Cloning and nucleotide sequence determination of the isopenicillin N synthetase gene from Streptomyces clavuligerus

Brenda K. Leskiw, Yair Aharonowitz, Moshe Mevarech, Saul Wolfe, Leo C. Vining, Donald W.S. Westlake, Susan E. Jensen*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

Abstract

The isopenicillin N synthetase (IPNS) gene from Streptomyces clavuligerus was isolated from an Escherichia coli plasmid library of S. clavuligerus genomic DNA fragments using a 44-mer mixed oligodeoxynucleotide probe. The nucleotide sequence of a 3-kb region of the cloned fragment from the plasmid, pBLl, was determined and analysis of the sequence showed an open reading frame that could encode a protein of 329 amino acids with an Mr of 36 917. When the S. clavuligerus DNA from pBLl was introduced into an IPNS-deficient mutant of S. clavuligerus on the Streptomyces vector pIJ941, the recombinant plasmid was able to complement the mutation and restore IPNS activity. The protein coding region of the S. clavuligerus IPNS gene shows about 63% and 62% similarity to the Cephalosporium acremonium and Penicillium chrysogenum IPNS nucleotide sequences, respectively, and the predicted amino acid sequence of the encoded protein showed about 56% similarity to both fungal sequences.

Original languageEnglish
Pages (from-to)187-196
Number of pages10
JournalGene
Volume62
Issue number2
DOIs
StatePublished - 29 Feb 1988

Keywords

  • Cephalosporium acremonium
  • Penicillium chrysogenum
  • Recombinant DNA
  • mutant complementation
  • pUC119 library
  • β-lactam biosynthesis

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