Isolation and characterization of the yeast aspartyl-tRNA synthetase gene.
A yeast genomic library in Escherichia coli, constructed by insertion of Sau3A restriction fragments into the hybrid Saccharomyces cerevisiae-E. coli plasmid pFL1, was screened by a radioimmunoassay (RIA) for colonies expressing yeast aspartyl-tRNA synthetase (AspRS). Four clones were isolated by this technique. Data obtained by Southern and restriction analysis of the inserts showed a common 3.8-kb BamHI restriction fragment which, when inserted into the plasmid pFL1, gave a positive RIA. Several controls showed that this 3.8-kb insert codes for the entire AspRS: (i) S. cerevisiae transformed by the PFL1 plasmid carrying the 3.8-kb fragment overproduces AspRS activity by a factor of ten compared to the wild-type yeast strain; and (ii) a new protein with electrophoretic behaviour similar to AspRS and immuno-reactive toward anti-AspRS appears in crude extracts of transformed yeast and E. coli.
D000604:Amino Acyl-tRNA Synthetases / Q000235:genetics*; D001226:Aspartate-tRNA Ligase / Q000235:genetics* / Q000378:metabolism; D004262:DNA Restriction Enzymes; D004271:DNA, Fungal / Q000302:isolation & purification; D004926:Escherichia coli / Q000235:genetics; D005796:Genes; D005800:Genes, Fungal; D007700:Kinetics; D010957:Plasmids; D012441:Saccharomyces cerevisiae / Q000201:enzymology / Q000235:genetics*
D004271:DNA, Fungal; D004262:DNA Restriction Enzymes; D000604:Amino Acyl-tRNA Synthetases; D001226:Aspartate-tRNA Ligase