Iron–sulfur clusters/semiquinones in Complex I

Abstract

NADH-quinone 1 oxidoreductase (Complex I) isolated from bovine heart mitochondria was, until recently, the major source for the study of this most complicated energy transducing device in the mitochondrial respiratory chain. Complex I has been shown to contain 43 subunits and possesses a molecular mass of about 1 million. Recently, Complex I genes have been cloned and sequenced from several bacterial sources including Escherichia coli, Paracoccus denitrificans, Rhodobacter capsulatus and Thermus thermophilus HB-8. These enzymes are less complicated than the bovine enzyme, containing a core of 13 or 14 subunits homologous to the bovine heart Complex I. From this data, important clues concerning the subunit location of both the substrate binding site and intrinsic redox centers have been gleaned. Powerful molecular genetic approaches used in these bacterial systems can identify structure/function relationships concerning the redox components of Complex I. Site-directed mutants at the level of bacterial chromosomes and over-expression and purification of single subunits have allowed detailed analysis of the amino acid residues involved in ligand binding to several iron-sulfur clusters. Therefore, it has become possible to examine which subunits contain individual iron-sulfur clusters, their location within the enzyme and what their ligand residues are. The discovery of g=2.00 EPR signals arising from two distinct species of semiquinone (SQ) in the activated bovine heart submitochondrial particles (SMP) is another line of recent progress. The intensity of semiquinone signals is sensitive to DeltamicroH+ and is diminished by specific inhibitors of Complex I. To date, semiquinones similar to those reported for the bovine heart mitochondrial Complex I have not yet been discovered in the bacterial systems. This mini-review describes three aspects of the recent progress in the study of the redox components of Complex I: (A) the location of the substrate (NADH) binding site, flavin, and most of the iron-sulfur clusters, which have been identified in the hydrophilic electron entry domain of Complex I; (B) experimental evidence indicating that the cluster N2 is located in the amphipathic domain of Complex I, connecting the promontory and membrane parts. Very recent data is also presented suggesting that the cluster N2 may have a unique ligand structure with an atypical cluster-ligation sequence motif located in the NuoB (NQO6/PSST) subunit rather than in the long advocated NuoI (NQO9/TYKY) subunit. The latter subunit contains the most primordial sequence motif for two tetranuclear clusters; (C) the discovery of spin-spin interactions between cluster N2 and two distinct Complex I-associated species of semiquinone. Based on the splitting of the g1 signal of the cluster N2 and concomitant strong enhancement of the semiquinone spin relaxation, one semiquinone species was localized 8-11 A from the cluster N2 within the inner membrane on the matrix side (N-side). Spin relaxation of the other semiquinone species is much less enhanc...

The Mitochondrial Production of Reactive Oxygen Species: Mechanisms and Implications in Human Pathology

“…It is also known that the SQ EPR signal in Complex I is rotenone-sensitive (24); the electron donor to the rst molecule of bound ubiquinone in the Complex is most probably the FeS cluster N2 (24). It is likely that this center is also the electron donor to oxygen either directly or via one-electron reduction of exogenous quinones (Fig.…”

Functional role of Coenzyme Q in the energy coupling of NADH‐CoQ oxidoreductase (Complex I): Stabilization of the semiquinone state with the application of inside‐positive membrane potential to proteoliposomes

“…These authors also proposed a possible role of cluster N1a as an anti-oxidant. N1a has, in general, a very low redox midpoint potential (E m7.0 = −370 mV) [19], and its edge-to-edge distance to FMN is 12.3Å and that to the closest iron-sulfur cluster N3 is 19.4Å. Therefore, N1a seems to play a role as a temporary electron reservoir.…”

“…In this exergonic redox reaction, the transfer of 2 electrons from one molecule of NADH to Q is coupled to the translocation of 4 protons from the negative side (abbreviated as N-side) to the positive side (P-side) of the mitochondrial inner membrane [6,32]. Complex I is the largest energy transducing complex (Molecular weight is ~1 megadalton [7], composed of 45 subunits [3] and contains 1 non-covalently bound FMN, and 8 or 9 iron-sulfur (Fe/S) clusters [19]. Recently the first atomic structure at 3.3Å resolution of the peripheral arm (hydrophilic domain) from Thermus thermophilus HB-8 complex I was determined by Sazanov and Hinchliffe [27] (See Fig.…”

The reaction of NADPH with bovine mitochondrial NADH:ubiquinone oxidoreductase revisited

“…A flavin molecule (FMN) accepts the two reducing equivalents from the hydride formed after the heterolytic cleavage of NADH. Subsequently electrons are transferred, one at the time, via a number of EPR-detectable iron-sulphur (Fe-S) clusters (Beinert and Albracht 1982;Ohnishi 1998) to ubiquinone (Q 10 ). This process is coupled to the extrusion from the mitochondrial matrix of ca.…”

“…Other research groups, most of them studying nonbovine systems, concluded that electron transfer from NADH to Q proceeds via FMN in the 51-kDa subunit all the way to the Fe-S cluster in the PSST subunit and from there to ubiquinone (Ohnishi 1998;Walker 1992;Friedrich 2001;Yagi and Matsuno-Yagi 2003;Hinchliffe and Sazanov 2005;Sazanov and Hinchliffe 2006;Zickermann et al 2009). Possible reasons for the discrepancy between that view and the ones presented here are reviewed in the accompanying paper.…”