The Experts below are selected from a list of 219 Experts worldwide ranked by ideXlab platform
Keith S. Wilson - One of the best experts on this subject based on the ideXlab platform.
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the Polypeptide Chain fold in tyrosine phenol lyase a pyridoxal 5 phosphate dependent enzyme
FEBS Letters, 1992Co-Authors: Alfred A Antson, Tatyana V Demidkina, Zbigniew Dauter, Emil H Harutyunyan, Boris V Strokopytov, Michail N. Isupov, Howard Terry, Garib N. Murshudov, Dmitry G Vassylyev, Keith S. WilsonAbstract:The tyrosine phenol lyase (EC 4.1.99.2) from Citrobacter intermedius has been crystallised in the apo form by vapour diffusion. The space group is P21212. The unit cell has dimensions a = 76.0 A, b = 138.3 A, c = 93.5 A and it contains two subunits of the tetrameric molecule in the asymmetric unit, Diffraction data for the native enzyme and two heavy atom derivatives have been collected with synchrotron radiation and an image plate scanners. The structure has been solved at 2.7 A resolution by isomorphous replacement with subsequent modification of the phases by averaging the density around the non-crystallographic symmetry axis. The electron density maps clearly show the relative orientation of the subunits and most of the trace of the Polypeptide Chain. Each subunit consists of two domains. The topology of the large domain appears to be similar to that of the aminotransferases.
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the Polypeptide Chain fold in tyrosine phenol lyase a pyridoxal 5 phosphate dependent enzyme
FEBS Letters, 1992Co-Authors: Alfred A Antson, Tatyana V Demidkina, Zbigniew Dauter, Emil H Harutyunyan, Boris V Strokopytov, Michail N. Isupov, Howard Terry, Garib N. Murshudov, Dmitry G Vassylyev, Keith S. WilsonAbstract:The tyrosine phenol lyase (EC 4.1.99.2) from Citrobacter intermedius has been crystallised in the apo form by vapour diffusion. The space group is P21212. The unit cell has dimensions a = 76.0 A, b = 138.3 A, c = 93.5 A and it contains two subunits of the tetrameric molecule in the asymmetric unit, Diffraction data for the native enzyme and two heavy atom derivatives have been collected with synchrotron radiation and an image plate scanners. The structure has been solved at 2.7 A resolution by isomorphous replacement with subsequent modification of the phases by averaging the density around the non-crystallographic symmetry axis. The electron density maps clearly show the relative orientation of the subunits and most of the trace of the Polypeptide Chain. Each subunit consists of two domains. The topology of the large domain appears to be similar to that of the aminotransferases.
Lev L Kisselev - One of the best experts on this subject based on the ideXlab platform.
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positioning of the mrna stop signal with respect to Polypeptide Chain release factors and ribosomal proteins in 80s ribosomes
FEBS Letters, 2002Co-Authors: K N Bulygin, Ludmila Frolova, M N Repkova, A G Venyaminova, D M Graifer, G G Karpova, Lev L KisselevAbstract:Abstract To study positioning of the mRNA stop signal with respect to Polypeptide Chain release factors (RFs) and ribosomal components within human 80S ribosomes, photoreactive mRNA analogs were applied. Derivatives of the UUCUAAA heptaribonucleotide containing the UUC codon for Phe and the stop signal UAAA, which bore a perfluoroaryl azido group at either the fourth nucleotide or the 3′-terminal phosphate, were synthesized. The UUC codon was directed to the ribosomal P site by the cognate tRNA Phe , targeting the UAA stop codon to the A site. Mild UV irradiation of the ternary complexes consisting of the 80S ribosome, the mRNA analog and tRNA resulted in tRNA-dependent crosslinking of the mRNA analogs to the 40S ribosomal proteins and the 18S rRNA. mRNA analogs with the photoreactive group at the fourth uridine (the first base of the stop codon) crosslinked mainly to protein S15 (and much less to S2). For the 3′-modified mRNA analog, the major crosslinking target was protein S2, while protein S15 was much less crosslinked. Crosslinking of eukaryotic (e) RF1 was entirely dependent on the presence of a stop signal in the mRNA analog. eRF3 in the presence of eRF1 did not crosslink, but decreased the yield of eRF1 crosslinking. We conclude that (i) proteins S15 and S2 of the 40S ribosomal subunit are located near the A site-bound codon; (ii) eRF1 can induce spatial rearrangement of the 80S ribosome leading to movement of protein L4 of the 60S ribosomal subunit closer to the codon located at the A site; (iii) within the 80S ribosome, eRF3 in the presence of eRF1 does not contact the stop codon at the A site and is probably located mostly (if not entirely) on the 60S subunit.
