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An intramolecular disulfide bond is required for the thermostability of methyl parathion hydrolase, OPHC2

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Abstract

OPHC2, a methyl parathion hydrolase (MPH) from Pseudomonas pseudoalcaligenes C2-1 (CGMCC 1150), can degrade a wide range of organophosphate pesticides. Compared with other MPHs, OPHC2 exhibits high thermostability. Its thermostability mechanism, however, remains unknown. In the present study, sequence analysis demonstrated that two cysteines (Cys110 and Cys146) exist in OPHC2, but not in other MPHs. The three-dimensional structural model of OPHC2 performed by computer-assisted homology modelling revealed a potential stacking network with residues Cys110 and Cys146, which probably formed an intramolecular disulfide bond. Furthermore, both sodium dodecyl sulphate-polyacrylamide gel electrophoresis and thiol-titration analyses indicated that OPHC2 contains a disulfide bond. Substitution of the disulfide bond-forming cysteines with alanine, leucine or methionine residues substantially decreased the thermostability of OPHC2, suggesting that disulfide bond formation affects conformational stability. These results, combined with three-dimensional structural modelling, demonstrated that the formation of a C110-C146 disulfide bond may stabilise the conformation of OPHC2, contributing to its thermostability.

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Acknowledgements

We thank Dr. Min Lin for critical reading of this manuscript. This work was supported by the Ministry of Science and Technology of the People's Republic of China (National “863” Project No. 2007AA100605).

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Authors and Affiliations

  1. Biotechnology Research Institute, Chinese Academy of Agricultural Sciences, No. 12 Zhongguancun South Street, Beijing, 100081, China

    Xiao-yu Chu, Jian Tian, Ning-feng Wu & Yun-liu Fan

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  1. Xiao-yu Chu
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  2. Jian Tian
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  3. Ning-feng Wu
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  4. Yun-liu Fan
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Correspondence to Ning-feng Wu.

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Chu, Xy., Tian, J., Wu, Nf. et al. An intramolecular disulfide bond is required for the thermostability of methyl parathion hydrolase, OPHC2. Appl Microbiol Biotechnol 88, 125–131 (2010). https://doi.org/10.1007/s00253-010-2738-5

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  • DOI: https://doi.org/10.1007/s00253-010-2738-5

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