Inhibitor effects on redox-linked protonations of the b haems of the mitochondrial bc1 complex

Inhibition of complex III promotes loss of Ca2+ dependence for mitochondrial superoxide formation and permeability transition evoked by peroxynitrite

“…1A-D) were indeed sensitive to the complex I inhibitor rotenone (0.5 M) under conditions in which inhibition of oxygen consumption Journal of Cell Science 120 (11) was also observed (not shown). In addition, the effects of rotenone were mimicked by myxothiazol (5 M), an agent binding to a site of complex III different from that of antimycin A or HQNO (Rich et al, 1990) thereby preventing the electron flow from the reduced coenzyme Q to cytochrome c 1 , and the ensuing formation of ubisemiquinone (Brand and Hermfisse, 1997). Thus, as previously shown (Guidarelli et al, 2000;Guidarelli et al, 2006), ubisemiquinone serves as an electron donor promoting the time-dependent formation of superoxide and H 2 O 2 in cells exposed to either high concentrations (e.g.…”

Exploration of Ligands to the Qi Site Semiquinone in the bc 1 Complex Using High-resolution EPR

“…The SQ at the Q i site has been studied most completely in the context of redox titrations of the signal detected by continuous wave-X-band EPR (8,9,14,15). From the pH dependence, it has been concluded that the anion is the stable species, and parameters have been proposed that describe the stability in terms of the disproportionation reaction, midpoint redox potential (E m ) values for the two half-cell reactions, and pK values for the dissociable forms.…”

“…Samples were incubated for 5 min and then stored under liquid N 2 until used for spectroscopy. 2) The bc 1 complex at 50 -200 M was suspended in a medium containing 100 mM KCl, 50 mM CHES, 5 mM MgSO 4 , 0.01% n-dodecyl-␤-D-maltoside, 15 g/ml phosphatidylcholine at pH 9.0, 18 -20°C in a redox titration cell which was kept anaerobic by argon gas. Redox mediators present were 40 M 1,2-naphthoquinone and 40 M 1,4-naphthoquinone.…”

“…3) The bc 1 complex at 50 -200 M was suspended in a medium containing 100 mM KCl, 50 mM CHES, 5 mM MgSO 4 , 0.01% n-dodecyl-␤-D-maltoside, 15 g/ml phosphatidylcholine, and excess myxothiazol at pH 9.0 and reduced by the addition of decylubihydroquinone in 2-fold molar excess over the bc 1 complex monomer concentration. Samples were immediately stored under liquid N 2 until used for spectroscopy.…”

Mitochondrial cytochrome b: evolution and structure of the protein

“…Correlations between an unusual response to one bc 1 inhibitor and the sequence of the cytochrome b protein are most convincing when they combine sequence analysis in related s p e o e s with the information derived from mutants In the case of the natural resistance of fish to fumculosln [90], the results of a selected screening of fumculosin sensitivity m animal m l t o c h o n d n a suggested that the substitution of the conserved alanme 126 with the bulky m e t h l o n m e in the fish protein (Fig 1) is probably responsible for a substantial increase m the tltre of this inhibitor relatwe to normally sensltwe s p e o e s (Table V and R e f 90) Natural resistance in the plant mItochondnal bc t complex [142] could also be correlated with the exchange of A126 with the bulkier V m the cytochrome b sequence (Fig 2 and R e f 90) The buried location of position 126 within the transmembrane sector of hehx C (Fig 1) may account for the 'hybrid' effects of fumculosln, which effects both center i and center o [90,114,[143][144][145][146] Note that the proteins having a bulky amino a o d at poslt~on 126 also show resistance to U H D B T (Table V), which xs a center o inhibitor that shares with funlculosm the property of effectlng both qulnone sites [113,114,143,144] An interesting property of funlculosln is its remarkable species specificity, even among mammals [143,147] The volume pattern and the comparison of the sequences of sensltwe and resistant species suggested previously that position 194 may also be revolved in funlculosln binding [90] By inspecting the aligned sequences, we noticed that the rabbit protein shows the substitution of alanine 194 with a bulkier vahne residue (Fig 1) Hence, rabbit mltochondrla were expected to be quite resistant to funlculosin, which would explain why rabbits are resistant to this drug in vwo [147] This ~s indeed the case, since the inhibitory potency of funlculosln on the ublqumol cytochrome c reductase actwlty is about 60-fold lower for m l t o c h o n d n a ~solated from rabbit than those from sensltwe mammals (Table V) The cytochrome b proteins of zebra and donkey also have vahne at position 194 (Fig 1 and R e f 32) and differ from that of pig, a...…”