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the Polypeptide Chain release factor erf1 specifically contacts the s 4 uga stop codon located in the a site of eukaryotic ribosomes
FEBS Journal, 2001Co-Authors: Laurent Chavatte, Ludmila Frolova, Lev L Kisselev, Alain FavreAbstract:It has been shown previously [Brown, C.M. & Tate, W.P. (1994) J. Biol. Chem. 269, 33164-33170.] that the Polypeptide Chain release factor RF2 involved in translation termination in prokaryotes was able to photocrossreact with mini-messenger RNAs containing stop signals in which U was replaced by 4-thiouridine (s4U). Here, using the same strategy we have monitored photocrosslinking to eukaryotic ribosomal components of 14-mer mRNA in the presence of tRNA(f)(Met), and 42-mer mRNA in the presence of tRNA(Asp) (tRNA(Asp) gene transcript). We show that: (a) both 14-mer and 42-mer mRNAs crossreact with ribosomal RNA and ribosomal proteins. The patterns of the crosslinked ribosomal proteins are similar with both mRNAs and sensitive to ionic conditions; (b) the crosslinking patterns obtained with 42-mer mRNAs show characteristic modification upon addition of tRNA(Asp) providing evidence for appropriate mRNA phasing onto the ribosome. Similar changes are not detected with the 14-mer mRNA.tRNA(f)(Met) pairs; (c) when eukaryotic Polypeptide Chain release factor 1 (eRF1) is added to the ribosome.tRNA(Asp) complex it crossreacts with the 42-mer mRNA containing the s(4)UGA stop codon located in the A site, but not with the s(4)UCA sense codon; this crosslink involves the N-terminal and middle domains of eRF1 but not the C domain which interacts with eukaryotic Polypeptide Chain release factor 3 (eRF3); (d) addition of eRF3 has no effect on the yield of eRF1-42-mer mRNA crosslinking and eRF3 does not crossreact with 42-mer mRNA. These experiments delineate the in vitro conditions allowing optimal phasing of mRNA on the eukaryotic ribosome and demonstrate a direct and specific contact of 'core' eRF1 and s(4)UGA stop codon within the ribosomal A site.
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Polypeptide Chain release factors
Molecular microbiology, 1997Co-Authors: Richard H Buckingham, Guido Grentzmann, Lev L KisselevAbstract:Newly synthesized Polypeptide Chains are released from peptidyl-tRNA when the ribosome encounters a stop signal on mRNA. Extra-ribosomal proteins (release factors) play an essential role in this process. Although the termination process was first discovered in the late 1960s, much of the mechanism has remained obscure. However, important steps have recently been made in both prokaryotic and eukaryotic organisms in unlocking the secrets of this vital stage in protein synthesis. In this review we summarize these advances and focus attention on the remaining areas of uncertainty, particularly with respect to the models that have been proposed for the action of the GTP-hydrolysing termination factors in prokaryotes and eukaryotes, i.e. RF3 and eRF3.
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eukaryotic Polypeptide Chain release factor erf3 is an erf1 and ribosome dependent guanosine triphosphatase
RNA, 1996Co-Authors: L Frolova, Le X Goff, Galina A Zhouravleva, E Davydova, Michel Philippe, Lev L KisselevAbstract:Termination of translation in eukaryotes is governed by two Polypeptide Chain release factors, eRF1 and eRF3 on the ribosome. eRF1 promotes stop-codon-dependent hydrolysis of peptidyl-tRNA, and eRF3 interacts with eRF1 and stimulates eRF1 activity in the presence of GTP. Here, we have demonstrated that eRF3 is a GTP-binding protein endowed with a negligible, if any, intrinsic GTPase activity that is profoundly stimulated by the joint action of eRF1 and the ribosome. Separately, neither eRF1 nor the ribosome display this effect. Thus, eRF3 functions as a GTPase in the quaternary complex with ribosome, eRF1, and GTP. From the in vitro uncoupling of the peptidyl-tRNA and GTP hydrolyses achieved in this work, we conclude that in ribosomes both hydrolytic reactions are mediated by the formation of the ternary eRF1-eRF3-GTP complex. eRF1 and the ribosome form a composite GTPase-activating protein (GAP) as described for other G proteins. A dual role for the revealed GTPase complex is proposed: in " GTP state," it controls the positioning of eRF1 toward stop codon and peptidyl-tRNA, whereas in "GDP state," it promotes release of eRFs from the ribosome. The initiation, elongation, and termination steps of protein synthesis seem to be similar with respect to GTPase cycles.
