The gene for indole-3-acetyl-L-aspartic acid hydrolase from Enterobacter agglomerans: Molecular cloning, nucleotide sequence, and expression in Escherichia coli

J. C. Chou, W. W. Mulbry, J. D. Cohen

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34 Scopus citations

Abstract

A 5.5-kb DNA fragment containing the indole-3-acetyl-aspartic acid (IAA-asp) hydrolase gene (iaaspH) was isolated from Enterobacter agglomerans strain GK12 using a hybridization probe based on the N-terminal amino acid sequence of the protein. The DNA sequence of a 2.4-kb region of this fragment was determined and revealed a 1311-nucleotide ORF large enough to encode the 45-kDa IAA-asp hydrolase. A 1.5-kb DNA fragment containing iaaspH was subcloned into the Escherichia coli expression plasmid pTTQ8 to yield plasmid pJCC2. Extracts of IPTG-induced E. coli cultures containing the pJCC2 recombinant plasmid showed IAA-asp hydrolase levels 5 to 10-fold higher than those in E. agglomerans extracts. Homology searches revealed that the IAA-asp hydrolase was similar to a variety of amidohydrolases. In addition, IAA-asp hydrolase showed 70% sequence identity to a putative thermostable carboxypeptidase of E. coli.

Original languageEnglish (US)
Pages (from-to)172-178
Number of pages7
JournalMolecular and General Genetics
Volume259
Issue number2
DOIs
StatePublished - 1998

Bibliographical note

Funding Information:
Acknowledgements This work was supported by grant DE-A102-94ER20153 from the US Department of Energy.

Keywords

  • Amidohydrolase carboxypeptidase
  • Enterobacter
  • Hydrolase
  • Indole-3-acetylaspartic acid

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