A reporter gene vector to investigate the regulation of glutamine synthetase in Bacteroides fragilis Bf1

Abstract

The Clostridium acetobutylicum eglA gene, encoding a β-1,4-endoglucanase (EG), was shown to be a useful reporter gene for the study of gene expression in Bacteroides fragilis. The eglA reporter gene has the advantages that it can be easily identified in both Escherichia coli and B. fragilis on agar media containing carboxymethylcellulose, and EG production can be rapidly quantified in liquid medium. Since the B. fragilis glutamine synthetase (GS) is inactivated in permeabilized cells and cell extracts, the eglA reporter gene was used to study the regulation of GS production in B. fragilis. Gene fusions containing the GS glnA promoter region fused to the promoterless eglA gene showed that glnA expression was regulated by nitrogen in B. fragilis at the transcriptional level. A glnA upstream region containing a near-perfect direct repeat sequence was essential for efficient GS expression and for regulation by nitrogen.