Ferredoxin : NADP(+) oxidoreductase is a subunit of the chloroplast cytochrome b(6)f complex

Purified detergent-soluble cytochrome b(6)f complex from chloroplast thylak oid membranes (spinach) and cyanobacteria (Mastigocladus laminosus) was hig hly active, transferring 300-350 electrons per cyt f/s. Visible absorbance spectra showed a red shift of the cytochrome f alpha -band and the Qy chlor ophyll a band in the cyanobacterial complex and an absorbance band in the f lavin 450-480-nm region of the chloroplast complex. An additional high mole cular weight (M-r similar to 35,000) polypeptide in the chloroplast complex was seen in SDS-polyacrylamide gel electrophoresis at a stoichiometry of s imilar to0.9 (cytochrome f)(-1). The extra polypeptide did not stain for he me and was much more accessible to protease than cytochrome f. Electrospray ionization mass spectrometry of CNBr fragments of the 35-kDa polypeptide w as diagnostic for ferredoxin:NADP(+) oxidoreductase (FNR), as were antibody reactivity to FNR and diaphorase activity. The absence of FNR in the cyano bacterial complex did not impair decyl-plastoquinolferricyanide activity. T he activity of the FNR in the chloroplast b(6)f complex was also shown by N ADPH reduction, in the presence of added ferredoxin, of 0.8 heme equivalent s of the cytochrome b(6) subunit. It was inferred that the b(6)f complex wi th bound FNR, one equivalent per monomer, provides the membrane protein con nection to the main electron transfer chain for ferredoxin-dependent cyclic electron transport.