PMID:18028317
| Citation |
Kachroo, AH, Kancherla, AK, Singh, NS, Varshney, U and Mahadevan, S (2007) Mutations that alter the regulation of the chb operon of Escherichia coli allow utilization of cellobiose. Mol. Microbiol. 66:1382-95 |
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| 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. |
| Links |
PubMed Online version:10.1111/j.1365-2958.2007.05999.x |
| Keywords |
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 |
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Significance
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This work has clarified several misconceptions regarding the cryptic nature of the chb operon with respect to celloiose utilization(peviously known as the cryptic cel operon)(Parker and Hall, 1990). Cellobiose utilization did not require mutations that alter or modify the native specificities of the chb permease and β-glucosidase as proposed earlier (Keyhani and Roseman, 1997). Furthermore, it has implications on the evolution of novel metabolic functions from pre-exisitng genes or operons. This work is consistent with recent bioinformatic studies (Babu et al., 2006; Lozada-Chávez et al., 2006) which suggest that mutations in the transcription regulatory networks (TRNs) are primarily responsible for the variation in bacterial phenotypes.
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- ↑ Plumbridge, J & Pellegrini, O (2004) Expression of the chitobiose operon of Escherichia coli is regulated by three transcription factors: NagC, ChbR and CAP. Mol. Microbiol. 52 437-49 PubMed
- ↑ Keyhani, NO & Roseman, S (1997) Wild-type Escherichia coli grows on the chitin disaccharide, N,N'-diacetylchitobiose, by expressing the cel operon. Proc. Natl. Acad. Sci. U.S.A. 94 14367-71 PubMed
- ↑ Parker, LL & Hall, BG (1990) Characterization and nucleotide sequence of the cryptic cel operon of Escherichia coli K12. Genetics 124 455-71 PubMed
- ↑ Madan Babu, M et al. (2006) Evolutionary dynamics of prokaryotic transcriptional regulatory networks. J. Mol. Biol. 358 614-33 PubMed
- ↑ Lozada-Chávez, I et al. (2006) Bacterial regulatory networks are extremely flexible in evolution. Nucleic Acids Res. 34 3434-45 PubMed
