Effects of inhibitors on the activity of the cytochrome b(6)f complex: evidence for the existence of two binding pockets in the lumenal site.
The effects of two inhibitors of electron transfer in the cytochrome b(6)f complex have been studied in whole cells of Chlamydomonas reinhardtii. DNP-INT affected equally the two steps of the concerted oxidation of plastoquinol at the Q(o) site; it decreased the rates of both cytochrome f reduction and cytochrome b(6) turnover, without affecting the amplitude of their redox signals. On the contrary, DBMIB inhibited only the rate of cytochrome f reduction while reducing, at the same time, the amplitude of cytochrome b(6) signals. The accessibility of DNP-INT to the Q(o) site was unaffected by preillumination, while that of DBMIB was greatly enhanced, even after a single turnover of the cytochrome b(6)f complex. Similar results were obtained with a mutant strain (FUD2) where the Q(o) site has an affinity for plastoquinol that is diminished by a factor of approximately 50 [Finazzi, G., et al. (1997) Biochemistry 36, 2867-2874]. However, the binding of the two inhibitors was differentially influenced by the mutation: a factor of approximately 250 was calculated for DNP-INT and a factor of only approximately 5 for DBMIB. This suggests that they bind within the Q(o) site in two distinct pockets, which are differentially involved in the process of quinol oxidation, in agreement with a recent model where two distinct positions for the reduced and semireduced quinones are considered [Crofts, A. R., and Berry, E. A. (1998) Curr. Opin. Struct. Biol. 8, 501-509].
Barbagallo
R P
RP
Centro di Studio del CNR sulla Biologia Cellulare e Molecolare delle Piante, via Celoria 26, 20133 Milano, Italy.
Finazzi
G
G
Forti
G
G
D000818:Animals; D001665:Binding Sites; D016825:Chlamydomonas reinhardtii / Q000201:enzymology / Q000502:physiology; D003573:Cytochrome b Group / Q000037:antagonists & inhibitors* / Q000378:metabolism; D045346:Cytochrome b6f Complex; D004579:Electron Transport; D004791:Enzyme Inhibitors / Q000378:metabolism / Q000494:pharmacology*; D008564:Membrane Potentials
D003573:Cytochrome b Group; D004791:Enzyme Inhibitors; D045346:Cytochrome b6f Complex