Isolation and Partial Characterization of the NADH Dehydrogenase Complex from Barley Chloroplast Thylakoids

Quiles, Marı́a José; Albacete, María Eugenia; Sabater, Bartolomé; Cuello, Juan

doi:10.1093/oxfordjournals.pcp.a029064

Cited by 23 publications

(19 citation statements)

References 29 publications

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“…With regard to the fraction resuspended from thylakoid with Triton X-100, immunoaffinity achieved 100-fold purification of the activity with a recovery of 22%. The activity of purified fractions on NADPH was only 3.6% of their activity on NADH, which is in accordance with previous results for the activities investigated in barley (14) and pea (15) and indicates that the NADH dehydrogenase activity of pure thylakoid Ndh complex was essentially NADH-specific, although crude thylakoid preparations also contain NADPH dehydrogenase activities (14). NADH dehydrogenase activity of purified Ndh complex was 100% inhibited after 30 min incubation with NDH-A antibody, confirming that the activity of purified fraction was due to the Ndh complex that includes the NDH-A polypeptide.…”

Section: Purification By Immunoaffinity Of the Ndh Complex Of Barleysupporting

confidence: 93%

Chlororespiration and Poising of Cyclic Electron Transport

Casano

Zapata

Martı́n

et al. 2000

Journal of Biological Chemistry

143

111

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Section: Purification By Immunoaffinity Of the Ndh Complex Of Barleysupporting

confidence: 93%

Chlororespiration and Poising of Cyclic Electron Transport

Casano

Zapata

Martı́n

et al. 2000

Journal of Biological Chemistry

143

111

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“…Total NADH:ferricyanide oxidoreductase activity reflects not only the activity of complex I but also that of other cellular NADH dehydrogenases (Quiles et al 1996). However, because most of the NADH:ferricyanide oxidoreductase activity in Chlamydomonas is due to the NADH dehydrogenase activity of the peripheral arm of complex I, the level of this activity is reduced in complex I mutants.…”

Section: Resultsmentioning

confidence: 99%

A Forward Genetic Screen Identifies Mutants Deficient for Mitochondrial Complex I Assembly in Chlamydomonas reinhardtii

et al. 2011

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Mitochondrial complex I is the largest multimeric enzyme of the respiratory chain. The lack of a model system with facile genetics has limited the molecular dissection of complex I assembly. Using Chlamydomonas reinhardtii as an experimental system to screen for complex I defects, we isolated, via forward genetics, amc1-7 nuclear mutants (for assembly of mitochondrial complex I) displaying reduced or no complex I activity. Blue native (BN)-PAGE and immunoblot analyses revealed that amc3 and amc4 accumulate reduced levels of the complex I holoenzyme (950 kDa) while all other amc mutants fail to accumulate a mature complex. In amc1, -2, -5-7, the detection of a 700 kDa subcomplex retaining NADH dehydrogenase activity indicates an arrest in the assembly process. Genetic analyses established that amc5 and amc7 are alleles of the same locus while amc1-4 and amc6 define distinct complementation groups. The locus defined by the amc5 and amc7 alleles corresponds to the NUOB10 gene, encoding PDSW, a subunit of the membrane arm of complex I. This is the first report of a forward genetic screen yielding the isolation of complex I mutants. This work illustrates the potential of using Chlamydomonas as a genetically tractable organism to decipher complex I manufacture.

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“…1). NADH dehydrogenase activity has also been detected in barley thylakoids (27), but as yet there is no information as to whether this activity is catalyzed by the Ndh complex (28).…”

Section: Discussionmentioning

confidence: 99%

The plastid ndh genes code for an NADH-specific dehydrogenase: Isolation of a complex I analogue from pea thylakoid membranes

Sazanov

Burrows

Nixon

1998

Proc. Natl. Acad. Sci. U.S.A.

191

125

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The plastid genomes of several plants contain ndh genes-homologues of genes encoding subunits of the proton-pumping NADH:ubiquinone oxidoreductase, or complex I, involved in respiration in mitochondria and eubacteria. From sequence similarities with these genes, the ndh gene products have been suggested to form a large protein complex (Ndh complex); however, the structure and function of this complex remains to be established. Herein we report the isolation of the Ndh complex from the chloroplasts of the higher plant Pisum sativum. The purification procedure involved selective solubilization of the thylakoid membrane with dodecyl maltoside, followed by two anion-exchange chromatography steps and one size-exclusion chromatography step. The isolated Ndh complex has an apparent total molecular mass of approximately 550 kDa and according to SDS͞PAGE consists of at least 16 subunits including NdhA, NdhI, NdhJ, NdhK, and NdhH, which were identified by N-terminal sequencing and immunoblotting. The Ndh complex showed an NADH-and deamino-NADH-specific dehydrogenase activity, characteristic of complex I, when either ferricyanide or the quinones menadione and duroquinone were used as electron acceptors. This study describes the isolation of the chloroplast analogue of the respiratory complex I and provides direct evidence for the function of the plastid Ndh complex as an NADH:plastoquinone oxidoreductase. Our results are compatible with a dual role for the Ndh complex in the chlororespiratory and cyclic photophosphorylation pathways.

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Isolation and Partial Characterization of the NADH Dehydrogenase Complex from Barley Chloroplast Thylakoids

Cited by 23 publications

References 29 publications

Chlororespiration and Poising of Cyclic Electron Transport

Chlororespiration and Poising of Cyclic Electron Transport

A Forward Genetic Screen Identifies Mutants Deficient for Mitochondrial Complex I Assembly in Chlamydomonas reinhardtii

The plastid ndh genes code for an NADH-specific dehydrogenase: Isolation of a complex I analogue from pea thylakoid membranes

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