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Polypeptide chain elongation factor 1 alpha (EF-1 alpha) from yeast: nucleotide sequence of one of the two genes for EF-1 alpha from Saccharomyces cerevisiae. 总被引:33,自引:5,他引:28
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S Nagata K Nagashima Y Tsunetsugu-Yokota K Fujimura M Miyazaki Y Kaziro 《The EMBO journal》1984,3(8):1825-1830
Messenger RNA for yeast cytosolic polypeptide chain elongation factor 1 alpha (EF-1 alpha) was partially purified from Saccharomyces cerevisiae. Double-stranded complementary DNA (cDNA) was synthesized and cloned in Escherichia coli with pBR327 as a vector. Recombinant plasmid carrying yEF-1 alpha cDNA was identified by cross-hybridization with the E. coli tufB gene and the yeast mitochondrial EF-Tu gene (tufM) under non-stringent conditions. A yeast gene library was then screened with the EF-1 alpha cDNA and several clones containing the chromosomal gene for EF-1 alpha were isolated. Restriction analysis of DNA fragments of these clones as well as the Southern hybridization of yeast genomic DNA with labelled EF-1 alpha cDNA indicated that there are two EF-1 alpha genes in S. cerevisiae. The nucleotide sequence of one of the two EF-1 alpha genes (designated as EF1 alpha A) was established together with its 5'- and 3'-flanking sequences. The sequence contained 1374 nucleotides coding for a protein of 458 amino acids with a calculated mol. wt. of 50 300. The derived amino acid sequence showed homologies of 31% and 32% with yeast mitochondrial EF-Tu and E. coli EF-Tu, respectively. 相似文献
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Two forms of elongation factor 1 alpha (EF-1 alpha O and 42Sp50), present in oocytes, but absent in somatic cells of Xenopus laevis
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S Deschamps J Morales A Mazabraud M le Maire H Denis D D Brown 《The Journal of cell biology》1991,114(6):1109-1111
We have purified and partially sequenced the EF-1 alpha protein from Xenopus laevis oocytes (EF-1 alpha O). We show that the two cDNA clones isolated by Coppared et al. (Coppard, N. J., K. Poulsen, H. O. Madsen, J. Frydenberg, and B. F. C. Clark. 1991. J. Cell Biol. 112:237-243) do not encode 42Sp50, as claimed by these authors, but two very similar forms of EF-1 alpha O (EF-1 alpha O and EF-1 alpha O1). 42Sp50 is the major protein component of a 42S nucleoprotein particle that is very abundant in previtellogenic oocytes of X. laevis, 42Sp50 differs from EF-1 alpha O not only by its amino acid sequence, but also by several properties already reported. In particular, 42Sp50 has a low EF-1 alpha activity. It is distributed uniformly in the cytoplasm of previtellogenic oocytes, in contrast to EF-1 alpha O which is concentrated in a small region of the cytoplasm, known as the mitochondrial mass or Balbiani body. 相似文献
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Cloning, nucleotide sequence, and expression of one of two genes coding for yeast elongation factor 1 alpha 总被引:18,自引:0,他引:18
P Cottrelle D Thiele V L Price S Memet J Y Micouin C Marck J M Buhler A Sentenac P Fromageot 《The Journal of biological chemistry》1985,260(5):3090-3096
One gene coding for yeast cytoplasmic elongation factor 1 alpha (EF-1 alpha) was isolated by colony hybridization using a cDNA probe prepared from purified EF-1 alpha mRNA. A recombinant plasmid, pLB1, with a 6-kilobase yeast DNA insert, was found by hybrid selection and translation experiments to carry the entire gene. The nucleotide sequence of the gene with its 5'- and 3'-flanking regions was determined. The 5' and 3' ends of EF-1 alpha mRNA were localized by the S1 nuclease mapping technique. The cloned gene, called TEF1, encodes a protein of 458 amino acids (Mr = 50,071) in a single, uninterrupted reading frame. The amino acid sequence shows a strong homology with several domains of Artemia salina EF-1 alpha cytoplasmic factor, as evidenced by diagonal dot matrix analysis. Protein sequence homology is comparatively much lower with the yeast mitochondrial elongation factor. S1 nuclease mapping of the mRNA, hybridization analysis of chromosomal DNA using intragenic or extragenic DNA probes, and gene disruption experiments demonstrated the existence of two genes coding for the cytoplasmic elongation factor EF-1 alpha/haploid genome. The presence of an intact chromosomal TEF1 gene is not essential for growth of haploid yeast cells. 相似文献
