Purification and properties of the photosystem I reaction center from chloroplasts

1. A reaction center from chloroplasts was purified by means of detergent treatment, differential centrifugation, column chromatography, and sucrose gradient. 2. The reaction center is active in NADP photoreduction by ascorbate. Ferredoxin, ferredoxin-NADP-reductase, and plastocyanin were required for the reaction. 3. The preparation contains five classes of polypeptide chains with apparent molecular weights of 70,000, 25,000, 20,000, 18,000 and 16,000 as determined by gel electrophoresis in sodium dodecyl sulfate. 4. Treatment with 0.5% sodium dodecyl sulfate abolished the NADP photoreduction activity and released the low molecular weight subunits, which were removed by sucrose gradient centrifugation from the high molecular weight ones. The P700 signal is associated with the 70,000 molecular weight polypeptide. 5. Antibody, prepared against the active reaction center, inhibited NADP photoreduction catalyzed by the purified reaction center as well as by isolated chloroplasts. The antibody interacted on immunodiffusion plates with any subchloroplast preparation capable of NADP photoreduction. It also interacted with the purified 70,000 molecular weight polypeptide. 6. It is concluded that both the primary oxidation and the primary reduction in Photosystem I are associated with the 70,000 molecular weight polypeptide.