Počet záznamů: 1  

Isolation and characterization of oxygen-evolving photosystem II particles and photosystem II core complex from the filamentous cyanobacterium Spirulina platensis

  1. 1. 0422357 - MBU-M 2014 RIV CZ eng J - Článek v odborném periodiku
    Šetlíková, Eva - Sofrová, D. - Kovář, V. - Budáč, Petr
    Isolation and characterization of oxygen-evolving photosystem II particles and photosystem II core complex from the filamentous cyanobacterium Spirulina platensis.
    Photosynthetica. Roč. 51, č. 4 (2013), s. 517-530 ISSN 0300-3604
    Grant CEP: GA ČR GA206/08/1683; GA MŠk(CZ) ED2.1.00/03.0110
    Institucionální podpora: RVO:61388971
    Klíčová slova: antibodies * fluorescence spectra * IMAC chromatography
    Kód oboru RIV: EE - Mikrobiologie, virologie
    Impakt faktor: 1.007, rok: 2013

    Photosystem (PS) II particles retaining a high rate of O-2 evolution were isolated from the mesophilic filamentous cyanobacterium, Spirulina platensis. To achieve high production of PSII complexes in the cells, irradiance from halogen incandescent lamps was used. Disruption of cells by vibration of glass beads proved to be the most suitable procedure for isolation of thylakoid membranes. The selectivity of detergents for PSII particle preparation rose in the order of Triton X-100 < decyl-beta-D-glucopyranoside < dodecyldimethyl-aminooxide < n-heptyl-beta-D-thioglucoside < N-dodecyl-N,N-dimethylammonio-3-propane sulphonate < n-octyl-beta-thioglycoside < octylglucoside < n-dodecyl-beta-D-maltoside. The last four detergents yielded extracts, from which pure PSII particles not contaminated by PSI complexes could be obtained by sucrose-gradient centrifugation (20-45%) at the 43% sucrose level. We assumed both the acceptor and donor sides of the isolated n-dodecyl-beta-D-maltoside (DM) particles to be intact due to high oxygen production by DM particles [1,500 meq(e(-)) mol(-1) (Chl) s(-1)] achieved in the presence of all artificial acceptors tested. The PSII particle fraction from the sucrose gradient was used with immobilized metal (Cu2+) affinity chromatography (IMAC) for the preparation of the PSII core complex. By washing the column with a MES buffer containing MgCl2 and CaCl2, the phycobiliproteins were stripped off. The PSII core complex was eluted in a buffer containing 1% DM, mannitol, MgCl2, NaCl, CaCl2, and E >-aminocaproic acid. SDS-PAGE of the core complex provided pure bands of D1 and D2 proteins and PsbO protein from thylakoid membrane, which were used to raise polyclonal antibodies in rabbits
    Trvalý link: http://hdl.handle.net/11104/0228579