Purification and Characterization of Haloalkane Dehalogenase of a New Substrate Class from a g-Hexachlorocyclohexane-Degrading Bacterium, Sphingomonas paucimobilis UT26

Authors

Nagata, Y., Miyauchi, K., Damborsky, J., Manova, K., Ansorgova, A. and Takagi, M.

Source

APPLIED AND ENVIRONMENTAL MICROBIOLOGY 63: 3707-3710 (1997)

Abstract

The linB gene product (LinB), 1,3,4,6-tetrachloro-1,4-cyclohexadiene halidohydrolase, which is involved in the degradation of gamma-hexachlorocyclohexane in Sphingomonas paucimobilis UT26 (Y. Nagata, T. Nariya, R. Ohtomo, M. Fukuda, K. Yano, and M. Takagi, J. Bacteriol. 175:6403-6410, 1993), was overproduced in E. coli and purified to homogeneity. The molecular mass of LinB was deduced to be 30 kDa by gel filtration chromatography and 32 kDa by electrophoresis on sodium dodecyl sulfate-polyacrylamide gel, indicating that LinB is a monomeric enzyme. The optimal pH for activity was 8.2. Not only monochloroalkanes (C-3 to C-10) but also dichloroalkanes, bromoalkanes, and chlorinated aliphatic alcohols were good substrates for LinB, suggesting that LinB is a haloalkane dehalogenase with a broad range of substrate specificity. These results indicate that LinB shares properties with another haloalkane dehalogenase, DhlA (S. Keuning, D. B. Janssen, and B. Witholt, J. Bacteriol. 163:635-639, 1985), which shows significant similarity to LinB in primary structure (D. B. Janssen, F. Pries, J. van der Ploeg, B. Kazemier, P. Terpstra, and B. Witholt, J. Bacteriol. 171:6791-6799, 1989) but not in substrate specificity. Principal component analysis of substrate activities of various haloalkane dehalogenases suggested that LinB probably constitutes a new substrate specificity class within this group of enzymes.

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Citation

Nagata, Y., Miyauchi, K., Damborsky, J., Manova, K., Ansorgova, A. and Takagi, M., 1997: Purification and Characterization of Haloalkane Dehalogenase of a New Substrate Class from a g-Hexachlorocyclohexane-Degrading Bacterium, Sphingomonas paucimobilis UT26. Applied and Environmental Microbiology 63: 3707-3710.

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