Molecular cloning of Escherichia coli K-12 ggt and rapid isolation of gamma-glutamyltranspeptidase

Biochem Biophys Res Commun. 1988 Jan 15;150(1):33-8. doi: 10.1016/0006-291x(88)90482-2.

Abstract

Based on the results of mapping of ggt, eight strains were selected from a gene library of E. coli. One of the strains harboring pLC9-12 was found to show 14 times higher gamma-glutamyltranspeptidase activity per cell than the wild type strain. The ggt was subcloned to the BamHI site of pUC18 and the recombinant plasmid pSH101 was obtained. Ggt- phenotype of gamma-glutamyltranspeptidase-deficient mutants was complemented by pSH101. The specific activity of the enzyme in cells harboring pSH101 was 37-fold higher than that in the wild type cells. gamma-Glutamyltranspeptidase was isolated from the periplasmic fraction of the cells by simple two steps and crystallized.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Cloning, Molecular*
  • DNA Restriction Enzymes
  • DNA, Bacterial / genetics
  • DNA, Recombinant
  • Electrophoresis, Polyacrylamide Gel
  • Escherichia coli / enzymology*
  • Escherichia coli / genetics
  • Molecular Weight
  • Plasmids
  • Transformation, Bacterial
  • gamma-Glutamyltransferase / biosynthesis
  • gamma-Glutamyltransferase / genetics*
  • gamma-Glutamyltransferase / isolation & purification

Substances

  • DNA, Bacterial
  • DNA, Recombinant
  • gamma-Glutamyltransferase
  • DNA Restriction Enzymes