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Identification and structure of the anti-sigma factor-binding domain of the Disulphide-stress regulated Sigma Factor sR from Streptomyces coelicolor
journal contribution
posted on 2023-06-08, 07:14 authored by Wei Li, Clare E M Stevenson, Nicolas Burton, Piotr Jakimowicz, Mark PagetMark Paget, Mark J Buttner, David M Lawson, Colin KleanthousThe extracytoplasmic function (ECF) sigma factor sR is a global regulator of redox homeostasis in the antibiotic-producing bacterium Streptomyces coelicolor, with a similar role in other actinomycetes such as Mycobacterium tuberculosis. Normally maintained in an inactive state by its bound anti-sigma factor RsrA, sR dissociates in response to intracellular disulphide-stress to direct core RNA polymerase to transcribe genes, such as trxBA and trxC that encode the enzymes of the thioredoxin disulphide reductase pathway, that re-establish redox homeostasis. Little is known about where RsrA binds on sR or how it suppresses sR-dependent transcriptional activity. Using a combination of proteolysis, surface-enhanced laser desorption ionisation mass spectrometry and pull-down assays we identify an N-terminal, 10 kDa domain (sRN) that encompasses region 2 of sR that represents the major RsrA binding site. We show that sRN inhibits transcription by an unrelated sigma factor and that this inhibition is relieved by RsrA binding, reaffirming that region 2 is involved in binding to core RNA polymerase but also demonstrating that the likely mechanism by which RsrA inhibits sR activity is by blocking this association. We also report the 2.4 Å resolution crystal structure of sRN that reveals extensive structural conservation with the equivalent region of s70 from Escherichia coli as well as with the cyclin-box, a domain-fold found in the eukaryotic proteins TFIIB and cyclin A. sRN has a propensity to aggregate, due to steric complementarity of oppositely charged surfaces on the domain, but this is inhibited by RsrA, an observation that suggests a possible mode of action for RsrA which we compare to other well-studied sigma factor-anti-sigma factor systems.
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Publication status
- Published
Journal
Journal of Molecular BiologyISSN
0022-2836External DOI
Issue
2Volume
323Page range
225-236Department affiliated with
- Biochemistry Publications
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- No
Peer reviewed?
- Yes
Legacy Posted Date
2012-02-06Usage metrics
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