Analysis and toxic overexpression in Escherichia coli of a staphylococcal gene encoding isoleucyl-tRNA synthetase

Gene. 1994 Apr 8;141(1):103-8. doi: 10.1016/0378-1119(94)90135-x.

Abstract

We have cloned and sequenced the Staphylococcus aureus Oxford ileS gene which encodes isoleucyl-tRNA synthetase (Ile-RS), the target for the antibiotic mupirocin. The gene was identified by hybridisation to oligodeoxyribonucleotide probes derived from internal Ile-RS amino acid (aa) sequences. The 2754-bp open reading frame encodes a 918-aa protein of 105 kDa which is homologous to other known Ile-RS from Gram- bacteria, archaebacteria, yeast and protozoa. Motifs which have been implicated in the functioning of the active site are strongly conserved. The gene was engineered for high-level expression in Escherichia coli. Ile-RS overproduction was toxic to the E. coli host, the magnitude of its observed effects being strain-dependent.

MeSH terms

  • Amino Acid Sequence
  • Base Sequence
  • Cloning, Molecular
  • Conserved Sequence
  • DNA Probes / chemical synthesis
  • DNA, Bacterial / isolation & purification
  • Escherichia coli / growth & development
  • Gene Expression Regulation, Bacterial / genetics
  • Gene Expression Regulation, Enzymologic / genetics*
  • Genes, Bacterial / genetics*
  • Genetic Complementation Test
  • Genomic Library
  • Isoleucine-tRNA Ligase / biosynthesis
  • Isoleucine-tRNA Ligase / genetics*
  • Isoleucine-tRNA Ligase / isolation & purification
  • Molecular Sequence Data
  • Operon / genetics
  • Sequence Alignment
  • Sequence Analysis, DNA
  • Staphylococcus aureus / enzymology*
  • Staphylococcus aureus / genetics

Substances

  • DNA Probes
  • DNA, Bacterial
  • Isoleucine-tRNA Ligase

Associated data

  • GENBANK/A01184
  • GENBANK/M30942
  • GENBANK/M59245
  • GENBANK/X74219