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The Role of Zinc in the Disulfide Stress-Regulated Anti-Sigma Factor RsrA from Streptomyces coelicolor
journal contribution
posted on 2023-06-08, 00:11 authored by Wei Li, Andrew R Bottrill, Maureen J Bibb, Mark J Buttner, Mark PagetMark Paget, Colin KleanthousThe regulation of disulphide stress in actinomycetes such as Streptomyces coelicolor is known to involve the zinc-containing anti-sigma factor RsrA that binds and inactivates the redox-regulated sigma factor sR. However, it is not known how RsrA senses disulphide stress nor what role the metal ion plays. Using in vitro assays, we show that while zinc is not required for sR binding it is required for functional anti-sigma factor activity, and that it plays a critical role in modulating the reactivity of RsrA cysteine thiol groups towards oxidation. Apo-RsrA is easily oxidised and, while the Zn-bound form is relatively resistant, the metal ion is readily expelled when the protein is treated with strong oxidants such as diamide. We also show, using a combination of proteolysis and mass spectrometry, that the first critical disulphide to form in RsrA involves Cys11 and one of either Cys41 or Cys44, all previously implicated in metal binding. Circular dichroism spectroscopy was used to follow structural changes during oxidation of RsrA, which indicated that concomitant with formation of this critical disulphide bond is a major restructuring of the protein where its a-helical content increases. Our data demonstrate that RsrA can only bind sR in the reduced state and that this state is stabilised by zinc. Redox stress induces disulphide bond formation amongst zinc-ligating residues, expelling the metal ion and stabilising a structure incapable of binding the sigma factor.
History
Publication status
- Published
Journal
Journal of Molecular BiologyISSN
0022-2836External DOI
Issue
2Volume
333Page range
461-472Department affiliated with
- Biochemistry Publications
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- No
Peer reviewed?
- Yes
Legacy Posted Date
2012-02-06Usage metrics
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