Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “Peptide Elongation Factors”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 19 recordsLinked to original sources
PubMed1988

Peptide elongation factor 1 from yeasts: purification and biochemical characterization of peptide elongation factors 1 alpha and 1 beta (gamma) from Saccharomyces carlsbergensis and Schizosaccharomyces pombe.

Cytoplasmic elongation factor 1 alpha (EF-1 alpha) [corrected] was purified to homogeneity in high yield from the two different yeasts Saccharomyces carlsbergensis (S. carls.) and Schizosaccharomyces pombe (S. pombe). The purification was easily achieved by CM-Sephadex column chromatography of the breakthrough fractions from DEAE-Sephadex chromatography of cell-free extracts. The basic proteins have a molecular weight of 47,000 for the S. carls. factor and of 49,000 for the S. pombe factor. While the purified yeast EF-1 alpha s function analogously to other eukaryotic factors and the E. coli EF-Tu in Phe-tRNA binding and polyphenylalanine synthesis, the yeast factor unusually hydrolyzed GTP on yeast ribosomes upon addition of Phe-tRNA in the absence of poly(U) as mRNA. This novelty is probably owing to the yeast ribosomes, which are assumed to lack elongation factor 3-equivalent component(s). Trypsin and chymotrypsin selectively cleaved the two yeast factors to generate resistant fragments with the same molecular weight of 43,000 (by trypsin) and of 44,000 (by chymotrypsin), respectively. Those cleavage sites were characteristically protected by the presence of several ligands bound to EF-1 alpha such as GDP, GTP, and aminoacyl-tRNA. Based on the sequence analysis of the fragments generated by the two proteases, the partial amino acid sequence of the S. carls. EF-1 alpha was deduced to be in accordance with the N-terminal region covering positions (1) to 94 and two Lys residues at the C-terminal end of the predicted total sequence of the Saccharomyces cerevisiae (S. cerev.) factor derived from DNA analysis, except for a few N-terminal residues, confirming the predicted S. cerev. sequence at the protein level. EF-1 beta and EF-1 beta gamma were isolated and highly purified as biologically active entities from the two yeasts. EF-1 beta s from the two yeasts have the same molecular weight of 27,000, whereas component gamma of the S. carls. EF-1 beta gamma showed a higher molecular weight (47,000) than that of the S. pombe factor (40,000). It was also shown that a stoichiometric complex was formed between EF-1 alpha and EF-1 beta gamma from S. pombe. Furthermore, a considerable amount of Phe-tRNA binding activity was distributed in the EF-1H (probably EF-1 alpha beta gamma) fraction from freshly prepared cell-free extracts of yeast.

Amino Acid Sequence↗
PubMed1985

Regulation of the activity of eukaryotic peptide elongation factor 1 by autocatalytic phosphorylation.

Highly purified peptide elongation factor 1 from rabbit reticulocytes liberates the terminal phosphate from [gamma-32P]GTP and incorporates it into its own protein. Approximately one phosphate residue becomes bound by one molecule of the factor. Only the eEF-1 alpha subunit of the factor (Mr 53 000) becomes phosphorylated as revealed by polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate followed by autoradiography and by the incubation of [gamma-32P]GTP with individual subunits of the elongation factor separated by chromatofocusing in the presence of 5 M urea. The phosphorylation also takes place, though to a lesser extent, if the factor is incubated with Na2H32PO4, probably due to the presence of endogenous GTP bound in the molecule of the factor. The content of endogenous GTP in various factor preparations was 0.21-0.43 mol/mol factor. Phosphorylation of the peptide elongation factor is ribosome-independent, acid-labile and apparently autocatalytic since no other proteins are required for this reaction. Preincubation of the factor with GTP or with inorganic phosphate results in the phosphorylation of the factor and is followed by an enhanced binding of phenylalanyl-tRNA to 80S ribosomes in the presence of poly(U). This is accompanied by a dephosphorylation of the factor protein and thus the reversible autophosphorylation of the factor apparently activates its binding site for aminoacyl-tRNA. This is supported by the observation that sodium fluoride, which inhibits the dephosphorylation of the factor, blocks the factor-catalyzed binding of aminoacyl-tRNA to ribosomes. The incorporation of phosphate into factor protein also inhibits the formation of an eEF-1 X GDP complex, which is inactive in protein synthesis. Thus GDP liberated by the GTPase activity of the factor cannot affect its binding site for aminoacyl-tRNA. This may be the other reason for the enhanced activity of the phosphorylated factor. The autocatalytic GTP-dependent phosphorylation of the peptide elongation factor 1 apparently modifies its function and may thus play a regulatory role in protein synthesis.