Pinak Chakrabarti - One of the best experts on this subject based on the ideXlab platform.
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Secondary structures at Polypeptide-Chain termini and their features.
Acta crystallographica. Section D Biological crystallography, 2002Co-Authors: Rajasri Bhattacharyya, Debnath Pal, Pinak ChakrabartiAbstract:An analysis of secondary structures (alpha-helices and beta-strands) in the two terminal regions of Polypeptide Chains reveals features different from those observed over the whole protein structure. Compared with the overall distribution, the helices in the N-terminal region tend to be smaller and have higher propensities to contain Gln and Leu, while the C-terminal helices are longer and have a greater proportion of Lys and Glu. As a strand, the C-terminal region is never found in the interior of parallel beta-sheets and has a higher propensity to be at the edge of antiparallel beta-sheets. In contrast, compared with the whole structure the N-terminal region has a higher propensity to be in the interior of parallel beta-sheets. Compared with the overall distributions, terminal helices and strands show distinct periodicities in length. The Schellman motif, which is a prevalent C-capping motif in helices, is not common in C-terminal helices. There are other observations that can be used in the design of helical peptides: more residues beyond the C-terminus of helices are used for capping interactions than residues before the N-terminus. Consideration of the distribution of terminal strands in the interior and at the edge of beta-sheets suggests a sequential folding mechanism beginning at the N-terminus of the Polypeptide Chain.
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Secondary structures at Polypeptide-Chain termini and their features
Acta Crystallographica Section D Biological Crystallography, 2002Co-Authors: Rajasri Bhattacharyya, Debnath Pal, Pinak ChakrabartiAbstract:An analysis of secondary structures (α-helices and β-strands) in the two terminal regions of Polypeptide Chains reveals features different from those observed over the whole protein structure. Compared with the overall distribution, the helices in the N-terminal region tend to be smaller and have higher propensities to contain Gln and Leu, while the C-terminal helices are longer and have a greater proportion of Lys and Glu. As a strand, the C-terminal region is never found in the interior of parallel β-sheets and has a higher propensity to be at the edge of antiparallel β-sheets. In contrast, compared with the whole structure the N-terminal region has a higher propensity to be in the interior of parallel β-sheets. Compared with the overall distributions, terminal helices and strands show distinct periodicities in length. The Schellman motif, which is a prevalent C-capping motif in helices, is not common in C-terminal helices. There are other observations that can be used in the design of helical peptides: more residues beyond the C-terminus of helices are used for capping interactions than residues before the N-terminus. Consideration of the distribution of terminal strands in the interior and at the edge of β-sheets suggests a sequential folding mechanism beginning at the N-terminus of the Polypeptide Chain.
Carlos Lopezotin - One of the best experts on this subject based on the ideXlab platform.
