bims-cytox1 Biomed News
on Cytochrome oxidase subunit 1
Issue of 2020‒11‒29
nine papers selected by
Gavin McStay
Staffordshire University
  1. Clinical features of mtDNA-related syndromes in adulthood.
  2. Visualizing Mitochondrial Ribosomal RNA and Mitochondrial Protein Synthesis in Human Cell Lines.
  3. The Analysis of Yeast Mitochondrial Translation.
  4. Blue-Native Electrophoresis to Study the OXPHOS Complexes.
  5. Essential role of accessory subunit LYRM6 in the mechanism of mitochondrial complex I.
  6. Elongational stalling activates mitoribosome-associated quality control.
  7. Mitochondrial Complexome Profiling.
  8. Analysis of Human Mutations in the Supernumerary Subunits of Complex I.
  9. Identification of Putative Mitochondrial Protease Substrates.
  1. Arch Biochem Biophys. 2020 Nov 20. pii: S0003-9861(20)30698-6. [Epub ahead of print] 108689
    Clinical features of mtDNA-related syndromes in adulthood.
    Montano V, Gruosso F, Simoncini C, Siciliano G, Mancuso M.
      Mitochondrial diseases are the most common inheritable metabolic diseases, due to defects in oxidative phosphorylation. They are caused by mutations of nuclear or mitochondrial DNA in genes involved in mitochondrial function. The peculiarity of "mitochondrial DNA genetics rules"in part explains the marked phenotypic variability, the complexity of genotype-phenotype correlations and the challenge of genetic counseling. The new massive genetic sequencing technologies have changed the diagnostic approach, enhancing mitochondrial DNA-related syndromes diagnosis and often avoiding the need of a tissue biopsy. Here we present the most common phenotypes associated with a mitochondrial DNA mutation with the recent advances in diagnosis and in therapeutic perspectives.
    Keywords:  MELAS; MERRF; Mitochondrial disorders; Primary mitochondrial myopathy
    DOI:  https://doi.org/10.1016/j.abb.2020.108689
  2. Methods Mol Biol. 2021 ;2192 159-181
    Visualizing Mitochondrial Ribosomal RNA and Mitochondrial Protein Synthesis in Human Cell Lines.
    Matthew Zorkau, Yasmin Proctor-Kent, Rolando Berlinguer-Palmini, Andrew Hamilton, Zofia M Chrzanowska-Lightowlers, Robert N Lightowlers.
      Human mitochondria contain their own DNA (mtDNA) that encodes 13 proteins all of which are core subunits of oxidative phosphorylation (OXPHOS) complexes. To form functional complexes, these 13 components need to be correctly assembled with approximately 70 nuclear-encoded subunits that are imported following synthesis in the cytosol. How this complicated coordinated translation and assembly is choreographed is still not clear. Methods are being developed to determine whether all members of a particular complex are translated in close proximity, whether protein synthesis is clustered in submitochondrial factories, whether these align with incoming polypeptides, and if there is evidence for co-translational translation that is regulated and limited by the interaction of the incoming proteins with synthesis of their mtDNA-encoded partners. Two methods are described in this chapter to visualize the distribution of mitochondrial ribosomal RNAs in conjunction with newly synthesized mitochondrial proteins. The first combines RNA Fluorescent In Situ Hybridization (FISH) and super-resolution immunocytochemistry to pinpoint mitochondrial ribosomal RNA. The second localizes nascent translation within the mitochondrial network through non-canonical amino acid labeling, click chemistry and fluorescent microscopy.
    Keywords:  Click chemistry; Fluorescence microscopy; Mitochondria; Mitochondrial RNA; Mitoribosome; Single-molecule RNA FISH; Stimulated emission depletion microscopy; Super-resolution microscopy; Translation
    DOI:  https://doi.org/10.1007/978-1-0716-0834-0_13
  3. Methods Mol Biol. 2021 ;2192 227-242
    The Analysis of Yeast Mitochondrial Translation.
    Andreas Carlström, Magdalena Rzepka, Martin Ott.
