The Experts below are selected from a list of 105 Experts worldwide ranked by ideXlab platform
Brian H. Robinson - One of the best experts on this subject based on the ideXlab platform.
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Diagnosis of mitochondrial energy metabolism defects in tissue culture. Induction of MnSOD and bcl‐2 in mitochondria from patients with complex I (NADH‐CoQ Reductase) deficiency
BioFactors (Oxford England), 1998Co-Authors: Brian H. Robinson, Sari Pitkänen, Xiaoping Luo, S. Bratinova, Jacqueline M. Bourgeois, D. C. Lehotay, Sandeep RahaAbstract:Complex I deficiency is perhaps the most common form of mitochondrial respiratory chain disease. We see more patient samples with this defect than any other. The variety of clinical phenotypes seen with complex I deficiency is continually expanding and encompasses nuclear-encoded defects which are in the majority and mtDNA-encoded defects which are in the minority. Complex I is made up of 41 different subunits, 34 of which are encoded in the nucleus and 7 are encoded in mtDNA. A number of defects resulting from point mutations in mtDNA encoded genes of complex I produce the prototype of maternally inherited diseases, Leber’s hereditary optic neuropathy (LHON). More severe symptoms of LHON plus dystonia with degeneration of the basal ganglia can be produced by the 14,459 mtDNA mutation in the ND6 gene of complex I. While induction of manganese superoxide dismutase in mitochondria appears to be an attempt by the cell to reduce free radical damage, we have shown that in the case of respiratory chain defects, this very induction is deleterious to the cell and results in excessive production of the damaging hydroxyl radical [1]. It has been shown in cultured mammalian cell systems that overexpression of the bcl-2 gene can protect against hydroxyl radical production and against hypoxia-induced cell death [4,6,7]. In general, the bcl-2 gene product is thought to be a protective gene against apoptosis [4,6,7]. Some of the bcl-2 protein is anchored in the outer mitochondrial membrane where it seems to exert an effect that interrupts a part of the apoptotic process which may involve induction of free radical production by mitochondria [4,6,7]. bcl-2 protein titre is increased in cultured skin fibroblasts treated with respiratory chain inhibitors [8] and we have found also that it is increased in response to genetically determined respiratory chain defects, especially those in complex I. Seemingly tied in with these protective effects may be an event in hypoxic cell death known as the opening of the mitochondrial permeability transition pore (MPTP) [9]. Evidence is growing that one of the early events in apoptotic cell death involves the opening of MPTP perhaps triggered by a phosphorylation event or by free radical action [9]. The functioning of the transition pore complex
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Familial Cardiomyopathy with Cataracts and Lactic Acidosis: A Defect in Complex I (NADH-Dehydrogenase) of the Mitochondria Respiratory Chain
Pediatric Research, 1996Co-Authors: Sari Pitkänen, Frank Merante, D Ross Mcleod, Derek Applegarth, Tony Tong, Brian H. RobinsonAbstract:Four patients in one generation of a multiply consanguineous pedigree died with cardiomyopathy, cataracts, and lactic acidemia. Postmortem heart and skeletal muscle tissues from one patient were analyzed. A low (12% control) activity of NADH-CoQ Reductase (complex I) in heart and normal activity in skeletal muscle mitochondria was found. Cultured skin fibroblasts obtained from two individuals in the pedigree showed elevated lactate to pyruvate ratios in the range of 2 to 3.5 times normal and decreased complex I + III activity (42 and 54% of control activities) in isolated mitochondria. Western blot analysis and enzymatic assay showed normal levels of CuZn-superoxide dismutase, but grossly elevated levels of the mitochondrial Mnsuperoxide dismutase. Southern blot analysis in heart muscle cells from the patient tested revealed multiple mitochondrial DNA deletions which indicate free oxygen radical damage. We hypothesize that a nuclear-encoded defect in the respiratory chain is responsible for excessive free oxygen radical production in these infants which contributes to the prenatal onset of cardiomyopathy and cataracts.
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Determination of the cDNA sequence for the human mitochondrial 75-kDa Fe-S protein of NADH-coenzyme Q Reductase.
European journal of biochemistry, 1991Co-Authors: Wendy Chow, Ian C Ragan, Brian H. RobinsonAbstract:A human-hepatoma cDNA lambda gt11 expression library was probed with an antibody to holoenzyme complex I (NADH-CoQ Reductase) of the respiratory chain. One of the 30 antibody positive clones was purified to homogeneity, amplified by the polymerase chain reaction (PCR), subcloned and sequenced. It proved to be highly similar to the cDNA sequence for the bovine 75-kDa Fe--S protein. Using the sequence obtained from this library, both sense and antisense oligonucleotides were constructed and used to probe a human kidney cDNA library using PCR amplification with oligonucleotides that flank the polylinker region of the lambda phage. Two further cDNA clones were obtained which overlapped and covered the entire cDNA sequence of 2526 bp. The encoded protein of 727 amino acids has 21 amino acids that differ from the bovine-protein sequence. Northern blot analysis of mRNA from fibroblasts of complex-I deficient patients revealed no abnormalities. We show that this Fe--S protein has significant similarity with (a) the gamma chain of the hydrogen hydrogenase of Alcaligenes eutrophus and (b) the A chain of the formate dehydrogenase of Methanobacterium formicum.