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Genes coding for the elongation factor EF-1 alpha in Artemia 总被引:8,自引:0,他引:8
J A Lenstra A Van Vliet A C Arnberg F J Van Hemert W M?ller 《European journal of biochemistry》1986,155(3):475-483
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Purification and characterization of an erythroid cell-specific factor that binds the murine alpha- and beta-globin genes. 总被引:7,自引:4,他引:3
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An erythroid cell-specific nuclear factor that binds tightly to a sequence motif (5'-GATAAGGA-3') shared by many erythroid cell-specific promoters was purified to homogeneity by DNA sequence affinity chromatography. Visualization of the purified factor, which we term EF-1, showed a simple pattern comprising a polypeptide doublet with Mrs of 18,000 and 19,000. We confirmed that these species account for EF-1-binding activity by eluting the polypeptides from sodium dodecyl sulfate-polyacrylamide gels and renaturing the appropriate binding activity. Using the purified polypeptides, we mapped seven factor-binding sites that are dispersed across the murine alpha- and beta-globin genes. The murine alpha-globin gene is flanked by at least two EF-1-binding sites. One site is centered at nucleotide (nt) -180 (with respect to the alpha-globin cap site). A fivefold-weaker site is located downstream of the alpha-globin poly(A) addition site, at nt +1049. We mapped five EF-1-binding sites near the murine beta-globin gene. The strongest site was centered at nt -210. Four additional sites were centered at nt -266 (adjacent to the binding site of a factor present in both murine erythroleukemia and Raji cells), -75 (overlapping the beta-globin CCAAT box), +543 (within the second intervening sequence), and -111. 相似文献
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The primary structure of the alpha subunit of human elongation factor 1. Structural aspects of guanine-nucleotide-binding sites 总被引:24,自引:0,他引:24
J H Brands J A Maassen F J van Hemert R Amons W M?ller 《European journal of biochemistry》1986,155(1):167-171
The primary structure of the alpha subunit of elongation factor 1 (EF-1 alpha) from human MOLT 4 cells was determined by cDNA sequencing. The data show that the conservation of the amino acid sequence is more than 80% when compared with yeast and Artemia EF-1 alpha. An inventory of amino acid sequences around the guanine-nucleotide-binding site in elongation factor Tu from Escherichia coli and homologous amino acid sequences in G proteins, initiation and elongation factors and proteins from the RAS family shows two regions containing conserved sequence elements. Region I has the sequence apolar-Xaa-Xaa-Xaa-Gly-Xaa-Xaa-Yaa-Xaa-Gly-LYs-Thr(Ser)- -Xaa-Xaa-Xaa-Xaa-X-apolar. Except for RAS proteins, Yaa is always an acidic amino acid residue. Region II is characterized by the invariant sequence apolar-apolar-Xaa-Xaa-Asn-Lys-Xaa-Asp. In order to facilitate sequence comparison we have used a graphic display, which is based on the hydrophilicity values of individual amino acids in a sequence. 相似文献
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The primary structure and the functional domains of an elongation factor-1 alpha from Mucor racemosus 总被引:9,自引:0,他引:9
We have determined the complete nucleotide sequence for TEF-1, one of three genes coding for elongation factor (EF)-1 alpha in Mucor racemosus. The deduced EF-1 alpha protein contains 458 amino acids encoded by two exons. The presence of an intervening sequence located near the 3' end of the gene was predicted by the nucleotide sequence data and confirmed by alkaline S1 nuclease mapping. The amino acid sequence of EF-1 alpha was compared to the published amino acid sequences of EF-1 alpha proteins from Saccharomyces cerevisiae and Artemia salina. These proteins shared nearly 85% homology. A similar comparison to the functionally analogous EF-Tu from Escherichia coli revealed several regions of amino acid homology suggesting that the functional domains are conserved in elongation factors from these diverse organisms. Secondary structure predictions indicated that alpha helix and beta sheet conformations associated with the functional domains in EF-Tu are present in the same relative location in EF-1 alpha from M. racemosus. Through this comparative structural analysis we have predicted the general location of functional domains in EF-1 alpha which interact with GTP and tRNA. 相似文献
