Isolation and crystallization of plant PSII

Jaroslava Kohoutová1,2 Magda Vojtová 1,2, EstelaPineda Molina3, Zbyněk Zdrahal4, Ondřej Šedo4, David Potěšil4, Jose A. Gavira3,,Tatyana Prudnikova1,2and Ivana Kutá Smatanová 1,2

1 University of South Bohemia in České Budějovice, 2 University of South Bohemia in České Budějovice, Faculty of Science, Branišovská 31, 37005 České Budějovice

2Academy of Sciences of the Czech Republic Center of Nanobiology and Structural Biology, Institute of Microbiology , Zámek 136, 373 33 Nové Hrady

3Laboratorio de Estudios Cristalográficos, Edf. López Neyra, P.T. Ciencias de la Salud, Avenida del Conocimiento s/n, 18100 Armilla (Granada) Spain

4Core Facility – Proteomics, CEITEC, Masaryk University, Kamenice 5,625 00 Brno

Photosystem II (PSII) is the heart of photosynthetic process. This multisubunit complex is embedded in the thylakoid membrane of plants, algae and cyanobacteria. The function of the photosystem II in different organisms is identical while the composition of subunits is different [1]. Recently the 3D X-ray structures of cyanobacterial PSII were determined to the maximum resolution of 1.9 Ǻ [2]. Compared to plant PSII cyanobacterial PSII consists of different extrinsic proteins and also the light-harvesting complex is not bound in thylakoid membrane to PSII core as it is in plant PSII.

No crystal structure of PSII from higher plants is available until now and to get structural data of PSII, crystallization of the PSII from higher plants employing several crystallization techniques and testing the role of different additives were reported. After testing of several crystallization conditions and techniques, the 3D crystals of PSII were obtained from PSII complexes isolated from Pisumsativum at the resolution of about 10.0 Ǻ [3], from Nicotiana tabaccum at 7.0 Ǻ [4] and from Spinacia oleracea at 6.5 Ǻ [5].We report here new optimized protocol for isolation of plant PSII with reproducible crystallization conditions and using of new crystallization techniques and testing methods.Obtained green crystals were analyzed by LC/MS and content of selected proteins from PSII was confirmed.

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3.         Kuta-Smatanova et al., Photosynth.Res. 2006, 90, 255-259.;

4.         Piano et al., Photosynth. Res. 2010, 106, , 221-226.;

5.         Adir, N., Acta Cryst. 1999, D55, 891–894.