Giorgio Lenaz - One of the best experts on this subject based on the ideXlab platform.
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Mitochondrial quinone Reductases: complex I.
Methods in enzymology, 2004Co-Authors: Giorgio Lenaz, Romana Fato, Alessandra Baracca, Maria Luisa GenovaAbstract:Publisher Summary This chapter provides an overview of mitochondrial quinone Reductases complex I. The NADH:quinone (Coenzyme Q, CoQ, ubiquinone) oxidoReductase (Complex I) is the most complicated enzyme of the respiratory chain in mitochondria and aerobic bacteria. Mitochondrial complex I is a multisubunit enzyme that uses the energy associated to NADH oxidation by CoQ to pump hydrogen ions across the inner membrane, thus significantly contributing to the formation of an electrochemical proton gradient and consequently to the efficiency of the oxidative phosphorylation process. Isolation of Complex I in an active form is not an easy task, so the best way to assay its activity is to study the enzyme in situ, in mitochondrial membranes or in submitochondrial particles. From the functional point of view, Complex I activity can be isolated from the other respiratory complexes by the action of specific inhibitors such as antimycin A and mucidin for complex III and cyanide for complex IV. The chapter provides the details of assay of redox activities of complex. An overview of NADH-CoQ Reductase is presented. The chapter elaborates the determination of NADH dehydrogenase and explains the metabolic flux control and other activities of complex I.
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Steady-state kinetics of the reduction of coenzyme Q analogs by complex I (NADH:ubiquinone oxidoReductase) in bovine heart mitochondria and submitochondrial particles.
Biochemistry, 1996Co-Authors: Romana Fato, E. Estornell, Salvatore Di Bernardo, Francesco Pallotti, And Giovanna Parenti Castelli, Giorgio LenazAbstract:The reduction kinetics of coenzyme Q (CoQ, ubiquinone) by NADH:ubiquinone oxidoReductase (complex I, EC 1.6.99.3) was investigated in bovine heart mitochondrial membranes using water-soluble homologs and analogs of the endogenous ubiquinone acceptor CoQ10 [the lower homologs from CoQ0 to CoQ3, the 6-pentyl (PB) and 6-decyl (DB) analogs, and duroquinone]. By far the best substrates in bovine heart submitochondrial particles are CoQ1 and PB. The kinetics of NADH−CoQ Reductase was investigated in detail using CoQ1 and PB as acceptors. The kinetic pattern follows a ping-pong mechanism; the Km for CoQ1 is in the range of 20 μM but is reversibly increased to 60 μM by extraction of the endogenous CoQ10. The increased Km in CoQ10-depleted membranes indicates that endogenous ubiquinone not only does not exert significant product inhibition but rather is required for the appropriate structure of the acceptor site. The much lower Vmax with CoQ2 but not with DB as acceptor, associated with an almost identical Km, sug...
Romana Fato - One of the best experts on this subject based on the ideXlab platform.
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Mitochondrial quinone Reductases: complex I.
Methods in enzymology, 2004Co-Authors: Giorgio Lenaz, Romana Fato, Alessandra Baracca, Maria Luisa GenovaAbstract:Publisher Summary This chapter provides an overview of mitochondrial quinone Reductases complex I. The NADH:quinone (Coenzyme Q, CoQ, ubiquinone) oxidoReductase (Complex I) is the most complicated enzyme of the respiratory chain in mitochondria and aerobic bacteria. Mitochondrial complex I is a multisubunit enzyme that uses the energy associated to NADH oxidation by CoQ to pump hydrogen ions across the inner membrane, thus significantly contributing to the formation of an electrochemical proton gradient and consequently to the efficiency of the oxidative phosphorylation process. Isolation of Complex I in an active form is not an easy task, so the best way to assay its activity is to study the enzyme in situ, in mitochondrial membranes or in submitochondrial particles. From the functional point of view, Complex I activity can be isolated from the other respiratory complexes by the action of specific inhibitors such as antimycin A and mucidin for complex III and cyanide for complex IV. The chapter provides the details of assay of redox activities of complex. An overview of NADH-CoQ Reductase is presented. The chapter elaborates the determination of NADH dehydrogenase and explains the metabolic flux control and other activities of complex I.
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Steady-state kinetics of the reduction of coenzyme Q analogs by complex I (NADH:ubiquinone oxidoReductase) in bovine heart mitochondria and submitochondrial particles.
