Ferredoxin:NADP+ oxidoreductase bound to cytochrome b₆f complex is active in plastoquinone reduction: implications for cyclic electron transport.
In this study, we have compared three isolation methods of cytochrome b₆f complex, obtained from spinach (Spinacia oleracea), differing in the preservation of the cytochrome b₆f-associated ferredoxin:NADP+ oxidoreductase (FNR). Although the complexes isolated by all the methods showed the presence of the FNR peptide(s), when incorporated into liposome membranes, the NADPH-PQ (plastoquinone) oxidoreductase activity was not detected for the cytochrome b₆f complex isolated with the original method including a NaBr wash. Some activity was found for the complex isolated with the omission of the wash, but the highest activity was detected for the complex isolated with the use of digitonin. The reaction rate of PQ reduction of the investigated complexes in liposomes was not significantly influenced by the addition of free FNR or ferredoxin. The reaction was inhibited by about 60% in the presence of 2 µM 2-n-nonyl-4-hydroxyquinoline N-oxide, an inhibitor of the cytochrome b₆ f complex at the Q(i) site, while it was not affected by triphenyltin or isobutyl cyanide that interacts with the recently identified heme c(i) . The obtained data indicate that FNR associated with the cytochrome b₆ f complex can participate in the cyclic electron transport as PSI-PQ or NADPH-PQ oxidoreductase. Moreover, we have shown that PQ can be non-enzymatically reduced by ascorbate in liposomes and this reaction might be involved in non-photochemical reduction pathways of the PQ-pool in chloroplasts.
Department of Plant Physiology and Biochemistry, Faculty of Biochemistry, Biophysics and Biotechnology, Jagiellonian University, Kraków, Poland.
Renata
R
Szymańska
Jolanta
J
Dłużewska
Ireneusz
I
Slesak
Jerzy
J
Kruk
D045346:Cytochrome b6f Complex / Q000302:isolation & purification / Q000378:metabolism*; D004579:Electron Transport; D005287:Ferredoxin-NADP Reductase / Q000032:analysis / Q000378:metabolism*; D005182:Flavin-Adenine Dinucleotide / Q000032:analysis; D008081:Liposomes / Q000737:chemistry; D009247:NADH, NADPH Oxidoreductases / Q000378:metabolism; D010084:Oxidation-Reduction; D010940:Plant Proteins / Q000378:metabolism; D010971:Plastoquinone / Q000737:chemistry*; D018724:Spinacia oleracea / Q000201:enzymology*
D008081:Liposomes; D010940:Plant Proteins; D005182:Flavin-Adenine Dinucleotide; D045346:Cytochrome b6f Complex; D005287:Ferredoxin-NADP Reductase; D009247:NADH, NADPH Oxidoreductases; C072396:NAD(P)H plastoquinone oxidoreductase; D010971:Plastoquinone