The 29.9kDa Subunit of Mitochondrial Complex I is Involved in the Enzyme Active/De-active Transitions

Ushakova, Alexandra V.; Duarte, Margarida; Vinogradov, Andrei D.; Videira, Arnaldo

doi:10.1016/j.jmb.2005.06.005

Cited by 15 publications

(17 citation statements)

References 40 publications

(48 reference statements)

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“…6C, lower panel), where the most intense ones were near the top of the first-dimension gel. Such high-molecular-weight species of membrane subunits were previously observed after density gradient centrifugation of Triton X-100 extracts from mitochondria of peripheral arm mutants like nuo30.4 (22), but also from nuo29.9 (87), which generates a surplus of the membrane assembly intermediate (84) as mentioned above. Most probably, the largest 20.8-kDa species represented unspecific aggregates of partially assembled hydrophobic membrane subunits, as previously suggested (22,23).…”

supporting

confidence: 55%

“…This is the first direct demonstration that the incorporation of complex I into supercomplexes as part of a proposed supramolecular network has major nonrespiratory significance and cannot be solely related to conceivable enzymatic advantages, like substrate channeling. Furthermore, the nuo29.9 mutant can assemble only low amounts of complex I, about 20% of that of the wild type (23,87), which is nonetheless incorporated into supercomplexes as in wild-type, nuo21, and nuo51 strains (Fig. 6B).…”

Section: Discussionmentioning

confidence: 95%

“…Their mitochondrial detergent extracts were analyzed in the same manner as those of the wild type through native gel electrophoresis. The mutants nuo21, nuo29.9, and nuo51 are able to assemble an intact complex I lacking the respective peripheral arm subunit, although the nuo29.9 mutant assembles reduced amounts (approximately 20%) of complex I (23,28,29,87). The fourth peripheral arm mutant used in this study, nuo30.4, cannot assemble a peripheral arm and accumulates membrane arm intermediates (22).…”

Section: Vol 6 2007 Inactive Respiratory Supercomplexes In a Nuo51 mentioning

confidence: 93%

“…6B, lower panel). Notably, the signal of the 30.4-kDa subunit was rather weak, since this protein, located in close vicinity to the 29.9-kDa subunit, is strongly reduced in the mutant nuo29.9 (87). No subcomplexes of complex I could be immunodetected, indicating that in the nuo29.9 mutant essentially all of the assembled peripheral arm joins with the membrane arm, occurring in surplus quantity, to form "complete" complex I. Additionally, it is likely that the excess hydrophobic membrane arm forms low-mobility membrane aggregates that contribute to the immunosignals in Fig.…”

Section: Vol 6 2007 Inactive Respiratory Supercomplexes In a Nuo51 mentioning

confidence: 97%

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Supramolecular Organization of the Respiratory Chain in Neurospora crassa Mitochondria

et al. 2007

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The existence of specific respiratory supercomplexes in mitochondria of most organisms has gained much momentum. However, its functional significance is still poorly understood. The availability of many deletion mutants in complex I (NADH:ubiquinone oxidoreductase) of Neurospora crassa, distinctly affected in the assembly process, offers unique opportunities to analyze the biogenesis of respiratory supercomplexes. Herein, we describe the role of complex I in assembly of respiratory complexes and supercomplexes as suggested by blue and colorless native polyacrylamide gel electrophoresis and mass spectrometry analyses of mildly solubilized mitochondria from the wild type and eight deletion mutants.As an important refinement of the fungal respirasome model, we found that the standard respiratory chain of N. crassa comprises putative complex I dimers in addition to I-III-IV and III-IV supercomplexes. Three Neurospora mutants able to assemble a complete complex I, lacking only the disrupted subunit, have respiratory supercomplexes, in particular I-III-IV supercomplexes and complex I dimers, like the wild-type strain. Furthermore, we were able to detect the I-III-IV supercomplexes in the nuo51 mutant with no overall enzymatic activity, representing the first example of inactive respirasomes. In addition, III-IV supercomplexes were also present in strains lacking an assembled complex I, namely, in four membrane arm subunit mutants as well as in the peripheral arm nuo30.4 mutant. In membrane arm mutants, high-molecular-mass species of the 30.4-kDa peripheral arm subunit comigrating with III-IV supercomplexes and/or the prohibitin complex were detected. The data presented herein suggest that the biogenesis of complex I is linked with its assembly into supercomplexes.

