Structure and Functional Properties of Bacillus subtilis Endospore Biogenesis Factor StoA
(2009) In Journal of Biological Chemistry 284(15). p.10056-10066- 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... (More)
- 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. (Less)
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https://lup.lub.lu.se/record/1401040
- author
- Crow, Allister ; Liu, Yiming LU ; Carlsson Möller, Mirja LU ; Le Brun, Nick E. and Hederstedt, Lars LU
- organization
- publishing date
- 2009
- type
- Contribution to journal
- publication status
- published
- subject
- in
- Journal of Biological Chemistry
- volume
- 284
- issue
- 15
- pages
- 10056 - 10066
- publisher
- American Society for Biochemistry and Molecular Biology
- external identifiers
-
- wos:000264892900048
- scopus:65649116921
- ISSN
- 1083-351X
- DOI
- 10.1074/jbc.M809566200
- language
- English
- LU publication?
- yes
- id
- 3448aa81-8db7-4925-ac88-28c783a53ad1 (old id 1401040)
- date added to LUP
- 2016-04-01 11:50:59
- date last changed
- 2022-01-26 19:09:06
@article{3448aa81-8db7-4925-ac88-28c783a53ad1,
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.}},
author = {{Crow, Allister and Liu, Yiming and Carlsson Möller, Mirja and Le Brun, Nick E. and Hederstedt, Lars}},
issn = {{1083-351X}},
language = {{eng}},
number = {{15}},
pages = {{10056--10066}},
publisher = {{American Society for Biochemistry and Molecular Biology}},
series = {{Journal of Biological Chemistry}},
title = {{Structure and Functional Properties of <em>Bacillus subtilis</em> Endospore Biogenesis Factor StoA}},
url = {{http://dx.doi.org/10.1074/jbc.M809566200}},
doi = {{10.1074/jbc.M809566200}},
volume = {{284}},
year = {{2009}},
}