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Structural basis of the sulphate starvation response in E-coli: Crystal structure and mutational analysis of the cofactor-binding domain of the cbl transcriptional regulator

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JournalJournal of Molecular Biology
DatePublished - 1 Dec 2006
Issue number3
Volume364
Number of pages14
Pages (from-to)309-322
Original languageEnglish

Abstract

Cb1 is a member of the large family of LysR-type transcriptional regulators (LTTRs) common in bacteria and found also in Archaea and algal chloroplasts. The function of Cb1 is required in Escherichia coli for expression of sulphate starvation-inducible (ssi) genes, associated with the biosynthesis of cysteine from organic sulphur sources (sulphonates). Here, we report the crystal structure of the cofactor-binding domain of Cb1 (c-Cb1) from E. coli. The overall fold of c-Cb1 is very similar to the regulatory domain (RD) of another LysR family member, CysB. The RD is composed of two subdomains enclosing a cavity, which is expected to bind effector molecules. We have constructed and analysed several full-length Cb1 variants bearing single residue substitutions in the RD that affect cofactor responses. Using in vivo and in vitro transcription assays, we demonstrate that pssuE, a Cb1 responsive promoter, is down-regulated not only by the cofactor, adenosine phosphosulphate (APS), but also by thiosulphate, and, that the same RD determinants are important for the response to both cofactors. We also demonstrate the effects of selected site-directed mutations on Cb1 oligomerization and discuss these in the context of the structure. Based on the crystal structure and molecular modelling, we propose a model for the interaction of Cb1 with adenosine phosphosulphate. (c) 2006 Elsevier Ltd. All rights reserved.

    Research areas

  • E. coli, Cbl regulator, LysR family, crystal structure, cofactor-binding, SALMONELLA-TYPHIMURIUM, CYSP PROMOTER, LYSR FAMILY, GENE-CLUSTER, PROTEIN, ACTIVATOR, SUBUNIT, SULFUR, SITE, IDENTIFICATION

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