Crystallization of haloalkane dehologenase DgaA isolated from Glaciecola Agarilytica NO2

D. Malakhova1,2, Iu. Iermak1,3, R. Chaloupkova4, I. Kuta Smatanova1,3, D. Stys1

1University of South Bohemia, Faculty of Science, Branišovska 31, 370 05 České Budějovice, Czech Republic

2University of South Bohemia, Faculty of Fisheries and Protection of Waters, Institute of Complex Systems, Zámek, 136, 373 33 Nové Hrady, Czech Republic

3Center of Nanobiology and Structural Biology, Institute of Microbiology, Academy of Sciences of the Czech Republic, Zámek 136, 37333 Nové Hrady, Czech Republic

4Loschmidt laboratories, Department of Experimental Biology and Research Centre for Toxic Compounds in the Environment, RECETOX, Masaryk University, 625 00, Brno, Czech Republic

dmalakhova@frov.jcu.cz

A novel haloalkane dehalogenase DgaA, isolated from Glaciecola Agarilytica NO2 was successfully crystallized.

Haloalkane dehalogenases represent an important class of microbial enzymes that catalyse an irreversible hydrolysis of halogenated compounds to the corresponding alcohol, halide ion and proton [1]. This enzymes are widely used for biodegradation and biosensing of environmental pollutants, decontamination of warfare agents, recycling by-products from chemical processes [2].

DgaA was isolated from psychrophilic and moderate halophillic organism Glaciecola Agarilytica NO2, found in the East Sea marine sediment in Korea [3].

In order to obtain the  structure of DgaA x-ray diffraction analysis of enzyme's crystals was employed. Initial screening for crystallization conditions has been performed on Gryphon crystallization robot (Art Robbins Instruments, USA) using sitting drop vapour diffusion method [4]. After initial crystallization, microcrystals were found in several conditions of JCSG - plus and Structure screen 1&2 (Molecular Dimensions Ltd, UK) commercial screens. Optimization procedures including protein and salt concentration variations resulted in appearance of crystals sufficient for x-ray diffraction experiments. Needle clusters and plate-like crystals with size about 1.3 × 0.059 mm and 0.2 × 0.065 mm, respectively, were obtained. The datasets were collected at resolution ranging from 1.8 to 2.4 Å.

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2. T. Koudelakova, S. Bidmanova, P. Dvorak, A. Pavelka, R. Chaloupkova, Z. Prokop, J. Damborsky, Biotechnology journal, 8, I.1, (2013), pp. 32-45.

3. J. J. Yong, S. J. Park, H. J. Kim, S. K. Rhee, International Journal of Systematic and Evolutionary Microbiology, 57, (2007), pp. 951-953.

4. A.  Ducruix, R. Giege, Crystallization of Nucleic Acids and Proteins. A Practical Approach, 2nd ed., Oxford University Press (2013).

This work was partially supported by the Ministry of Education, Youth and Sport of the Czech Republic – projects CENAKVA (No. CZ 1. 05/2.1. 00/01.0024) and CENAKVA II (No. LO1205 under NPU I program) and GAJU grant (134/2013/ Z 2014 FUUP), and by the GACR 207/12/0775.