Structure and Functional Properties of Bacillus subtilis Endospore Biogenesis Factor StoA

Crow, Allister, Liu, Y., Moller, M. C., Le Brun, Nick and Hederstedt, L. (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

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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
Related URLs:
Depositing User: Rachel Smith
Date Deposited: 23 Mar 2011 10:36
Last Modified: 18 Sep 2018 15:30
URI: https://ueaeprints.uea.ac.uk/id/eprint/26975
DOI: 10.1074/jbc.M809566200

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