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confidence: 55%

Section: Discussionmentioning

confidence: 95%

Section: Vol 6 2007 Inactive Respiratory Supercomplexes In a Nuo51 mentioning

confidence: 93%

Section: Vol 6 2007 Inactive Respiratory Supercomplexes In a Nuo51 mentioning

confidence: 97%

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Supramolecular Organization of the Respiratory Chain in Neurospora crassa Mitochondria

et al. 2007

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“…Therefore, it seems likely that this transition involves at least one of the accessory subunits, e.g. 39 kDa (NDUFA9) [32], B14 (NDUFA6) and SDAP-a (NDUFAB1) [34] or B13 (NDUFA5) [35] which are located in close proximity to the hydrophilic loop of ND3 [23,24,28]. At least two different conformational states of complex I were observed in recent cryo-electron microscopy studies [23,24].…”

Section: Mitochondrial Complex Imentioning

confidence: 97%

Modulation of the conformational state of mitochondrial complex I as a target for therapeutic intervention

Galkin

Moncada

2017

Interface Focus.

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In recent years, there have been significant advances in our understanding of the functions of mitochondrial complex I other than the generation of energy. These include its role in generation of reactive oxygen species, involvement in the hypoxic tissue response and its possible regulation by nitric oxide (NO) metabolites. In this review, we will focus on the hypoxic conformational change of this mitochondrial enzyme, the so-called active/deactive transition. This conformational change is physiological and relevant to the understanding of certain pathological conditions including, in the cardiovascular system, ischaemia/reperfusion (I/R) damage. We will discuss how complex I can be affected by NO metabolites and will outline some potential mitochondria-targeted therapies in I/R damage.

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Topography and chemical reactivity of the active–inactive transition-sensitive SH-group in the mitochondrial NADH:ubiquinone oxidoreductase (Complex I)

Gostimskaya

Cecchini

Vinogradov

2006

Biochimica et Biophysica Acta (BBA) - Bioenergetics

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The spatial arrangement and chemical reactivity of the activation-dependent thiol in the mitochondrial Complex I was studied using the membrane penetrating N-ethylmaleimide (NEM) and non-penetrating anionic 5,5'-dithiobis-(2-nitrobenzoate) (DTNB) as the specific inhibitors of the enzyme in mitochondria and inside-out submitochondrial particles (SMP). Both NEM and DTNB rapidly inhibited the de-activated Complex I in SMP. In mitochondria NEM caused rapid inhibition of Complex I, whereas the enzyme activity was insensitive to DTNB. In the presence of the channel-forming antibiotic alamethicin, mitochondrial Complex I became sensitive to DTNB. Neither active nor de-activated Complex I in SMP was inhibited by oxidized glutathione (10 mM, pH 8.0, 75 min). The data suggest that the active/de-active transition sulfhydryl group of Complex I which is sensitive to inhibition by NEM is located at the inner membrane-matrix interface. These data include the sidedness dependency of inhibition, effect of pH, ionic strength, and membrane bilayer modification on enzyme reactivity towards DTNB and its neutral analogue.

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The 29.9kDa Subunit of Mitochondrial Complex I is Involved in the Enzyme Active/De-active Transitions

Cited by 15 publications

References 40 publications

Supramolecular Organization of the Respiratory Chain in Neurospora crassa Mitochondria

Supramolecular Organization of the Respiratory Chain in Neurospora crassa Mitochondria

Modulation of the conformational state of mitochondrial complex I as a target for therapeutic intervention

Topography and chemical reactivity of the active–inactive transition-sensitive SH-group in the mitochondrial NADH:ubiquinone oxidoreductase (Complex I)

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