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Purification and crystallization of Photosystem I complex from a phycobilisome-less mutant of the cyanobacterium Synechococcus PCC 7002

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Abstract

An active photosystem (PSI) complex was isolated from a phycobilisome-less mutant of the mesophilic cyanobacterium Synechococcus PCC 7002 by a mild procedure. Purification of PS I was achieved using a sucrose density gradient and an isoelectric focussing subsequent to the extraction of PSI from thylakoids with dodecyl-β-maltoside. Electron microscopy and gel filtration HPLC suggested that the isolated complex represents a trimeric form of PSI. The trimeric form was resistant to pH or detergent exchange. A ‘molecular weight’ of 690 kDa to 760 kDa has been determined for the complex by gel filtration HPLC in several detergents or mixtures of detergents.

The PSI complex contains the polypeptides of the psaA, psaB, psaC, psaD, psaE, psaL gene products and two small polypeptides as determined by SDS-PAGE and N-terminal sequencing; its antenna size is 77±2 Chl a/P700. The full set of Fe-S clusters (FA, FB and FX) was observed by EPR-spectroscopy. A preliminary characterization of crystals obtained from this preparation was carried out using SDS-PAGE, optical and EPR spectroscopy.

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Abbreviations

BA:

benzamidine

CAS:

6-amino-n-caproic acid

C8-G:

octyl-β-D-glucopyranoside

C12-M:

lauryl-β-D-maltoside

C10-M:

decyl-β-D-maltoside

C8-TG:

octyl-β-D-thioglucoside

Chl a :

chlorophyll a

EPR:

electron paramagnetic resonance

FA, FB, FX :

iron-sulfur centers

HPLC:

high perfomance liquid chromatography

kDa:

kilodalton(s)

LDAO:

lauryldimethylamine oxide

MES:

2-(N-morpholino)ethanesulfonic acid

PSI:

Photosystem I

PS II:

Photosystem II

P700 :

primary electron donor

SB12:

sulfobetain 12

SDS-PAGE:

sodium dodecyl sulfate polyacrylamide gel electrophoresis

Tris:

tris(hydroxymethyl)-aminomethane

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Authors and Affiliations

  1. Abteilung Molekulare Membranbiologie, Max-Planck-Institut für Biophysik, Heinrich Hoffmann Straße 7, D-6000, Frankfurt am Main 71, Germany

    G. Tsiotis & H. Michel

  2. Institut de Biologie Physico-Chimique, 13, Rue Pierre et Marie Curie, F-75005, Paris, France

    W. Nitschke

  3. Max-Planck-Institut für Biophysik, Abteilung Physiologie, Kennedy-Allee 70, D-6000, Frankfurt am Main 71, Germany

    W. Haase

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  1. G. Tsiotis
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  2. W. Nitschke
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  3. W. Haase
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  4. H. Michel
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Tsiotis, G., Nitschke, W., Haase, W. et al. Purification and crystallization of Photosystem I complex from a phycobilisome-less mutant of the cyanobacterium Synechococcus PCC 7002. Photosynth Res 35, 285–297 (1993). https://doi.org/10.1007/BF00016559

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  • DOI: https://doi.org/10.1007/BF00016559

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