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Query: UNIPROT:Q07644 (polypeptide)
 72,197 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Termination of protein synthesis is promoted in ribosomes by proper stop codon discrimination by class 1 polypeptide release factors (RFs). A large set of prokaryotic RFs differing in stop codon specificity, RF1 for UAG and UAA, and RF2 for UGA and UAA, was analyzed by means of a recently developed computational method allowing identification of the specificity-determining positions (SDPs) in families composed of proteins with similar but not identical function. Fifteen SDPs were identified within the RF1/2 superdomain II/IV known to be implicated in stop codon decoding. Three of these SDPs had particularly high scores. Five residues invariant for RF1 and RF2 [invariant amino acid residues (IRs)] were spatially clustered with the highest-scoring SDPs that in turn were located in two zones within the SDP/IR area. Zone 1 (domain II) included PxT and SPF motifs identified earlier by others as 'discriminator tripeptides'. We suggest that IRs in this zone take part in the recognition of U, the first base of all stop codons. Zone 2 (domain IV) possessed two SDPs with the highest scores not identified earlier. Presumably, they also take part in stop codon binding and discrimination. Elucidation of potential functional role(s) of the newly identified SDP/IR zones requires further experiments.
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PMID:Common and specific amino acid residues in the prokaryotic polypeptide release factors RF1 and RF2: possible functional implications. 1616 10

About 30 years ago, experiments found that there are polarity and hydrophobicity (P and H) correlations and affinity between amino acids and their anticodons. Although it is shown that these experimental findings are important for explaining the origins of the genetic code, the great potential of P and H interactions in investigating other bio-functions have not been fully explored. Here, through raising, discussing and answering seven relevant questions hidden in tRNA aminoacylation, the formation of peptide bonds, and the ending of translations, etc., we show our theoretical findings that the P and H correlations and affinity take vital roles in the protein synthesis process. We found the relationship between the 3' end ACCN sequences of tRNA molecules and the activated amino acids and its biological significance, the rRNAs' consensus sequences 5'NCC...TGG3' or 5'TGG...NCC3' which may perform as functional segments of rRNAs to help triggering the reaction of peptide formation, and common nature of releasing factors that the first amino acid residue of releasing factors ERF, RF1 and RF2 are all Methionine, except a few Alanine, which may be necessary for releasing the translated polypeptide and stopping the translating process. In the terms of P and H correlations and affinity, we provide explanations of why only using the poly (G) as mRNA template cannot get the poly (Gly) in experiments deciphering the genetic code, why Gly often appears in beta turns and why translational bypassing might occur when translating 5'GGAUGA on mRNA. Since amino acids and nucleotides are the subunits, respectively, for composing proteins and nucleic acids, these findings will help in further understanding interactions among the bio-macromolecules. These findings are also helpful for investigating rRNAs, further understanding the protein synthesis process and analysing similar bio-problems, and should be proved useful for experimental biologists.
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PMID:Polarity and hydrophobicity interactions in protein synthesis process. 1625 10

During protein synthesis, translational release factors catalyze the release of the polypeptide chain when a stop codon on the mRNA reaches the A site of the ribosome. The detailed mechanism of this process is currently unknown. We present here the crystal structures of the ribosome from Thermus thermophilus with RF1 and RF2 bound to their cognate stop codons, at resolutions of 5.9 Angstrom and 6.7 Angstrom, respectively. The structures reveal details of interactions of the factors with the ribosome and mRNA, including elements previously implicated in decoding and peptide release. They also shed light on conformational changes both in the factors and in the ribosome during termination. Differences seen in the interaction of RF1 and RF2 with the L11 region of the ribosome allow us to rationalize previous biochemical data. Finally, this work demonstrates the feasibility of crystallizing ribosomes with bound factors at a defined state along the translational pathway.
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PMID:Crystal structures of the ribosome in complex with release factors RF1 and RF2 bound to a cognate stop codon. 1637 66

In eubacteria, termination of translation is signaled by any one of the stop codons UAA, UAG, and UGA moving into the ribosomal A site. Two release factors, RF1 and RF2, recognize and bind to the stop codons with different affinities and trigger the hydrolysis of the ester bond that links the polypeptide with the P-site tRNA. Cryo-electron microscopy (cryo-EM) results obtained in this study show that ribosome-bound RF1 is in an open conformation, unlike the closed conformation observed in the crystal structure of the free factor, allowing its simultaneous access to both the decoding center and the peptidyl-transferase center. These results are similar to those obtained for RF2, but there is an important difference in how the factors bind to protein L11, which forms part of the GTPase-associated center of the large ribosomal subunit. The difference in the binding position, C-terminal domain for RF2 versus N-terminal domain for RF1, explains a body of L11 mutation studies that revealed differential effects on the activity of the two factors. Very recent data obtained with small-angle X-ray scattering now reveal that the solution structure of RF1 is open, as here seen on the ribosome by cryo-EM, and not closed, as seen in the crystal.
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PMID:Interactions of the release factor RF1 with the ribosome as revealed by cryo-EM. 1647 44

