Cloning, sequencing and expression of a cDNA encoding mammalian valyl-tRNA synthetase.
A fragment of the cDNA encoding a rat valyl-tRNA synthetase (TrsVal)-like protein was cloned from a rat cDNA library in lambda gt11 using an oligodeoxyribonucleotide (oligo) probe. Three independent plaque clones containing the human TrsVal cDNA were then isolated from a lambda gt10 human erythroleukemia cDNA library using the rat cDNA fragment as the hybridization probe. Sequence analyses of the cDNA fragments provided a 3.2-kb sequence with an open reading frame that contained the 'HIGH' synthetase signature sequence and the tRNA 3'-end-binding motif, KMSKS, and putative Val-binding motif, EWCISRQ. The sequence was extended to the 3' end of the cDNA by the polymerase chain reaction using an internal primer and an oligo(dT) adapter. The deduced 1051-amino-acid sequence shares 65% identity with yeast TrsVal, and contains a highly basic N-terminal region, a newly evolved protease-sensitive region in sequence close to the C terminus, and several sites for protein kinase C phosphorylation. A 3-kb cDNA fragment was sub-cloned into plasmid pSVL and expressed in COS-7 cells; up to a sevenfold increase in TrsVal activity was obtained. These results confirm the cloning and sequencing of a human TrsVal-encoding cDNA.
Vilalta
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A
Department of Chemistry, Georgetown University, Washington, DC 20057.
Donovan
D
D
Wood
L
L
Vogeli
G
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Yang
D C
DC
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D003001:Cloning, Molecular; D015723:Gene Library; D006801:Humans; D008969:Molecular Sequence Data; D051381:Rats; D017386:Sequence Homology, Amino Acid; D014162:Transfection; D014637:Valine-tRNA Ligase / Q000235:genetics*
D014637:Valine-tRNA Ligase