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Crow, Allister, Liu, Yiming, Carlsson Moller, Mirja, Le Brun, Nick 
ORCID: https://orcid.org/0000-0001-9780-4061 and Hederstedt, Lars
(2009)
Structure and functional properties of Bacillus subtilis endospore biogenesis factor StoA.
The Journal of Biological Chemistry, 284 (15).
pp. 10056-10066.
ISSN 1083-351X
Abstract
Bacillus subtilis StoA is an extracytoplasmic thiol-disulfide oxidoreductase (TDOR) important for the synthesis of the endospore peptidoglycan cortex protective layer. Here we demonstrate that StoA is membrane-associated in B. subtilis and report the crystal structure of the soluble protein lacking its membrane anchor. This showed that StoA adopts a thioredoxin-like fold with N-terminal and internal additions that are characteristic of extracytoplasmic TDORs. The CXXC active site of the crystallized protein was found to be in a mixture of oxidized and reduced states, illustrating that there is little conformational variation between redox states. The midpoint reduction potential was determined as -248 mV versus normal hydrogen electrode at pH 7 consistent with StoA fulfilling a reductive role in endospore biogenesis. pKa values of the active site cysteines, Cys-65 and Cys-68, were determined to be 5.5 and 7.8. Although Cys-68 is buried within the structure, both cysteines were found to be accessible to cysteine-specific alkylating reagents. In vivo studies of site-directed variants of StoA revealed that the active site cysteines are functionally important, as is Glu-71, which lies close to the active site and is conserved in many reducing extracytoplasmic TDORs. The structure and biophysical properties of StoA are very similar to those of ResA, a B. subtilis extracytoplasmic TDOR involved in cytochrome c maturation, raising important general questions about how these similar but non-redundant proteins achieve specificity. A detailed comparison of the two proteins demonstrates that relatively subtle differences, largely located around the active sites of the proteins, are sufficient to confer specificity.
| Item Type: | Article |
|---|---|
| Faculty \ School: | Faculty of Science > School of Chemistry (former - to 2024) |
| UEA Research Groups: | Faculty of Science > Research Groups > Biophysical Chemistry (former - to 2017) Faculty of Science > Research Groups > Centre for Ocean and Atmospheric Sciences Faculty of Science > Research Centres > Centre for Molecular and Structural Biochemistry Faculty of Science > Research Groups > Chemistry of Life Processes |
| Depositing User: | Rachel Smith |
| Date Deposited: | 23 Mar 2011 10:36 |
| Last Modified: | 24 Sep 2024 09:19 |
| URI: | https://ueaeprints.uea.ac.uk/id/eprint/26975 |
| DOI: | 10.1074/jbc.M809566200 |
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