      The mitochondrial genome encodes only a handful of proteins, but methods to track their synthesis are highly limited. Saccharomyces cerevisiae is a model organism that offers possibilities to expand the classical systems to analyze mitochondrial translation. In this chapter, we present two approaches of monitoring mitochondrial protein synthesis. Labeling of mitochondrially translated products with radioactive amino acids can be performed either in intact cells or in isolated mitochondria. However, these classical methods have disadvantages that can affect cell physiology and hence are not suitable for all types of research questions. Some of these limitations can be overcome by the use of reporter genes that are inserted into yeast genetic screens mitochondrial DNA via biolistic transformation. These reporter genes can be used for yeast genetic screen and to monitor regulation and efficiency of mitochondrial translation with a variety of methods.
    Keywords:  Mitochondria; Protein synthesis; Reporter genes; Translation; Yeast
    DOI:  https://doi.org/10.1007/978-1-0716-0834-0_17
  4. Methods Mol Biol. 2021 ;2192 287-311
    Blue-Native Electrophoresis to Study the OXPHOS Complexes.
    Erika Fernandez-Vizarra, Massimo Zeviani.
      Blue-native polyacrylamide gel electrophoresis (BN-PAGE) is a technique optimized for the analysis of the five components of the mitochondrial oxidative phosphorylation (OXPHOS) system. BN-PAGE is based on the preservation of the interactions between the individual subunits within the integral complexes. To achieve this, the complexes are extracted from the mitochondrial inner membrane using mild detergents and separated by electrophoresis in the absence of denaturing agents. The electrophoretic procedures can then be combined with a variety of downstream detection techniques. Since its development in the 1990s, BN-PAGE has been applied in the study of mitochondria from all kinds of organisms and extensive amounts of data have been produced using this technique, being key for the understanding of many aspects of OXPHOS physiopathology.
    Keywords:  Blue-native gel electrophoresis; First-dimension BN-PAGE; In gel activity assays; Mitochondrial complexes I, II, III, IV, and V; Oxidative phosphorylation system; Second-dimension BN-PAGE
    DOI:  https://doi.org/10.1007/978-1-0716-0834-0_20
  5. Nat Commun. 2020 Nov 26. 11(1): 6008
    Essential role of accessory subunit LYRM6 in the mechanism of mitochondrial complex I.
    Etienne Galemou Yoga, Kristian Parey, Amina Djurabekova, Outi Haapanen, Karin Siegmund, Klaus Zwicker, Vivek Sharma, Volker Zickermann, Heike Angerer.
      Respiratory complex I catalyzes electron transfer from NADH to ubiquinone (Q) coupled to vectorial proton translocation across the inner mitochondrial membrane. Despite recent progress in structure determination of this very large membrane protein complex, the coupling mechanism is a matter of ongoing debate and the function of accessory subunits surrounding the canonical core subunits is essentially unknown. Concerted rearrangements within a cluster of conserved loops of central subunits NDUFS2 (β1-β2S2 loop), ND1 (TMH5-6ND1 loop) and ND3 (TMH1-2ND3 loop) were suggested to be critical for its proton pumping mechanism. Here, we show that stabilization of the TMH1-2ND3 loop by accessory subunit LYRM6 (NDUFA6) is pivotal for energy conversion by mitochondrial complex I. We determined the high-resolution structure of inactive mutant F89ALYRM6 of eukaryotic complex I from the yeast Yarrowia lipolytica and found long-range structural changes affecting the entire loop cluster. In atomistic molecular dynamics simulations of the mutant, we observed conformational transitions in the loop cluster that disrupted a putative pathway for delivery of substrate protons required in Q redox chemistry. Our results elucidate in detail the essential role of accessory subunit LYRM6 for the function of eukaryotic complex I and offer clues on its redox-linked proton pumping mechanism.
    DOI:  https://doi.org/10.1038/s41467-020-19778-7
  6. Science. 2020 Nov 27. 370(6520): 1105-1110
    Elongational stalling activates mitoribosome-associated quality control.
    Nirupa Desai, Hanting Yang, Viswanathan Chandrasekaran, Razina Kazi, Michal Minczuk, V Ramakrishnan.
      The human mitochondrial ribosome (mitoribosome) and associated proteins regulate the synthesis of 13 essential subunits of the oxidative phosphorylation complexes. We report the discovery of a mitoribosome-associated quality control pathway that responds to interruptions during elongation, and we present structures at 3.1- to 3.3-angstrom resolution of mitoribosomal large subunits trapped during ribosome rescue. Release factor homolog C12orf65 (mtRF-R) and RNA binding protein C6orf203 (MTRES1) eject the nascent chain and peptidyl transfer RNA (tRNA), respectively, from stalled ribosomes. Recruitment of mitoribosome biogenesis factors to these quality control intermediates suggests additional roles for these factors during mitoribosome rescue. We also report related cryo-electron microscopy structures (3.7 to 4.4 angstrom resolution) of elongating mitoribosomes bound to tRNAs, nascent polypeptides, the guanosine triphosphatase elongation factors mtEF-Tu and mtEF-G1, and the Oxa1L translocase.