Animals↗
PubMed1978

Decreased activity of peptide-elongation factors after treatment with cholesterol esterase.

1. Peptide-elongation factors were purified from rat liver and treated with cholesterol esterase and phospholipase A2 immobilized on Sepharose 4B. 2. Binding of L-[3H]-phenylalanyl-tRNA to 40S ribosomal subunits was decreased by approx. 70% and to polyribosomes by 30% in the presence of the binding factor incubated with cholesterol esterase. Treatment of this factor with immobilized phospholipase A2 decreased the binding to smaller ribosomal subunits by only about 15%. 3. Poly(U)-dependent phenylalanine polymerization by ribosomal subunits was decreased to approx. 30% of its original value by treatment of both elongation factors with cholesterol esterase. 4. The normal activity of esterase-treated elongation factor in both the binding reaction and peptide-elongation assay was fully recovered by the addition of cholesteryl 14-methyl-hexadecanoate. 5. Different classes of lipids present in peptide-elongation factor 1 have apparently different functions. Whereas phospholipids are required to maintain the strcture of heavy aggregates of this factor, the presence of cholesteryl 14-methylhexadecanoate is obviously necessary for the normal function of peptide-elongation factors.

Animals↗
PubMed1985

The role of cholesteryl 14-methylhexadecanoate in the function of eukaryotic peptide elongation factor 1.

The binding of [3H]cholesteryl 14-methylhexadecanoate by a highly purified peptide elongation factor 1 from rabbit reticulocytes is significantly enhanced by GTP and CTP, much less by guanosine 5'-[beta, gamma-methylene]-triphosphate and not at all by ATP or UTP. Removal of endogenous cholesteryl 14-methylhexadecanoate present in the molecule of the factor [Hradec, J. et al. (1971) Biochem. J. 123, 959-966] by digestion with immobilized cholesterol esterase resulted in an almost complete loss of GTPase activity and this could be restored to nearly normal values by the addition of the ester. The same holds true for the GTP-dependent autophosphorylation of the protein-synthesis factor. Cholesteryl 14-methylhexadecanoate was bound only by the beta subunit of the factor. Addition of the alpha subunit, which was inactive on its own, stimulated the binding of the ester to the beta subunit in a sigmoid dependence. The binding of the ester was significantly stimulated by aminoacyl-tRNA but this effect was fully abolished by sodium fluoride, indicating a relation of cholesteryl 14-methylhexadecanoate to the dephosphorylation of the peptide elongation factor. Treatment of the factor with cholesterol esterase decreased its activity in the poly(U)-dependent binding of phenylalanyl-tRNA to ribosome and this activity was again restored by the addition of cholesteryl 14-methylhexadecanoate. The ester thus interacts with the GTP-dependent autophosphorylation of peptide elongation factor 1 and in this way modulates the activity of the factor. A putative scheme is presented explaining the mode of action of cholesteryl 14-methylhexadecanoate.

Animals↗
PubMed1980

Cholesteryl esters are bound by a peptide-initiation and a peptide-elongation factor.

Significantly higher quantities of cholesteryl 14-methylhexadecanoate than of cholesteryl laurate and cholesteryl palmitate are bound by a homogeneous peptide-initiation factor and purified peptide-elongation factor 1. Cholesteryl 14-methylhexadecanoate may function as a specific allosteric modifier changing the conformation of protein synthesis factors and thus modulating the activity of their binding sites.

Binding Sites↗
PubMed1984

CTP can replace GTP in reactions catalyzed by eukaryotic peptide elongation factor 1.

In several reactions catalyzed by highly purified peptide elongation factor 1 from rabbit reticulocytes, GTP may be fully replaced by CTP but not by ATP or UTP. This holds true for the factor-dependent binding of aminoacyl-tRNA to ribosomes, GTPase activity, GTP-dependent autophosphorylation of the factor protein and binding of cholesteryl 14-methylhexadecanoate by the factor.