Bacterial release factors RF1 and RF2 are methylated on the Gln residue of a universally conserved tripeptide motif GGQ, which interacts with the peptidyl transferase center of the large ribosomal subunit, triggering hydrolysis of the ester bond in peptidyl-tRNA and releasing the newly synthesized polypeptide from the ribosome. In vitro experiments have shown that the activity of RF2 is stimulated by Gln methylation. The viability of Escherichia coli K12 strains depends on the integrity of the release factor methyltransferase PrmC, because K12 strains are partially deficient in RF2 activity due to the presence of a Thr residue at position 246 instead of Ala. Here, we study in vivo RF1 and RF2 activity at termination codons in competition with programmed frameshifting and the effect of the Ala-246 --> Thr mutation. PrmC inactivation reduces the specific termination activity of RF1 and RF2(Ala-246) by approximately 3- to 4-fold. The mutation Ala-246 --> Thr in RF2 reduces the termination activity in cells approximately 5-fold. After correction for the decrease in level of RF2 due to the autocontrol of RF2 synthesis, the mutation Ala-246 --> Thr reduced RF2 termination activity by approximately 10-fold at UGA codons and UAA codons. PrmC inactivation had no effect on cell growth in rich media but reduced growth considerably on poor carbon sources. This suggests that the expression of some genes needed for optimal growth under such conditions can become growth limiting as a result of inefficient translation termination.
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PMID:Methylation of bacterial release factors RF1 and RF2 is required for normal translation termination in vivo. 1793 46

During translation termination, release factor (RF) protein catalyzes a hydrolytic reaction in the large subunit peptidyl transferase center to release the finished polypeptide chain. While the mechanism of catalysis of peptide release remains obscure, important contributing factors have been identified, including conserved active-site nucleotides and a GGQ tripeptide motif in the RF. Here we describe pre-steady-state kinetic and nucleophile competition experiments to examine RF contributions to the rate and specificity of peptide release. We find that while unacylated tRNA stimulates release in a nondiscriminating manner, RF1 is very specific for water. Further analysis reveals that amino acid Q235 of the RF1 GGQ motif is critical for the observed specificity. These data lead to a model where RFs make two distinct contributions to catalysis--a relatively nonspecific activation of the catalytic center and specific selection of water as a nucleophile facilitated by Q235.
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PMID:Two distinct components of release factor function uncovered by nucleophile partitioning analysis. 1799 9

Novel mutations in prfA, the gene for the polypeptide release factor RF1 of Escherichia coli, were isolated using a positive genetic screen based on the parD (kis, kid) toxin-antitoxin system. This original approach allowed the direct selection of mutants with altered translational termination efficiency at UAG codons. The isolated prfA mutants displayed a approximately 10-fold decrease in UAG termination efficiency with no significant changes in RF1 stability in vivo. All three mutations, G121S, G301S and R303H, were situated close to the nonsense codon recognition site in RF1:ribosome complexes. The prfA mutants displayed increased sensitivity to the RelE toxin encoded by the relBE system of E. coli, thus providing in vivo support for the functional interaction between RF1 and RelE. The prfA mutants also showed increased sensitivity to the Kid toxin. Since this toxin can cleave RNA in a ribosome-independent manner, this result was not anticipated and provided first evidence for the involvement of RF1 in the pathway of Kid toxicity. The sensitivity of the prfA mutants to RelE and Kid was restored to normal levels upon overproduction of the wild-type RF1 protein. We discuss these results and their utility for the design of novel antibacterial strategies in the light of the recently reported structure of ribosome-bound RF1.
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PMID:Novel Escherichia coli RF1 mutants with decreased translation termination activity and increased sensitivity to the cytotoxic effect of the bacterial toxins Kid and RelE. 1901 62

Quality control: The incorporation of a wrong amino acid into a growing polypeptide chain induces a correction step in which the release factor (RF1) hydrolyzes the peptide from the incorrectly matched peptidyl-tRNA (see picture). The nascent erroneous polypeptide is released from the ribosome and degraded.
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PMID:Correction of errors during protein synthesis. 1930 38

The bacterial translation factor RF3 promotes translation termination by recycling the tRNA-mimicking release factors, RF1 and RF2, after mature polypeptide release. RF3 also enhances the premature peptidyl-tRNA drop-off reaction in the presence of RRF and EF-G. Despite the recently resolved X-ray crystal structure of RF3, the molecular details of the bimodal functionality of RF3 remain obscure. In this report, we demonstrate a novel class of RF3 mutations specifically defective in the tRNA drop-off reaction. These mutations suggest differential molecular pathways closely related to the guanine nucleotide modes of RF3.
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PMID:A novel class of bacterial translation factor RF3 mutations suggests specific structural domains for premature peptidyl-tRNA drop-off. 2004 13

In termination of protein synthesis, the bacterial release factors RF1 and RF2 bind to the ribosome through specific recognition of messenger RNA stop codons and trigger hydrolysis of the bond between the nascent polypeptide and the transfer RNA at the peptidyl-tRNA site, thereby releasing the newly synthesized protein. The release factors are highly specific for a U in the first stop-codon position and recognize different combinations of purines in the second and third positions, with RF1 reading UAA and UAG and RF2 reading UAA and UGA. With recently determined crystal structures of termination complexes, it has become possible to decipher the energetics of stop-codon reading by computational analysis and to clarify the origin of the high release-factor binding accuracy. Here we report molecular dynamics free-energy calculations on different cognate and non-cognate termination complexes. The simulations quantitatively explain the basic principles of decoding in all three codon positions and reveal the key elements responsible for specificity of the release factors. The overall reading mechanism involves hitherto unidentified interactions and recognition switches that cannot be described in terms of a tripeptide anticodon model. Further simulations of complexes with tRNA(Trp), the tRNA recognizing the triplet codon for Trp, explain the observation of a 'leaky' stop codon and highlight the fundamentally different third position reading by RF2, which leads to a high stop-codon specificity with strong discrimination against the Trp codon. The simulations clearly illustrate the versatility of codon reading by protein, which goes far beyond tRNA mimicry.
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PMID:Principles of stop-codon reading on the ribosome. 2051 19


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