Haloalkane dehalogenases make up an important class of hydrolytic enzymes which catalyse the cleavage of carbon-halogen bonds in halogenated aliphatic compounds. There is growing interest in these enzymes owing to their potential use in environmental and industrial applications. The haloalkane dehalogenase DhaA from Rhodococcus rhodochrous NCIMB 13064 can slowly detoxify the industrial pollutant 1,2,3-trichloropropane (TCP). Structural analysis of this enzyme complexed with target ligands was conducted in order to obtain detailed information about the structural limitations of its catalytic properties. In this study, the crystallization and preliminary X-ray analysis of complexes of wild-type DhaA with 2-propanol and with TCP and of complexes of the catalytically inactive variant DhaA13 with the dye coumarin and with TCP are described. The crystals of wild-type DhaA were plate-shaped and belonged to the triclinic space group P1, while the variant DhaA13 can form prism-shaped crystals belonging to the orthorhombic space group P2(1)2(1)2(1) as well as plate-shaped crystals belonging to the triclinic space group P1. Diffraction data for crystals of wild-type DhaA grown from crystallization solutions with different concentrations of 2-propanol were collected to 1.70 and 1.26 Å resolution, respectively. A prism-shaped crystal of DhaA13 complexed with TCP and a plate-shaped crystal of the same variant complexed with the dye coumarin diffracted X-rays to 1.60 and 1.33 Å resolution, respectively. A crystal of wild-type DhaA and a plate-shaped crystal of DhaA13, both complexed with TCP, diffracted to atomic resolutions of 1.04 and 0.97 Å, respectively.

Institute of Physical Biology University of South Bohemia Ceske Budejovice Nove Hrady Czech Republic

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Bergfors, T. M. (2003). J. Struct. Biol. 142, 66–76. PubMed

Bidmanova, S., Chaloupkova, R., Damborsky, J. & Prokop, Z. (2010). Anal. Bioanal. Chem. 398, 1891–1898. PubMed

Bradford, M. M. (1976). Anal. Biochem. 72, 248–254. PubMed

Campbell, D. W., Muller, C. & Reardon, K. F. (2006). Biotechnol. Lett. 28, 883–887. PubMed

Carrea, G. & Riva, S. (2000). Angew. Chem. Int. Ed. 39, 2226–2254. PubMed

Collaborative Computational Project, Number 4 (1994). Acta Cryst. D50, 760–763. PubMed

Dauter, Z. (1999). Acta Cryst. D55, 1703–1717. PubMed

Diederichs, K. (2006). Acta Cryst. D62, 96–101. PubMed

Diederichs, K. & Karplus, P. A. (1997). Nature Struct. Biol. 4, 269–275. PubMed

Ducruix, A. & Giegé, R. (1999). Crystallization of Nucleic Acids and Proteins: A Practical Approach, 2nd ed. Oxford University Press.

Janssen, D. B. (2004). Curr. Opin. Chem. Biol. 8, 150–159. PubMed

Janssen, D. B. (2007). Adv. Appl. Microbiol. 61, 233–252. PubMed

Jesenska, A., Sykora, J., Olzynska, A., Brezovsky, J., Zdrahal, Z., Damborsky, J. & Hof, M. (2009). J. Am. Chem. Soc. 131, 494–501. PubMed

Kabsch, W. (1993). J. Appl. Cryst. 26, 795–800.

Kabsch, W. (2010a). Acta Cryst. D66, 125–132. PubMed PMC

Kabsch, W. (2010b). Acta Cryst. D66, 133–144. PubMed PMC

Klibanov, A. M. (2001). Nature (London), 409, 241–246. PubMed

Matthews, B. W. (1968). J. Mol. Biol. 33, 491–497. PubMed

Nardini, M. & Dijsktra, B. W. (1999). Curr. Opin. Struct. Biol. 9, 732–737. PubMed

Ollis, D. L., Cheah, E., Cygler, M., Dijkstra, B., Frolow, F., Franken, S. M., Harel, M., Remington, S. J., Silman, I., Schrag, J., Sussman, J. L., Verschueren, K. H. G. & Goldman, A. (1992). Protein Eng. 5, 197–211. PubMed

Otwinowski, Z. & Minor, W. (1997). Methods Enzymol. 276, 307–326. PubMed

Pavlova, M., Klvana, M., Prokop, Z., Chaloupkova, R., Banas, P., Otyepka, M., Wade, R. C., Tsuda, M., Nagata, Y. & Damborsky, J. (2009). Nature Chem. Biol. 5, 727–733. PubMed

Prokop, Z., Damborsky, J., Nagata, Y. & Janssen, D. B. (2004). Patent WO 2006/079295 A2.

Prokop, Z., Damborsky, J., Oplustil, F., Jesenska, A. & Nagata, Y. (2005). Patent WO 2006/128390 A1.

Prokop, Z., Oplustil, F., DeFrank, J. & Damborsky, J. (2006). Biotechnol. J. 1, 1370–1380. PubMed

Prokop, Z., Sato, Y., Brezovsky, J., Mozga, T., Chaloupkova, R., Koudelakova, T., Jerabek, P., Stepankova, V., Natsume, R., Leeuwen, J. G. E., Janssen, D. B., Florian, J., Nagata, Y., Senda, T. & Damborsky, J. (2010). Angew. Chem. Int. Ed. 49, 6111–6115. PubMed

Stsiapanava, A., Dohnalek, J., Gavira, J. A., Kuty, M., Koudelakova, T., Damborsky, J. & Kuta Smatanova, I. (2010). Acta Cryst. D66, 962–969. PubMed

Stsiapanava, A., Koudelakova, T., Lapkouski, M., Pavlova, M., Damborsky, J. & Kuta Smatanova, I. (2008). Acta Cryst. F64, 137–140. PubMed PMC

Stucki, G. & Thuer, M. (1995). Environ. Sci. Technol. 29, 2339–2345. PubMed

Teng, T.-Y. (1990). J. Appl. Cryst. 23, 387–391.

Vagin, A. & Teplyakov, A. (2010). Acta Cryst. D66, 22–25. PubMed

Weiss, M. S. (2001). J. Appl. Cryst. 34, 130–135.

Crystallographic analysis of new psychrophilic haloalkane dehalogenases: DpcA from Psychrobacter cryohalolentis K5 and DmxA from Marinobacter sp. ELB17