Crystallization and preliminary X-ray diffraction analysis of new psychrophilic haloalkane dehalogenases DmxA and DpcA and DpcA complex with 1-bromohexane

Katsiaryna Tratsiak^1,2, Radka Chaloupkova³, Pavlina Rezacova⁴, Michal Kuty^1,5, Jiri Damborsky³, Ivana Kuta Smatanova^1,5

¹University of South Bohemia in Ceske Budejovice, Faculty of Fisheries and Protection of Waters, South Bohemian Research Center of Aquaculture and Biodiversity of Hydrocenoses and School of complex systems, Zamek 136, 373 33 Nove Hrady, Czech Republic.

²University of South Bohemia in Ceske Budejovice, Faculty of Science, Branisovska 31, 37005 Ceske Budejovice, Czech Republic.

³Loschmidt Laboratories, Department of Experimental Biology and Research Centre for Toxic Compounds in the Environment, Faculty of Science, Masaryk University, Kamenice 5/A4, 62500 Brno, Czech Republi).

⁴Institute of Organic Chemistry and Biochemistry, Flemingovo nam. 2, 166 10 Prague 6, Czech Republic.

⁵Academy of Sciences of the Czech Republic, Inst. of Nanobiology and Structural Biology GCRC, Zamek 136, 373 33 Nove Hrady, Czech Republic.

Haloalkane dehalogenases (EC 3.8.1.5; HLDs) are microbial enzymes with catalytic activity for the hydrolytic conversion of xenobiotic and highly toxic halogenated aliphatic compounds to the corresponding alcohols (Janssen et al. 2005). The active site is buried inside the protein and lined with hydrophobic residues. The reaction proceeds via a covalent substrate-enzyme complex (Schanstra et al. 1997).

To date, HLD activity has been experimentally confirmed in only about a dozen different proteins (Chovancova et al. 2007). A novel HLD were found in Gram negative psychrophilic bacteria: DpcA from Psychrobacter cryohalolentis K5 and DmxA from Marinobacter sp. ELB 17.

Both haloalkane dehalogenase DpcA and DmxA were crystallized in optimal conditions and diffraction data were collected using in-house source (IOCB, Prague) as well as the synchrotron beamline (BESSY, Berlin). To understand the changes in the active site of the enzyme DpcA, the native enzyme was co-crystallized with substrate 1-Bromohexane. Structure determination is currently in progress now.

This research was supported by the GACR (P207/12/0775), ME CR (CZ.1.05/2.1.00/01.0024) and by the AS CR (AV0Z60870520).

Janssen, D. B., Dinkla, I. J. T., Poelarends, G. J. & Terpstra, P. (2005).Environ. Microbiol. 7, 1868–1882.

Schanstra, J. P., Kingma, J, Janssen. D.B, (1997), Protein Engineering, 10 (1), pp.53–61.

Chovancova, E., Kosinski, J., Bujnicki, J. M. & Damborsky, J. (2007), Proteins, 67, 305–316.