Adenosine Triphosphate↗
PubMed1978

Protein synthesis in tumor host. II. Increased activity of peptide elongation factor 1 in experimental rat tumors and in host liver.

Both peptide elongation factors were purified from Zajdela and Walker tumors and from the host and normal liver. The activity of peptide elongation factor 1 from tumor tissues in promoting the binding of phenylalanyl-tRNA to ribosomes was significantly higher than that of normal liver. Also preparations from the host liver were markedly more active when compared with the corresponding factor from normal liver. Poly(U)-dependent phenylalanine polymerization was enhanced in subcellular systems containing elongation factor 1 from tumors or host liver. No differences were found between preparations of elongation factor 2 isolated from various sources. Tumor ribosomes showed an increased activity of both the acceptor and donor binding site of peptidyl transferase. In ribosomes from the tumor-host liver the activity of the acceptor site of this enzyme was decreased while that of the donor site remained unaltered. The enhanced activity of peptide elongation factor 1 from tumors and host liver is apparently the main reason of enhanced protein synthesis in these tissues. The enhanced activity of this factor is not specific for tumor growth as it occurs also in other pathological conditions.

Animals↗
PubMed2001

Peptide elongation factor eEF1A-2/S1 expression in cultured differentiated myotubes and its protective effect against caspase-3-mediated apoptosis.

Peptide elongation factor eEF1A-2/S1, which shares 92% homology with eEF1A-1/EF-1alpha, is exclusively expressed in brain, heart, and skeletal muscle. In these tissues, eEF1A-2/S1 is the only type 1A elongation factor expressed in adulthood because a transition from eEF1A-1/EF-1alpha to eEF1A-2/S1 occurs in early postnatal development. In this article, we report that the expression of eEF1A-2/S1 protein is activated upon myogenic differentiation. Furthermore, we show that upon serum deprivation-induced apoptosis, eEF1A-2/S1 protein disappears and is replaced by its homolog eEF1A-1/EF-1alpha in dying myotubes; cell death is characterized by the activation of caspase-3. In addition, we show that the continuous expression of eEF1A-2/S1 resulting from adenoviral gene transfer protects differentiated myotubes from apoptosis by delaying their death, thus suggesting a prosurvival function for eEF1A-2/S1 in skeletal muscle. In contrast, myotube death is accelerated by the introduction of the homologous gene, eEF1A-1/EF-1alpha, whereas cells transfected with antisense eEF1A-1/EF-1alpha are protected from apoptosis. These results demonstrate that the two sister genes, eEF1A-1/EF-1alpha and eEF1A-2/S1, regulate myotube survival with the former exerting prodeath activity and the latter a prosurvival effect.

Adenoviridae↗
PubMed1989

Sequence analysis of the peptide-elongation factor EF-2 gene, downstream from those of ribosomal proteins H-S12 and H-S7, from the archaebacterial extreme halophile, Halobacterium halobium.

The gene for the peptide-elongation factor 2 (EF-2) was cloned from the archaebacterial extreme halophile Halobacterium halobium and sequenced. The 1013 nucleotides upstream from this gene was two open reading frames similar to ribosomal proteins S12 and S7 from Escherichia coli. Sequence alignment studies showed the halobacterial elongation factor 2 to be equivalent to eukaryotic EF-2 and eubacterial EF-G. Sequence similarity to the eukaryotic elongation factor was much higher than to the eubacterial factor. Conserved sequence regions were present within the factor and are likely to constitute functionally important domains. These include the sites of GTP binding and ADP ribosylation by diphtheria toxin.

Animals↗
PubMed1988

Purification and characterization of peptide-elongation factor 2 (aEF-2) from an extremely halophilic archaebacterium Halobacterium halobium.

A procedure is described for the purification of the archaebacterial peptide-elongation factor 2 (aEF-2) from an extremely halophilic archaebacterium Halobacterium halobium. The enrichment was about 530-fold, the obtained preparation practically homogeneous as judged by SDS-PAGE. The poly(U)-dependent poly(Phe) synthesis was completely dependent on aEF-2 in the presence of partially purified aEF-1, and the activity was equivalent to a poly(Phe)-synthesizing system containing unfractionated S-100 enzymes. aEF-2 consists of a single peptide with a relative molecular mass of 125,000 +/- 3000 and 100,000 +/- 3000 as determined by SDS-PAGE and gel filtration on Sephadex G-200 respectively. The isoelectric point was 5.7. The amino acid composition analysis indicated the predominance of acidic amino acids (aspartic acid and glutamic acid) and the low content of hydrophobic amino acid (phenylalanine) as compared with those of eukaryotes and prokaryotes. The factor was stable in a pH range from 6 to 8. 2-Mercaptoethanol and GTP but not GDP markedly protected aEF-2 from heat denaturation at 52 degrees C. aEF-2 became inactivated and insensitive to ADP-ribosylation by diphtheria toxin at low ionic strength but could be renatured by increasing ionic strength. Obviously higher concentrations of salts contribute to the conformational stability of aEF-2.

