Analysis of DNA Binding and Transcriptional Activation by the LysR-Type Transcriptional Regulator CbbR of Xanthobacter flavus
AUTOR(ES)
van Keulen, Geertje
FONTE
American Society for Microbiology
RESUMO
The LysR-type transcriptional regulator CbbR controls the expression of the cbb and gap-pgk operons in Xanthobacter flavus, which encode the majority of the enzymes of the Calvin cycle required for autotrophic CO2 fixation. The cbb operon promoter of this chemoautotrophic bacterium contains three potential CbbR binding sites, two of which partially overlap. Site-directed mutagenesis and subsequent analysis of DNA binding by CbbR and cbb promoter activity were used to show that the potential CbbR binding sequences are functional. Inverted repeat IR1 is a high-affinity CbbR binding site. The main function of this repeat is to recruit CbbR to the cbb operon promoter. In addition, it is required for negative autoregulation of cbbR expression. IR3 represents the main low-affinity binding site of CbbR. Binding to IR3 occurs in a cooperative manner, since mutations preventing the binding of CbbR to IR1 also prevent binding to the low-affinity site. Although mutations in IR3 have a negative effect on the binding of CbbR to this site, they result in an increased promoter activity. This is most likely due to steric hindrance of RNA polymerase by CbbR since IR3 partially overlaps with the −35 region of the cbb operon promoter. Mutations in IR2 do not affect the DNA binding of CbbR in vitro but have a severe negative effect on the activity of the cbb operon promoter. This IR2 binding site is therefore critical for transcriptional activation by CbbR.
ACESSO AO ARTIGO
http://www.pubmedcentral.nih.gov/articlerender.fcgi?artid=142840Documentos Relacionados
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