Multiple molecular forms of cysteinyl-tRNA synthetase from rat liver: Purification and subunit structure

Pan, Foo; Lee, H.H.; Pai, S. H.; Yu, Tien‐Shin; Guoo, J. Y.; Duh, G.M.

doi:10.1016/0005-2744(76)90080-2

Cited by 21 publications

(4 citation statements)

References 26 publications

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“…A dimeric form is consistent with the previous observations that the S. cerevisiae enzyme (Motorin et al, 1997), the rat liver enzyme (Pan et al, 1976), and the two isoforms of the rabbit cytoplasmic cysteinyl-tRNA synthetase (Motorin and Waller, 1998) exist as homodimers. The full-length protein, purified after expression in E. coli, eluted at a position corresponding to a molecular weight of 250000.…”

Section: Biochemical Analysis Of the Two Isoforms Of Human Cytoplasmisupporting

confidence: 92%

Isolation of Two cDNAs Encoding Functional Human Cytoplasmic Cysteinyl-tRNA Synthetase

Davidson¹,

Caffarella²,

Vitseva³

et al. 2001

Biological Chemistry

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Cysteinyl-tRNA synthetase catalyzes the addition of cysteine to its cognate tRNA. The available eukaryotic sequences for this enzyme contain several insertions that are absent from bacterial sequences. To gain insights into the differences between the bacterial and eukaryotic forms, we previously studied the E. coli cysteinyl-tRNA synthetase. In this study, we sought to clone and express the full-length gene for the human cytoplasmic cysteinyl-tRNA synthetase. Although a gene encoding the human enzyme has been described, the predicted protein sequence, consisting of 638 amino acids, lacks homology with other eukaryotic enzymes in the carboxyl-terminus. This suggested that a further investigation was necessary to obtain the definitive sequence for the human enzyme. Here we report the isolation of a full-length cDNA that encodes a protein of 748 amino acids. The predicted protein sequence shows considerable similarity to other eukaryotic cysteinyl-tRNA synthetases in the carboxyl-terminus. We also found that approximately 20% of the mRNA encoding the cytoplasmic cysteinyl-tRNA synthetase contained an insertion of 8 bases in the 3' coding region of the mRNA. This insertion arises from an alternative splicing between the last two exons of the gene. The alternative splicing alters the reading frame and results in the replacement of the carboxy-terminal 44 amino acids with a novel sequence of 22 amino acids. Expression of the full-length and alternative forms of the enzyme in E. coli generated functional proteins that were active in aminoacylation of human cytoplasmic tRNA(Cys) with cysteine.

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Section: Biochemical Analysis Of the Two Isoforms Of Human Cytoplasmisupporting

confidence: 92%

Isolation of Two cDNAs Encoding Functional Human Cytoplasmic Cysteinyl-tRNA Synthetase

Davidson¹,

Caffarella²,

Vitseva³

et al. 2001

Biological Chemistry

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“…The homocomplex of cysteine studied by Pan et al (1976), however, is much smaller than the heterocomplexes normally observed. The valyl-tRNA synthetase complex studied by the other workers appears to involve an electrostatic interaction between tRNA and enzyme and is readily disrupted by quite low ionic strengths while the core complex normally observed is stable to much higher ionic strengths.…”

mentioning

confidence: 78%

“…Dickman & Boll (1977), for example, claim that purification methods "mild enough not to break the types of bonds expected" alter the ratio of the components and hence suggest a mixture of homocomplexes. There have been a number of reports of the existence of homocomplexes containing multiple copies of a single synthetase (Pan et al, 1976;Dietrich et al, 1978;Zaccai et al, 1979) which have sometimes been quoted as evidence for the existence of a set of inseparable homocomplexes with similar properties rather than a heterocomplex.…”

mentioning

confidence: 99%

Molecular weights of mitochondrial and cytoplasmic aminoacyl-tRNA synthetases of beef liver and their complexes

Ej¹,

Gb²,

Pd³

1983

Biochemistry

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In eukaryotes, multienzyme complexes containing five to nine aminoacyl-tRNA synthetase activities have frequently been reported. In this study, we report the existence, in bovine liver cytoplasm, of a multienzyme complex containing at least 16 activities which can be disrupted by homogenization to give rise to smaller complexes and noncomplexed synthetases. Determination of the size and component activity of these complexes and of the molecular weights of all 20 free synthetases suggests that the smaller complexes and free activities normally identified arise from the larger complex by well-defined stages during homogenization. We also show that similar, though not identical, complexes are found in bovine liver mitochondria and give the molecular weights of 16 mitochondrial synthetases.

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“…Although the revised human CRS fits with the smaller rabbit CRS, the source of the larger one has not been explained. Finally, different CRS isoforms were also identified from rat (18), despite the fact that only one CRS gene was identified (19). For these reasons, the structural gene for human CRS needs to be re-examined.…”

Section: Introductionmentioning

confidence: 99%

An elongation factor-associating domain is inserted into human cysteinyl-tRNA synthetase by alternative splicing

Kim¹,

Kim²,

Lee³

et al. 2000

Nucleic Acids Research

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The amino acid sequence of human cytoplasmic cysteinyl-tRNA synthetase (CRS) was examined by analyzing sequences of genomic and expressed sequence tag fragments. From theses analyses, a few interesting possibilities were suggested for the structure of human CRS. First, different isoforms of CRS may result from alternative splicing. Second, the largest one would comprise 831 amino acids. Third, a new exon was identified encoding an 83 amino acid domain that is homologous to parts of elongation factor-1 subunits as well as other proteins involved in protein synthesis. Northern blot analysis showed three different mRNAs for CRS (of approximately 3.0, 2.7 and 2.0 kb) from human testis while only the 2.7 kb mRNA was commonly detected in other tissues. Expression of the exon 2-containing transcript in testis was confirmed by RT-PCR and northern blotting. CRS containing the exon 2-encoded peptide retained catalytic activity comparable to that lacking this peptide. This peptide was responsible for the specific interaction of CRS with elongation factor-1gamma.

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Multiple molecular forms of cysteinyl-tRNA synthetase from rat liver: Purification and subunit structure

Cited by 21 publications

References 26 publications

Isolation of Two cDNAs Encoding Functional Human Cytoplasmic Cysteinyl-tRNA Synthetase

Isolation of Two cDNAs Encoding Functional Human Cytoplasmic Cysteinyl-tRNA Synthetase

Molecular weights of mitochondrial and cytoplasmic aminoacyl-tRNA synthetases of beef liver and their complexes

An elongation factor-associating domain is inserted into human cysteinyl-tRNA synthetase by alternative splicing

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