Bioscience, Biotechnology, and Biochemistry
Online ISSN : 1347-6947
Print ISSN : 0916-8451
Cloning, Purification, and Properties of a Cofactor-independent Glutamate Racemase from Lactobacillus brevis ATCC 8287
Makoto YagasakiKazuhisa IwataShuichi IshinoMasaki AzumaAkio Ozaki
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JOURNAL FREE ACCESS

1995 Volume 59 Issue 4 Pages 610-614

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Abstract

A glutamate racemase gene of Lactobacillus brevis ATCC 8287 was cloned into Escherichia coli TM93 by the phenotypic complementation of a phosphoenolpyruvate carboxylase deficiency on minimum agar medium containing D-glutamate. The gene was localized to a 1.4-kb HindIII-EcoRI DNA fragment and the total nucleotide sequence of the fragment was analyzed. The gene has typical promoter and SD sequences which appeared to function in E. coli. The deduced amino acid sequence of the enzyme had 276 amino acids and the molecular weight was calculated as 29, 426. Two cysteine residues and their surrounding regions of the enzyme are homologous to those of other cofactor-independent racemases. The glutamate racemase was puritied from recombinant E. coli to homogeneity and characterized. The enzyme required no cofactors for the activity, and retained its activity even in 2M (300g/l) L-glutamate.

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