Charge translocation by mitochondrial NADH:ubiquinone oxidoreductase (complex I) from Yarrowia lipolytica measured on solid-supported membranes

2016 ◽  
Vol 479 (2) ◽  
pp. 277-282 ◽  
Author(s):  
Ilka Siebels ◽  
Stefan Dröse
Keyword(s):  
Yarrowia Lipolytica ◽  
Complex I ◽  
Nadh:Ubiquinone Oxidoreductase ◽  
Supported Membranes ◽  
Charge Translocation ◽  
Solid Supported Membranes

External alternative NADH:ubiquinone oxidoreductase redirected to the internal face of the mitochondrial inner membrane rescues complex I deficiency in Yarrowia lipolytica

2001 ◽  
Vol 114 (21) ◽  
pp. 3915-3921 ◽  
Author(s):  
Stefan J. Kerscher ◽  
Andrea Eschemann ◽  
Pamela M. Okun ◽  
Ulrich Brandt
Keyword(s):  
Yarrowia Lipolytica ◽  
Complex I ◽  
Functional Expression ◽  
Proton Pumping ◽  
Nadh:Ubiquinone Oxidoreductase ◽  
Inner Membrane ◽  
Mitochondrial Inner Membrane ◽  
The Matrix ◽  
Cytosolic Side ◽  
Respiratory Membrane

Alternative NADH:ubiquinone oxidoreductases are single subunit enzymes capable of transferring electrons from NADH to ubiquinone without contributing to the proton gradient across the respiratory membrane. The obligately aerobic yeast Yarrowia lipolytica has only one such enzyme, encoded by the NDH2 gene and located on the external face of the mitochondrial inner membrane. In sharp contrast to ndh2 deletions, deficiencies in nuclear genes for central subunits of proton pumping NADH:ubiquinone oxidoreductases (complex I) are lethal. We have redirected NDH2 to the internal face of the mitochondrial inner membrane by N-terminally attaching the mitochondrial targeting sequence of NUAM, the largest subunit of complex I. Lethality of complex I mutations was rescued by the internal, but not the external version of alternative NADH:ubiquinone oxidoreductase. Internal NDH2 also permitted growth in the presence of complex I inhibitors such as 2-decyl-4-quinazolinyl amine (DQA). Functional expression of NDH2 on both sides of the mitochondrial inner membrane indicates that alternative NADH:ubiquinone oxidoreductase requires no additional components for catalytic activity. Our findings also demonstrate that shuttle mechanisms for the transfer of redox equivalents from the matrix to the cytosolic side of the mitochondrial inner membrane are insufficient in Y. lipolytica.

A single external enzyme confers alternative NADH:ubiquinone oxidoreductase activity in Yarrowia lipolytica

1999 ◽  
Vol 112 (14) ◽  
pp. 2347-2354 ◽  
Author(s):  
S.J. Kerscher ◽  
J.G. Okun ◽  
U. Brandt
Keyword(s):  
Yarrowia Lipolytica ◽  
Complex I ◽  
Nadh:Ubiquinone Oxidoreductase ◽  
Inner Mitochondrial Membrane ◽  
Gene Encoding ◽  
Oxidoreductase Activity ◽  
Mitochondrial Inner Membrane ◽  
Energy Conserving ◽  
And Function ◽  
Complex I Activity

NADH:ubiquinone oxidoreductases catalyse the first step within the diverse pathways of mitochondrial NADH oxidation. In addition to the energy-conserving form commonly called complex I, fungi and plants contain much simpler alternative NADH:ubiquinone oxido-reductases that catalyze the same reaction but do not translocate protons across the inner mitochondrial membrane. Little is known about the distribution and function of these enzymes. We have identified YLNDH2 as the only gene encoding an alternative NADH:ubiquinone oxidoreductase (NDH2) in the obligate aerobic yeast Yarrowia lipolytica. Cells carrying a deletion of YLNDH2 were fully viable; full inhibition by piericidin A indicated that complex I activity was the sole NADH:ubiquinone oxidoreductase activity left in the deletion strains. Studies with intact mitochondria revealed that NDH2 in Y. lipolytica is oriented towards the external face of the mitochondrial inner membrane. This is in contrast to the situation seen in Saccharomyces cerevisiae, Neurospora crassa and in green plants, where internal alternative NADH:ubiquinone oxidoreductases have been reported. Phylogenetic analysis of known NADH:ubiquinone oxidoreductases suggests that during evolution conversion of an ancestral external alternative NADH:ubiquinone oxidoreductase to an internal enzyme may have paved the way for the loss of complex I in fermenting yeasts like S. cerevisiae.

Mutants of Chlamydomonas reinhardtii Deficient in Mitochondrial Complex I: Characterization of Two Mutations Affecting the nd1 Coding Sequence

Genetics ◽  
2001 ◽  
Vol 158 (3) ◽  
pp. 1051-1060
Author(s):  
Claire Remacle ◽  
Denis Baurain ◽  
Pierre Cardol ◽  
René F Matagne
Keyword(s):  
Mitochondrial Genome ◽  
Chlamydomonas Reinhardtii ◽  
Complex I ◽  
Slow Growth ◽  
Nadh:Ubiquinone Oxidoreductase ◽  
Sequencing Analysis ◽  
Mitochondrial Complex ◽  
Genetical Analysis ◽  
Complex I Activity

Abstract The mitochondrial rotenone-sensitive NADH:ubiquinone oxidoreductase (complex I) comprises more than 30 subunits, the majority of which are encoded by the nucleus. In Chlamydomonas reinhardtii, only five components of complex I are coded for by mitochondrial genes. Three mutants deprived of complex I activity and displaying slow growth in the dark were isolated after mutagenic treatment with acriflavine. A genetical analysis demonstrated that two mutations (dum20 and dum25) affect the mitochondrial genome whereas the third mutation (dn26) is of nuclear origin. Recombinational analyses showed that dum20 and dum25 are closely linked on the genetic map of the mitochondrial genome and could affect the nd1 gene. A sequencing analysis confirmed this conclusion: dum20 is a deletion of one T at codon 243 of nd1; dum25 corresponds to a 6-bp deletion that eliminates two amino acids located in a very conserved hydrophilic segment of the protein.

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