    DOI:  https://doi.org/10.1126/science.abc7782
  7. Methods Mol Biol. 2021 ;2192 269-285
    Mitochondrial Complexome Profiling.
    Heiko Giese, Jana Meisterknecht, Juliana Heidler, Ilka Wittig.
      Complexome profiling combines blue native gel electrophoresis (BNE) and quantitative mass spectrometry to define an entire protein interactome of a cell, an organelle, or a biological membrane preparation. The method allows the identification of protein assemblies with low abundance and detects dynamic processes of protein complex assembly. Applications of complexome profiling range from the determination of complex subunit compositions, assembly of single protein complexes, and supercomplexes to comprehensive differential studies between patients or disease models. This chapter describes the workflow of complexome profiling from sample preparation, mass spectrometry to data analysis with a bioinformatics tool.
    Keywords:  Assembly; Blue native electrophoresis; Complexome profiling; Mass spectrometry; Membrane protein complexes; Mitochondria
    DOI:  https://doi.org/10.1007/978-1-0716-0834-0_19
  8. Life (Basel). 2020 Nov 20. pii: E296. [Epub ahead of print]10(11):
    Analysis of Human Mutations in the Supernumerary Subunits of Complex I.
    Quynh-Chi L Dang, Duong H Phan, Abigail N Johnson, Mukund Pasapuleti, Hind A Alkhaldi, Fang Zhang, Steven B Vik.
      Complex I is the largest member of the electron transport chain in human mitochondria. It comprises 45 subunits and requires at least 15 assembly factors. The subunits can be divided into 14 "core" subunits that carry out oxidation-reduction reactions and proton translocation, as well as 31 additional supernumerary (or accessory) subunits whose functions are less well known. Diminished levels of complex I activity are seen in many mitochondrial disease states. This review seeks to tabulate mutations in the supernumerary subunits of humans that appear to cause disease. Mutations in 20 of the supernumerary subunits have been identified. The mutations were analyzed in light of the tertiary and quaternary structure of human complex I (PDB id = 5xtd). Mutations were found that might disrupt the folding of that subunit or that would weaken binding to another subunit. In some cases, it appeared that no protein was made or, at least, could not be detected. A very common outcome is the lack of assembly of complex I when supernumerary subunits are mutated or missing. We suggest that poor assembly is the result of disrupting the large network of subunit interactions that the supernumerary subunits typically engage in.
    Keywords:  Leigh syndrome; NADH dehydrogenase; complex I assembly; complex I deficiency; complex I structure; electron transport chain; mammalian complex I; mitochondria; mitochondrial dysfunction; supernumerary subunits
    DOI:  https://doi.org/10.3390/life10110296
  9. Methods Mol Biol. 2021 ;2192 313-329
    Identification of Putative Mitochondrial Protease Substrates.
    Eduard Hofsetz, Pitter F Huesgen, Aleksandra Trifunovic.
      Mitochondrial proteases constitute a fundamental part of the organellar protein quality control system to ensure the timely removal of damaged or obsolete proteins. The analysis of proteases is often limited to the identification of bona fide substrates that are degraded in the presence and become more abundant in the absence of the respective protease. However, proteases in numerous organisms from bacteria to humans can process specific substrates to release shortened proteins with potentially altered activities. Here, we describe an adaptation of the substrate-trapping approach, as well as the N-terminal profiling protocol Terminal Amine Isotope Labeling of Substrates (TAILS) for the identification of bona fide substrates and mitochondrial proteins that undergo complete or partial proteolysis.
    Keywords:  Immunoprecipitations; Mitochondria; N Termini profiling; Proteases; Proteolysis; Substrate-trapping; Substrates; TAILS
    DOI:  https://doi.org/10.1007/978-1-0716-0834-0_21
This page is being maintained by Thomas Krichel. It was last updated on 2021‒07‒23 at 10:19:28.