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Apis mellifera cytoplasmic elongation factor 1 alpha (EF-1 alpha) is closely related to Drosophila melanogaster EF-1 alpha 总被引:2,自引:0,他引:2
Using low stringency hybridisation with a Drosophila melanogaster EF-1 alpha gene fragment we have isolated a genomic DNA clone encoding elongation factor 1 alpha (EF-1 alpha) from Apis mellifera. The hybridising Apis mellifera sequence could be delineated to two small EcoRI fragments that were also revealed by genomic Southern hybridisation. By comparison with the corresponding Drosophila melanogaster data the complete translational reading frame has been deduced. It is interrupted by two intervening sequences of 220 and about 790 nucleotides. Comparison with known eucaryotic EF-1 alpha sequences further confirms that certain amino acid sequences seem to be invariable within the EF-1 alpha protein family. 相似文献
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M Sandbaken J A Lupisella B DiDomenico K Chakraburtty 《Biochimica et biophysica acta》1990,1050(1-3):230-234
The unique yeast translational factor EF-3 participates in the elongation cycle by stimulating the function of EF-1 alpha in binding aminoacyl-tRNA to the ribosome. We have isolated the structural gene encoding EF-3 from the yeast Saccharomyces cerevisiae. The YEF3 gene is found in one copy per haploid genome and is essential for vegetative growth. DNA sequence analysis reveals that the YEF3 gene contains an open reading frame of 1044 codons. The deduced amino acid sequence has two repeats of a nucleotide-binding motif. Each of these repeats shows similarity to the nucleotide-binding motif of hydrophilic, membrane-associated ATPases including human multidrug resistant protein MDR. Factor 3 manifests ribosome-dependent ATP hydrolysis. Introduction of the YEF3 gene on a high copy number plasmid into yeast strains increases the ribosome-dependent ATPase activity and EF-3 protein levels by 3-5-fold. Yeast strains containing elevated EF-3 protein levels also exhibit increased sensitivity to the aminoglycoside antibiotics hygromycin and paromomycin. These drugs are known to increase translational errors. These observations suggest that EF-3 may affect translational accuracy. 相似文献
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Translation elongation factor-1 alpha interacts with the 3' stem-loop region of West Nile virus genomic RNA. 总被引:4,自引:0,他引:4
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The conserved 3'-terminal stem-loop (3' SL) of the West Nile virus (WNV) genomic RNA was previously used to probe for cellular proteins that may be involved in flavivirus replication and three cellular proteins were detected that specifically interact with the WNV 3' SL RNA (J. L. Blackwell and M. A. Brinton, J. Virol. 69:5650-5658, 1995). In this study, one of these cellular proteins was purified to apparent homogeneity by ammonium sulfate precipitation and liquid chromatography. Amino acid sequence Western blotting, and supershift analyses identified the cellular protein as translation elongation factor-1 alpha (EF-1 alpha). Competition gel mobility shift assays demonstrated that the interaction between EF-1 alpha and WNV 3' SL RNA was specific. Dephosphorylation of EF-1 alpha by calf intestinal alkaline phosphatase inhibited its binding to WNV 3' SL RNA. The apparent equilibrium dissociation constant for the interaction between EF-1 alpha and WNV 3' SL RNA was calculated to be 1.1 x 10(-9) M. Calculation of the stoichiometry of the interaction indicated that one molecule of EF-1 alpha binds to each molecule of WNV 3' SL RNA. Using RNase footprinting and nitrocellulose filter binding assays, we detected a high-activity binding site on the main stem of the WNV 3' SL RNA. Interaction with EF-1 alpha at the high-activity binding site was sequence specific, since nucleotide substitution in this region reduced the binding activity of the WNV 3' SL RNA for EF-1 alpha by approximately 60%. Two low-activity binding sites were also detected, and each accounted for approximately 15 to 20% of the binding activity. Intracellular association between the host protein and the viral RNA was suggested by coimmunoprecipitation of WNV genomic RNA and EF-1 alpha, using an anti-EF-1 alpha antibody. 相似文献