Adenosine Diphosphate Ribose↗
PubMed1988

Role of yeast peptide elongation factor 3 (EF-3) at the AA-tRNA binding step.

The stimulatory effect of peptide elongation factor 3 (EF-3), which is uniquely required for the yeast elongation cycle, on the step of binding of aminoacyl-tRNA (AA-tRNA) to ribosomes has been investigated in detail. Yeast EF-1 alpha apparently functions in a stoichiometric manner in the binding reaction of AA-tRNA to the ribosomes. The addition of EF-3 and ATP to this binding system strikingly stimulated the binding reaction, and the stimulated reaction proceeded catalytically with respect to both EF-1 alpha and EF-3, accompanied by ATP hydrolysis, indicating that EF-3 stimulated the AA-tRNA binding reaction by releasing EF-1 alpha from the ribosomal complex, thus recycling it. This binding stimulation by EF-3 was in many respects distinct from that by EF-1 beta gamma. The idea that EF-3 may participate in the regeneration of GTP from ATP and the formed GDP, as indicated by the findings that the addition of EF-3 along with ATP allowed the AA-tRNA binding and Phe polymerization reactions to proceed even in the presence of GDP in place of GTP, was not verified by the results of direct measurement of [32P]GTP formation from [gamma-32P]ATP and GDP under various conditions. Examination of the stability of the bound AA-tRNA disclosed the different binding states of AA-tRNA on ribosomes between in the cases of the complexes formed with EF-1 alpha alone, or factor-independently, and with EF-1 alpha and EF-3.(ABSTRACT TRUNCATED AT 250 WORDS)

Fungal Proteins↗
PubMed1988

The yeast peptide elongation factor 3 (EF-3) carries an active site for ATP hydrolysis which can interact with various nucleoside triphosphates in the absence of ribosomes.

ATP (GTP) hydrolysis was clearly demonstrated by using at most 16 pmol of yeast peptide elongation factor 3 (EF-3) in the absence of ribosomes. However, the highly active yeast ribosomes (up to 48 pmol) displayed virtually no ATPase (or GTPase) activity in the absence of EF-3. Several lines of evidence indicated that both the catalytic and binding sites of the ATPase reside in the elongation factor itself, not on the ribosomes. The patterns of protection by various nucleoside triphosphates against tryptic digestion of EF-3, reflecting the wide substrate specificity of the ATPase, confirmed that the active center of the endogenous ATPase is located on the factor itself and not on contaminants. The intrinsic activity was stimulated up to two orders of magnitude by the presence of the yeast ribosomes fully active in polyphenylalanine synthesis. The activation was achieved by enhancing the catalytic activity (kcat) to a much greater extent than the binding affinity (Km). On the other hand, the ribosome-activated ATPase activity was revealed to inherit its wide substrate specificity from the intrinsic property of EF-3, which shows an affinity to various XTPs, including pyrimidine- and purine-nucleoside triphosphates, irrespective of 2'-hydroxylation of the sugar moiety. From experiments on protection against tryptic digestion, we determined that intricate conformational changes of the factor molecule occur upon interaction with the substrate XTP and ribosomes.

Adenosine Triphosphatases↗
PubMed1990

Soluble factor requirements for the Tetrahymena peptide elongation system and the ribosomal ATPase as a counterpart of yeast elongation factor 3 (EF-3).

