A Pair of Membrane-Embedded Acidic Residues in the NuoK Subunit ofEscherichia coliNDH-1, a Counterpart of the ND4L Subunit of the Mitochondrial Complex I, Are Required for High Ubiquinone Reductase Activity†

Biochemistry ◽  
2004 ◽  
Vol 43 (3) ◽  
pp. 773-781 ◽  
Author(s):  
Marko Kervinen ◽  
Jukka Pätsi ◽  
Moshe Finel ◽  
Ilmo E. Hassinen
Keyword(s):  
Complex I ◽  
Reductase Activity ◽  
Mitochondrial Complex I ◽  
Mitochondrial Complex ◽  
Acidic Residues

scholarly journals The specificity of mitochondrial complex I for ubiquinones

1996 ◽  
Vol 313 (1) ◽  
pp. 327-334 ◽  
Author(s):  
Mauro ESPOSTI DEGLI ◽  
Anna NGO ◽  
Gabrielle L. McMULLEN ◽  
Anna GHELLI ◽  
Francesca SPARLA ◽  
...  
Keyword(s):  
Membrane Potential ◽  
Electron Transport ◽  
Complex I ◽  
Reductase Activity ◽  
Redox Activity ◽  
Mitochondrial Complex I ◽  
Transport Activity ◽  
Mitochondrial Complex ◽  
Potential Generation ◽  
Electron Transport Activity

We report the first detailed study on the ubiquinone (coenzyme Q; abbreviated to Q) analogue specificity of mitochondrial complex I, NADH:Q reductase, in intact submitochondrial particles. The enzymic function of complex I has been investigated using a series of analogues of Q as electron acceptor substrates for both electron transport activity and the associated generation of membrane potential. Q analogues with a saturated substituent of one to three carbons at position 6 of the 2,3-dimethoxy-5-methyl-1,4-benzoquinone ring have the fastest rates of electron transport activity, and analogues with a substituent of seven to nine carbon atoms have the highest values of association constant derived from NADH:Q reductase activity. The rate of NADH:Q reductase activity is potently but incompletely inhibited by rotenone, and the residual rotenone-insensitive rate is stimulated by Q analogues in different ways depending on the hydrophobicity of their substituent. Membrane potential measurements have been undertaken to evaluate the energetic efficiency of complex I with various Q analogues. Only hydrophobic analogues such as nonyl-Q or undecyl-Q show an efficiency of membrane potential generation equivalent to that of endogenous Q. The less hydrophobic analogues as well as the isoprenoid analogue Q-2 are more efficient as substrates for the redox activity of complex I than for membrane potential generation. Thus the hydrophilic Q analogues act also as electron sinks and interact incompletely with the physiological Q site in complex I that pumps protons and generates membrane potential.

Leber hereditary optic neuropathy mutations in the ND6 subunit of mitochondrial complex I affect ubiquinone reduction kinetics in a bacterial model of the enzyme

2007 ◽  
Vol 409 (1) ◽  
pp. 129-137 ◽  
Author(s):  
Jukka Pätsi ◽  
Marko Kervinen ◽  
Moshe Finel ◽  
Ilmo E. Hassinen
Keyword(s):  
Optic Neuropathy ◽  
Complex I ◽  
Reductase Activity ◽  
Substrate Inhibition ◽  
Reduction Rate ◽  
Mitochondrial Complex I ◽  
Mitochondrial Complex ◽  
Leber Hereditary Optic Neuropathy ◽  
Nadh:Quinone Oxidoreductase ◽  
Hereditary Optic Neuropathy

LHON (Leber hereditary optic neuropathy) is a maternally inherited disease that leads to sudden loss of central vision at a young age. There are three common primary LHON mutations, occurring at positions 3460, 11778 and 14484 in the human mtDNA (mitochondrial DNA), leading to amino acid substitutions in mitochondrial complex I subunits ND1, ND4 and ND6 respectively. We have now examined the effects of ND6 mutations on the function of complex I using the homologous NuoJ subunit of Escherichia coli NDH-1 (NADH:quinone oxidoreductase) as a model system. The assembly level of the NDH-1 mutants was assessed using electron transfer from deamino-NADH to the ‘shortcut’ electron acceptor HAR (hexammine ruthenium), whereas ubiquinone reductase activity was determined using DB (decylubiquinone) as a substrate. Mutant growth in minimal medium with malate as the main carbon source was used for initial screening of the efficiency of energy conservation by NDH-1. The results indicated that NuoJ-M64V, the equivalent of the common LHON mutation in ND6, had a mild effect on E. coli NDH-1 activity, while nearby mutations, particularly NuoJ-Y59F, NuoJ-V65G and NuoJ-M72V, severely impaired the DB reduction rate and cell growth on malate. NuoJ-Met64 and NuoJ-Met72 position mutants lowered the affinity of NDH-1 for DB and explicit C-type inhibitors, whereas NuoJ-Y59C displayed substrate inhibition by oxidized DB. The results are compatible with the notion that the ND6 subunit delineates the binding cavity of ubiquinone substrate, but does not directly take part in the catalytic reaction. How these changes in the enzyme's catalytic properties contribute to LHON pathogenesis is discussed.

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