Isolation and crystallization of the recombinant PsbR protein of higher-plant photosystem II

T. Holubeva^1,2, M. Plevaka^1,2, J.Heller^1,2, J. Ludwig³, O. Sedo⁴, Z. Zdrahal⁴, I. Kuta Smatanova^1,5 and J. Kohoutova¹

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

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

³University of Bonn, IZMB / Molekulare Bioenergetik, Kirschallee 1, D-53115 Bonn, Germany

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

⁵Academy of Sciences of the Czech Republic, Institute of Nanobiology and Structural Biology

GCRC, Zamek 136, 373 33 Nove Hrady

Photosystem II consists of more than 20 proteins, which catalyze the light-driven oxidation of water to molecular oxygen in cyanobacteria, algae, and plants. PSII can be divided into two functional domains: the electron transfer domain, comprising the integral membrane helices and cofactors and the oxygen evolving complex (OEC), located on the lumenal face of the complex including the extrinsic proteins PsbO (33 kDa), PsbP (23 kDa), PsbQ (17 kDa) and PsbR (10 kDa), the loop regions of several membrane proteins and the catalytic inorganic manganese, calcium, and chloride cluster [1].

PsbR is a low-molecular weight protein, which is found in higher-plant photosystem II and anticipated to play a role in water oxidation, yet the physiological significance, exact location and structure of the purified protein has remained obscure [1,2].

Therefore, further biochemical studies should elucidate the position of PsbR relative to other components of plant OEC. As primary structure of spinach PsbR protein is known, we were able to produce psbR gene from mRNA isolated from Spinacia oleracea, create an expression vector with its anchor as HisPsbR and without anchor as PsbR using molecular biology techniques. The further goal is to optimize purification conditions in order to get stable proteins that will be used for structural analysis.

This research was supported by the ME CR (COST LD11011, CZ.1.05/2.1.00/01.0024), by the AS CR (AV0Z60870520) and GAJU 170/2010/P.

1.Roose, J. L. et al., (2007). Photosynthesis Research 92, 369-387.

2.Suorsa, M. et al., (2006). Journal of Biological Chemistry 281, 145-150.