Biochemistry, 1996Co-Authors: Romana Fato, E. Estornell, Salvatore Di Bernardo, Francesco Pallotti, And Giovanna Parenti Castelli, Giorgio LenazAbstract:The reduction kinetics of coenzyme Q (CoQ, ubiquinone) by NADH:ubiquinone oxidoReductase (complex I, EC 1.6.99.3) was investigated in bovine heart mitochondrial membranes using water-soluble homologs and analogs of the endogenous ubiquinone acceptor CoQ10 [the lower homologs from CoQ0 to CoQ3, the 6-pentyl (PB) and 6-decyl (DB) analogs, and duroquinone]. By far the best substrates in bovine heart submitochondrial particles are CoQ1 and PB. The kinetics of NADH−CoQ Reductase was investigated in detail using CoQ1 and PB as acceptors. The kinetic pattern follows a ping-pong mechanism; the Km for CoQ1 is in the range of 20 μM but is reversibly increased to 60 μM by extraction of the endogenous CoQ10. The increased Km in CoQ10-depleted membranes indicates that endogenous ubiquinone not only does not exert significant product inhibition but rather is required for the appropriate structure of the acceptor site. The much lower Vmax with CoQ2 but not with DB as acceptor, associated with an almost identical Km, sug...
Maria Luisa Genova - One of the best experts on this subject based on the ideXlab platform.
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Mitochondrial quinone Reductases: complex I.
Methods in enzymology, 2004Co-Authors: Giorgio Lenaz, Romana Fato, Alessandra Baracca, Maria Luisa GenovaAbstract:Publisher Summary This chapter provides an overview of mitochondrial quinone Reductases complex I. The NADH:quinone (Coenzyme Q, CoQ, ubiquinone) oxidoReductase (Complex I) is the most complicated enzyme of the respiratory chain in mitochondria and aerobic bacteria. Mitochondrial complex I is a multisubunit enzyme that uses the energy associated to NADH oxidation by CoQ to pump hydrogen ions across the inner membrane, thus significantly contributing to the formation of an electrochemical proton gradient and consequently to the efficiency of the oxidative phosphorylation process. Isolation of Complex I in an active form is not an easy task, so the best way to assay its activity is to study the enzyme in situ, in mitochondrial membranes or in submitochondrial particles. From the functional point of view, Complex I activity can be isolated from the other respiratory complexes by the action of specific inhibitors such as antimycin A and mucidin for complex III and cyanide for complex IV. The chapter provides the details of assay of redox activities of complex. An overview of NADH-CoQ Reductase is presented. The chapter elaborates the determination of NADH dehydrogenase and explains the metabolic flux control and other activities of complex I.
Takeshi Kinoshita - One of the best experts on this subject based on the ideXlab platform.
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mode of antibacterial action of retrochalcones from glycyrrhiza inflata
Phytochemistry, 1998Co-Authors: Hiroyuki Haraguchi, Katsuhito Tanimoto, Yukiyoshi Tamura, Kenji Mizutani, Takeshi KinoshitaAbstract:Abstract The antibacterial mechanism of retrochalcones isolated from the roots of Glycyrrhiza inflata was studied. Licochalcone A and C inhibited electron transport in the bacterial respiratory chain. Licochalcone A-D and echinatin, retrochalcones isolated from the roots of Glycyrrhiza inflata, showed antimicrobial activity. Among them, licochalcone A and C had potent activity against some Grampositive bacteria. These retrochalcones inhibited oxygen consumption in susceptible bacterial cells. The oxidation of NADH in bacterial membrane preparations was also inhibited by them. NADH-cytochrome c Reductase was inhibited by licochalcones, while cytochrome c oxidase was not. NADH-CoQ Reductase and NADH-FMN oxidoReductase were not inhibited. The site of respiratory inhibition of licochalcones was thought to be between CoQ and cytochrome c in the bacterial respiratory electron transport chain.
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Mode of antibacterial action of retrochalcones from Glycyrrhiza inflata
Phytochemistry, 1998Co-Authors: Hiroyuki Haraguchi, Katsuhito Tanimoto, Yukiyoshi Tamura, Kenji Mizutani, Takeshi KinoshitaAbstract:Licochalcone A-D and echinatin, retrochalcones isolated from the roots of Glycyrrhiza inflata, showed antimicrobial activity. Among them, licochalcone A and C had potent activity against some Gram-positive bacteria. These retrochalcones inhibited oxygen consumption in susceptible bacterial cells. The oxidation of NADH in bacterial membrane preparations was also inhibited by them. NADH-cytochrome c Reductase was inhibited by licochalcones, while cytochrome c oxidase was not. NADH-CoQ Reductase and NADH-FMN oxidoReductase were not inhibited. The site of respiratory inhibition of licochalcones was thought to be between CoQ and cytochrome c in the bacterial respiratory electron transport chain.
