth) and mutants of its key residues involved in catalysis show that the activity and selectivity of the protein depends on the stereochemistry and dynamics of residues H24, D97, N118, and N14. D97-H24 interaction is critical for activity because it increases the nucleophilicity of H24. The N118 and N14 have orthogonally competing interactions with H24, both of which reduce the nucleophilicity of H24 and are likely to be offset by positioning of a peptidyl-tRNA substrate. The region proximal to H24 and the lid region exhibit slow motions that may assist in accommodating the substrate. Helix α3 exhibits a slow wobble with intermediate time scale motions of its N-cap residue N118, which may work as a flypaper to position the scissile ester bond of the substrate. Overall, the dynamics of interactions between the side chains of N14, H24, D97, and N118, control the catalysis of substrate by this enzyme.
“…Previous structural and mutagenesis studies showed that helix α6 directly interacts with the TΨC arm of the tRNA moiety of the substrate, and this interaction is necessary for the Pth activity. 60,61 Based on these facts, we infer that Pth is able to interact with the wide variety of tRNA species of its substrates by adjusting the position of helix α6.…”
Peptidyl‐tRNA hydrolase (Pth) cleaves the ester bond between the peptide and the tRNA of peptidyl‐tRNA molecules, which are the products of defective translation, to recycle the tRNA for further rounds of protein synthesis. Pth is ubiquitous in nature, and its activity is essential for bacterial viability. Here, we have determined the crystal structure of Pth from Thermus thermophilus (TtPth) at 1.00 Å resolution. This is the first structure of a Pth from a thermophilic bacterium and the highest resolution Pth structure reported so far. The present atomic resolution data enabled the calculation of anisotropic displacement parameters for all atoms, which revealed the directionality of the fluctuations of key regions for the substrate recognition. Comparisons between TtPth and mesophilic bacterial Pths revealed that their structures are similar overall. However, the structures of the N‐ and C‐terminal, loop‐helix α4, and helix α6 regions are different. In addition, the helix α1 to strand β4 region of TtPth is remarkably different from those of the mesophilic bacterial Pths, because this region is 9 or 10 amino acid residues shorter than those of the mesophilic bacterial Pths. This shortening seems to contribute to the thermostability of TtPth. To further understand the determinants for the thermostability of TtPth, we compared various structural factors of TtPth with those of mesophilic bacterial Pths. The data suggest that the decreases in accessible surface area and thermolabile amino acid residues, and the increases in ion pairs, hydrogen bonds, and proline residues cooperatively contribute to the thermostability of TtPth.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