Peptide elongation factor 3 (EF-3), which is widely present in yeasts and fungi (Eumycota), does not occur in another lower eukaryote, the unicellular protozoan Tetrahymena pyriformis, as was shown by the following findings: (a) there is no activity to satisfy the EF-3 requirement of yeast ribosomes in the post-ribosomal supernatant fraction from Tetrahymena, and (b) the Tetrahymena ribosomes displayed their full capacity for polyphenylalanine synthesis with purified EF-1 alpha and EF-2 alone from either Tetrahymena or yeast, and their activity on the Tetrahymena ribosomes was not further enhanced by the addition of yeast EF-3, in contrast to the case of the yeast ribosomes. However, as a substitute for the ribosome-activated nucleotidase activity of EF-3, Tetrahymena ribosomes were shown to harbor strong, firmly bound ATPase and GTPase activities, which probably involve the same active site. The ribosome-bound ATPase activity was inhibited by a polyclonal antibody raised against yeast EF-3 with the same inactivation profile as that of polyphenylalanine synthesis on Tetrahymena ribosomes, indicating that the ribosomal ATPase plays an essential role in the elongation process on Tetrahymena ribosomes as previously revealed in the yeast system. It was also shown that the ribosomal nucleotidase plays a pivotal role in the elongation cycle in other eukaryotes.

Adenosine Triphosphatases↗
PubMed2004

Immuno-characterization of the switch of peptide elongation factors eEF1A-1/EF-1alpha and eEF1A-2/S1 in the central nervous system during mouse development.

During early postnatal development, a switch occurs between eEF1A-1/EF-1alpha and eEF1A-2/S1, homologous peptide elongation factors, in brain, heart, and skeletal muscle; eEF1A-2/S1 becomes the major form expressed in maturity. By immunofluorescent labeling, we detected both homologues in the developing brains of wild-type and wasted mutant mice, carrying a deletion in the eEF1A-2/S1 gene; we found that brain expression of eEF1A-2/S1 protein is restricted to mature, terminally differentiated neurons, and coincides with the disappearance of eEF1A-1/EF-1alpha 20 days after birth. Furthermore, no elongation factor 1A is present in wasted mutant mice neurons following the developmental switch, indicating that the genetic regulation silencing eEF1A-1/EF-1alpha is still functional.

Animals↗
PubMed2000

Rapid up-regulation of peptide elongation factor EF-1alpha protein levels is an immediate early event during oxidative stress-induced apoptosis.

Hydrogen peroxide (H(2)O(2)) induces apoptosis in cultured cells, in a dose-dependent manner. Treatment with H(2)O(2) causes decreased mitochondrial respiration, along with DNA degradation and the formation of an oligonucleosomal ladder, all hallmarks of apoptotic cell death. In this report, we investigate alterations in expression of a peptide elongation factor, EF-1alpha, during oxidative challenge. EF-1alpha protein levels undergo rapid increase upon treatment with H(2)O(2); however, whereas sublethal doses of H(2)O(2) stimulate only transient increases of EF-1alpha protein levels, lethal doses produce sustained elevation of EF-1alpha levels. Furthermore, pretreatment of H9c2(2-1) cells with transcriptional inhibitors fails to abolish the oxidant-induced increase in EF-1alpha, and Northern blotting analysis reveals that EF-1alpha mRNA levels remain steady throughout the H(2)O(2) treatment period, suggesting that the up-regulation of EF-1alpha is mediated posttranscriptionally. Transient transfection with an antisense EF-1alpha cDNA protects against hydrogen peroxide-mediated cytotoxicity in proportion to the degree of repression of EF-1alpha protein levels, suggesting that up-regulation of EF-1alpha plays a role in expediting the execution of the apoptotic program in response to oxidative stress.

Actins↗
PubMed1994

Staurosporine and gossypol are inhibitors of the function of peptide elongation factor 1 alpha from rabbit reticulocytes.

As it has been found, the incubation of [gamma-32P]ATP with elongation factor--1 alpha purified from rabbit reticulocytes resulted in the phosphorylation of several substrate proteins /Tuhácková, Z. (1992) In: Rec. Adv. Cell. Mol. Biol. 4, 79-86, Peeters Press, Leuven/ (1). In the present paper chromatofocusing of the purified eEF-1 alpha demonstrates that the ATP-dependent protein kinase activity is associated with a single protein catalyzing the GTP-dependent binding of aminoacyl-tRNA to ribosomes. Both of these activities are inhibited by staurosporine and gossypol. The inhibition by GDP but not by GTP indicates a possible involvement of conformation changes also in the modulation of the protein kinase activity displayed by eEF-1 alpha.

Alkaloids↗