Isolation, overexpression, and biochemical characterization of the two isoforms of glutamic acid decarboxylase from Escherichia coli.

Abstract:

:Escherichia coli glutamic acid decarboxylase is a pyridoxal phosphate-dependent enzyme that catalyzes the alpha-decarboxylation of glutamate to yield 4-amino-butyrate and CO2. The E. coli chromosome contains two genes encoding for this enzyme, gadA and gadB, which map at distinct loci. Their protein products differ in only five amino acid residues, four of which are located in the N-terminal region (Smith et al., 1992, J. Bacteriol. 174, 5820-5826). Herein, we report the sequences of the two gad genes, including their regulatory regions. Both genes were separately cloned into the vector pQE60, for overexpression under the control of the lac promoter. In this way, we have succeded in separately expression large quantities of each pure isoform. The two isoforms were characterized biochemically and all evidence, including that from analysis of the complex pre-steady-state kinetic behavior of the enzymes, indicates that the functional properties of the two isoenzymes are identical.

journal_name

Protein Expr Purif

authors

De Biase D,Tramonti A,John RA,Bossa F

doi

10.1006/prep.1996.0121

subject

Has Abstract

pub_date

1996-12-01 00:00:00

pages

430-8

issue

4

eissn

1046-5928

issn

1096-0279

pii

S1046592896901217

journal_volume

8

pub_type

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