Isolation of Cytochrome b 6 f Complex from Grana and Stroma Membranes from Spinach Chloroplasts

Abstract

The cytochrome b6f complex is located in the appressed granal membranes and nonappressed stroma thylakoids. The procedure presents isolation of the complex from both types of thylakoids by washing with NaBr, detergent treatment, ammonium sulfate fractionation, and sucrose gradient centrifugation. Optimal concentration of the detergent is lower for grana than for stroma vesicles. The cytochrome b6f complex from stroma lamellae locates at a higher density in the sucrose gradient than the granal complex. Electrophoretic analyses indicate that both complexes are dimeric and contain four large subunits and at least three small subunits with masses below 4 kDa. Plastocyanin and 15 kDa protein are also identified in the complexes but in variable amounts.