Binding dynamics at the quinone reduction (Qi) site influence the equilibrium interactions of the iron sulfur protein and hydroquinone oxidation (Qo) site of the cytochrome bc1 complex.
Multiple instances of low-potential electron-transport pathway inhibitors that affect the structure of the cytochrome (cyt) bc(1) complex to varying degrees, ranging from changes in hydroquinone (QH(2)) oxidation and cyt c(1) reduction kinetics to proteolytic accessibility of the hinge region of the iron-sulfur-containing subunit (Fe/S protein), have been reported. However, no instance has been documented of any ensuing change on the environment(s) of the [2Fe-2S] cluster. In this work, this issue was addressed in detail by taking advantage of the increased spectral and spatial resolution obtainable with orientation-dependent electron paramagnetic resonance (EPR) spectroscopic analysis of ordered membrane preparations. For the first time, perturbation of the low-potential electron-transport pathway by Q(i)-site inhibitors or various mutations was shown to change the EPR spectra of both the cyt b hemes and the [2Fe-2S] cluster of the Fe/S protein. In particular, two interlinked effects of Q(i)-site modifications on the Fe/S subunit, one changing the local environment of its [2Fe-2S] cluster and a second affecting the mobility of this subunit, are revealed. Remarkably, different inhibitors and mutations at or near the Q(i) site induce these two effects differently, indicating that the events occurring at the Q(i) site affect the global structure of the cyt bc(1). Furthermore, occupancy of discrete Q(i)-site subdomains differently impede the location of the Fe/S protein at the Q(o) site. These findings led us to propose that antimycin A and HQNO mimic the presence of QH(2) and Q at the Q(i) site, respectively. Implications of these findings in respect to the Q(o)-Q(i) sites communications and to multiple turnovers of the cyt bc(1) are discussed.
Cooley
Jason W
JW
Department of Biology, Plant Science Institute, University of Pennsylvania, Philadelphia, Pennsylvania 19104, USA.
Ohnishi
Tomoko
T
Daldal
Fevzi
F
D000968:Antimycin A / Q000378:metabolism; D016227:Benzoquinones / Q000378:metabolism; D001665:Binding Sites / Q000235:genetics; D001667:Binding, Competitive; D002462:Cell Membrane / Q000737:chemistry / Q000378:metabolism; D004578:Electron Spin Resonance Spectroscopy; D004579:Electron Transport; D014450:Electron Transport Complex III / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000737:chemistry / Q000378:metabolism; D004795:Enzyme Stability; D006418:Heme / Q000378:metabolism; D006873:Hydroquinones / Q000378:metabolism*; D006912:Hydroxyquinolines / Q000378:metabolism; D007506:Iron-Sulfur Proteins / Q000737:chemistry* / Q000378:metabolism*; D010084:Oxidation-Reduction; D011090:Polyenes / Q000737:chemistry / Q000378:metabolism; D011809:Quinones / Q000378:metabolism*; D016354:Rhodobacter capsulatus / Q000201:enzymology / Q000235:genetics
D016227:Benzoquinones; D004791:Enzyme Inhibitors; D006873:Hydroquinones; D006912:Hydroxyquinolines; D007506:Iron-Sulfur Proteins; D011090:Polyenes; D011809:Quinones; C001333:2-(n-heptyl)-4-hydroxyquinoline N-oxide; C025232:semiquinone radicals; D006418:Heme; D000968:Antimycin A; C041573:stigmatellin; D014450:Electron Transport Complex III; C031927:hydroquinone