Co-production of two recombinant proteins in Escherichia coli using the promoter region of the divergently expressed curli genes.
Journal - Journal of biotechnology
Expression of multiple proteins in a single host is desirable in biotechnological processes. The curli intergenic region in Escherichia coli contains promoter elements for the expression of the divergent csgBAC and csgDEFG operons. Using this bidirectionally active promoter region, we demonstrate high level production of two different recombinant proteins. The curli intergenic region may thus be used to produce multiple enzymes involved in biosynthetic pathways and biotransformations in a cost-effective manner.Copyright © 2012 Elsevier B.V. All rights reserved.
Biofilm formation in Escherichia coli cra mutants is impaired due to down-regulation of curli biosynthesis.
Journal - Archives of microbiology (Germany )
Cra is a pleiotropic regulatory protein that controls carbon and energy flux in enteric bacteria. Recent studies have shown that Cra also regulates other cell processes and influences biofilm formation. The purpose of the present study was to investigate the role of Cra in biofilm formation in Escherichia coli. Congo red-binding studies suggested that curli biosynthesis is impaired in cra mutants. Microarray analysis of wild-type and mutant E. coli cultivated in conditions promoting biofilm formation revealed that the curli biosynthesis genes, csgBAC and csgDEFG, are poorly expressed in the mutant, suggesting that transcription of genes required for curli production is regulated by Cra. Four putative Cra-binding sites were identified in the curli intergenic region, which were experimentally validated by performing electromobility shift assays. Site-directed mutagenesis of three Cra-binding sites in the promoter region of the csgDEFG operon suggests that Cra activates transcription of this operon upon binding to operator regions both downstream and upstream of the transcription start site. Based on the Cra-binding sites identified in this and other studies, the Cra consensus sequence is refined.