RRC ID 35194
Author Kachroo AH, Kancherla AK, Singh NS, Varshney U, Mahadevan S.
Title Mutations that alter the regulation of the chb operon of Escherichia coli allow utilization of cellobiose.
Journal Mol. Microbiol.
Abstract Wild-type strains of Escherichia coli are normally unable to metabolize cellobiose. However, cellobiose-positive (Cel(+)) mutants arise upon prolonged incubation on media containing cellobiose as the sole carbon source. We show that the Cel(+) derivatives carry two classes of mutations that act concertedly to alter the regulation of the chb operon involved in the utilization of N,N'-diacetylchitobiose. These consist of mutations that abrogate negative regulation by the repressor NagC as well as single base-pair changes in the transcriptional regulator chbR that translate into single-amino-acid substitutions. Introduction of chbR from two Cel(+) mutants resulted in activation of transcription from the chb promoter at a higher level in the presence of cellobiose, in reporter strains carrying disruptions of the chromosomal chbR and nagC. These transformants also showed a Cel(+) phenotype on MacConkey cellobiose medium, suggesting that the wild-type permease and phospho-beta-glucosidase, upon induction, could recognize, transport and cleave cellobiose respectively. This was confirmed by expressing the wild-type genes encoding the permease and phospho-beta-glucosidase under a heterologous promoter. Biochemical characterization of one of the chbR mutants, chbRN238S, showed that the mutant regulator makes stronger contact with the target DNA sequence within the chb promoter and has enhanced recognition of cellobiose 6-phosphate as an inducer compared with the wild-type regulator.
Volume 66(6)
Pages 1382-95
Published 2007-12
DOI 10.1111/j.1365-2958.2007.05999.x
PII MMI5999
PMID 18028317
MeSH Base Sequence Cellobiose / metabolism* Circular Dichroism Disaccharides / metabolism Electrophoretic Mobility Shift Assay Escherichia coli / genetics* Escherichia coli / metabolism* Escherichia coli Proteins / genetics Escherichia coli Proteins / metabolism Gene Expression Regulation, Bacterial Models, Biological Models, Genetic Mutagenesis, Insertional Mutation Operon / genetics* Promoter Regions, Genetic / genetics Repressor Proteins / genetics Repressor Proteins / metabolism Sequence Analysis, DNA Sequence Deletion Transcription Factors / genetics Transcription Factors / metabolism
IF 3.816
Times Cited 20
WOS Category MICROBIOLOGY BIOCHEMISTRY & MOLECULAR BIOLOGY
Resource
Prokaryotes E. coli ME8333(JF496)