Characterization of the Ubiquinone Reduction Site of Mitochondrial Complex I Using Bulky Synthetic Ubiquinones

Abstract

A wide variety of alkyl derivatives of Q2 (6-geranyl-2, 3-dimethoxy-5-methyl-1,4-benzoquinone) and DB (6-n-decyl-2, 3-dimethoxy-5-methyl-1,4-benzoquinone), in which methoxy groups of the 2- and/or 3-positions of the quinone ring were replaced by other bulky alkoxy groups from ethoxy to butoxy, were prepared by novel synthetic procedures. Electron-accepting activities of the bulky quinones were investigated with bovine heart mitochondrial complex I and its counterpart of Paracoccus denitrificans(NDH-1) to elucidate structural and functional features of the quinone reduction site of the enzymes. The bulky quinone analogues served as sufficient electron acceptors from the physiological quinone reduction site of bovine complex I. Considering the very poor activities of even the ethoxy derivatives as substrates for other respiratory enzymes such as mitochondrial complexes II and III [He, D. Y., Gu, L. Q., Yu, L., and Yu, C. A. (1994) Biochemistry 33, 880-884], this result indicated that the quinone reduction site of bovine complex I is spacious enough to accommodate bulky exogenous substrates. In contrast to bovine complex I, bulky quinone analogues served as poor electron acceptors with Paracoccus NDH-1. These observations indicated that bovine complex I recognizes the substrate structure with poor specificity. The substituent effects in the 2- and 3-positions of the quinone ring on the electron-transfer activity with bovine complex I differed significantly between Q2 and DB series despite having the same total number of carbon atoms in the side chain. The inhibitory effect involving Q2 due to its geranyl side chain was markedly diminished by structural modifications of the quinone ring moiety. These findings indicate that the side chain plays a specific role in the redox reaction and that the quinone ring and side-chain moieties contribute interdependently to binding interaction. Moreover, structural dependency of the proton-pumping activity of the quinone analogues was comparable to that of the electron-transfer activity with bovine complex I, indicating that the mechanism of redox-driven proton-pumping does not differ depending upon the substrate structure.

Three Classes of Inhibitors Share a Common Binding Domain in Mitochondrial Complex I (NADH:Ubiquinone Oxidoreductase)

“…The idea of a fairly large ubiquinone binding domain also fits well with recently published data showing that this pocket is sufficiently spacious to accommodate rather bulky exogenous ubiquinones (52).…”

The Ubiquinone-binding Site in NADH:Ubiquinone Oxidoreductase from Escherichia coli

“…This fits well with the proposed proximity of this subunit with NuoB, NuoD, and NuoH (42). Because the binding of the various inhibitors is affected by the presence of other inhibitors, it was concluded that these binding sites partially overlap (43). Here, we have shown that the binding of [ 3 H]azido-Q is not influenced by addition of inhibitors, indicating that the inhibitor binding site(s) may not be identical to the quinone binding site(s).…”

Altered Quinone Biosynthesis in the Long-lived clk-1Mutants of Caenorhabditis elegans

“…Succinate-quinone reductase activities were measured as described previously (21). DMQ 2 and 3-hydroxy-UQ 2 were synthesized as described previously (22). All the assays were performed at 20°C.…”