Characterization of a D1-D2-cyt b-559 complex containing 4 chlorophyll a/2 pheophytin a isolated with the use of MgSO4.
A D1-D2-cyt b-559 complex containing 4 chlorophyll alpha, 1 beta-carotene and 1 cytochrome b-559 per 2 pheophytin a has been isolated from spinach with 30% yield using a Q-Sepharose Fast-Flow anion-exchange column equilibrated with 0.1% Triton X-100, 10 mM MgSO4 and 50 mM Tris-HCl (pH 7.2). The preparation was then stabilized with 0.1% dodecyl-beta-D-maltoside. This method gave a yield 10 times higher than that using a Fractogel TSK-DEAE 650(S) column equilibrated with 0.1% Triton X-100, 30 mM NaCl and 50 mM Tris-HCl (pH 7.2). The PS II RC complex was characterized using absorption and fluorescence spectroscopy at 277 and 77 K. A selective reversible bleaching under reducing conditions with maximum at 682 nm, associated with pheophytin a reduction, and light-induced absorption differences with a lifetime of 1.0 ms, ascribed to the triplet state of P680 were measured and indicated that the isolated D1-D2-cyt b-559 complex is active in charge separation. The results are compared with the data obtained for a PS II RC preparation containing 6 chlorophyll alpha, 2 beta-carotene and 1 cyt b-559 per 2 pheophytin a.
Yruela
I
I
Max-Planck Institut für Strahlenchemie, Mülheim an der Ruhr, Germany.
van Kan
P J
PJ
Müller
M G
MG
Holzwarth
A R
AR
D002734:Chlorophyll / Q000032:analysis* / Q000737:chemistry; D000077194:Chlorophyll A; D003573:Cytochrome b Group / Q000737:chemistry / Q000302:isolation & purification*; D005960:Glucosides / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration; D045342:Light-Harvesting Protein Complexes; D046911:Macromolecular Substances; D008278:Magnesium Sulfate; D008970:Molecular Weight; D017830:Octoxynol / Q000494:pharmacology; D010674:Pheophytins / Q000032:analysis* / Q000737:chemistry; D010777:Photochemistry; D045322:Photosynthetic Reaction Center Complex Proteins / Q000737:chemistry*; D045332:Photosystem II Protein Complex; D010944:Plants / Q000737:chemistry*; D013050:Spectrometry, Fluorescence; D013053:Spectrophotometry
D003573:Cytochrome b Group; D005960:Glucosides; D045342:Light-Harvesting Protein Complexes; D046911:Macromolecular Substances; D010674:Pheophytins; D045322:Photosynthetic Reaction Center Complex Proteins; D045332:Photosystem II Protein Complex; D002734:Chlorophyll; C061694:pheophytin a; C040358:dodecyl maltoside; D008278:Magnesium Sulfate; D017830:Octoxynol; C019313:cytochrome b559; D000077194:Chlorophyll A