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human polyserase 2 a novel enzyme with three tandem serine protease domains in a single Polypeptide Chain
Journal of Biological Chemistry, 2005Co-Authors: Santiago Cal, María Llamazares, Víctor Quesada, Cecilia Garabaya, Araceli Diazperales, Carlos LopezotinAbstract:We have cloned a human cDNA encoding a new serine protease that has been called polyserase-2 (polyserine protease-2) because it is the second identified human enzyme with several tandem serine protease domains in its amino acid sequence. The first serine protease domain contains all characteristic features of these enzymes, whereas the second and third domains lack one residue of the catalytic triad of serine proteases and are predicted to be catalytically inactive. This complex domain organization is also present in the sequences of mouse and rat polyserase-2 and resembles that of polyserase-1, which also contains three serine protease domains in its amino acid sequence. However, polyserase-2 lacks additional domains present in polyserase-1, including a type II transmembrane motif and a low-density lipoprotein receptor A module. Enzymatic analysis demonstrated that both full-length polyserase-2 and its first serine protease domain hydrolyzed synthetic peptides used for assaying serine proteases. Nevertheless, the activity of the isolated domain was greater than that of the entire protein, suggesting that the two catalytically inactive serine protease domains of polyserase-2 may modulate the activity of the first domain. Northern blot analysis showed that polyserase-2 is expressed in fetal kidney; adult skeletal muscle, liver, placenta, prostate, and heart; and tumor cell lines derived from lung and colon adenocarcinomas. Finally, analysis of post-translational processing mechanisms of polyserase-2 revealed that, contrary to those affecting to the membrane-bound polyserase-1, this novel polyprotein is a secreted enzyme whose three protease domains remain as an integral part of a single Polypeptide Chain.
Michel Philippe - One of the best experts on this subject based on the ideXlab platform.
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eukaryotic Polypeptide Chain release factor erf3 is an erf1 and ribosome dependent guanosine triphosphatase
RNA, 1996Co-Authors: L Frolova, Le X Goff, Galina A Zhouravleva, E Davydova, Michel Philippe, Lev L KisselevAbstract:Termination of translation in eukaryotes is governed by two Polypeptide Chain release factors, eRF1 and eRF3 on the ribosome. eRF1 promotes stop-codon-dependent hydrolysis of peptidyl-tRNA, and eRF3 interacts with eRF1 and stimulates eRF1 activity in the presence of GTP. Here, we have demonstrated that eRF3 is a GTP-binding protein endowed with a negligible, if any, intrinsic GTPase activity that is profoundly stimulated by the joint action of eRF1 and the ribosome. Separately, neither eRF1 nor the ribosome display this effect. Thus, eRF3 functions as a GTPase in the quaternary complex with ribosome, eRF1, and GTP. From the in vitro uncoupling of the peptidyl-tRNA and GTP hydrolyses achieved in this work, we conclude that in ribosomes both hydrolytic reactions are mediated by the formation of the ternary eRF1-eRF3-GTP complex. eRF1 and the ribosome form a composite GTPase-activating protein (GAP) as described for other G proteins. A dual role for the revealed GTPase complex is proposed: in " GTP state," it controls the positioning of eRF1 toward stop codon and peptidyl-tRNA, whereas in "GDP state," it promotes release of eRFs from the ribosome. The initiation, elongation, and termination steps of protein synthesis seem to be similar with respect to GTPase cycles.
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termination of translation in eukaryotes is governed by two interacting Polypeptide Chain release factors erf1 and erf3
The EMBO Journal, 1995Co-Authors: Galina A Zhouravleva, L Frolova, Le X Goff, Le R Guellec, S G Ingevechtomov, L Kisselev, Michel PhilippeAbstract:Termination of translation in higher organisms is a GTP-dependent process. However, in the structure of the single Polypeptide Chain release factor known so far (eRF1) there are no GTP binding motifs. Moreover, in prokaryotes, a GTP binding protein, RF3, stimulates translation termination. From these observations we proposed that a second eRF should exist, conferring GTP dependence for translation termination. Here, we have shown that the newly sequenced GTP binding Sup35-like protein from Xenopus laevis, termed eRF3, exhibits in vitro three important functional properties: (i) although being inactive as an eRF on its own, it greatly stimulates eRF1 activity in the presence of GTP and low concentrations of stop codons, resembling the properties of prokaryotic RF3; (ii) it binds and probably hydrolyses GTP; and (iii) it binds to eRF1. The structure of the C-domain of the X.laevis eRF3 protein is highly conserved with other Sup35-like proteins, as was also shown earlier for the eRF1 protein family. From these and our previous data, we propose that yeast Sup45 and Sup35 proteins belonging to eRF1 and eRF3 protein families respectively are also yeast termination factors. The absence of structural resemblance of eRF1 and eRF3 to prokaryotic RF1/2 and RF3 respectively, may point to the different evolutionary origin of the translation termination machinery in eukaryotes and prokaryotes. It is proposed that a quaternary complex composed of eRF1, eRF3, GTP and a stop codon of the mRNA is involved in termination of Polypeptide synthesis in ribosomes.