bims-cytox1 Biomed news
on Cytochrome oxidase subunit 1
Issue of 2018‒05‒06
five papers selected by
Gavin McStay
New York Institute of Technology


  1. Biochim Biophys Acta. 2018 Apr 25. pii: S0005-2728(18)30083-5. [Epub ahead of print]1859(8): 555-566
    Rocha MC, Springett R.
      Cytochrome oxidase is the terminal oxidase of the mitochondrial electron transport chain and pumps 4 protons per oxygen reduced to water. Spectral shifts in the α-band of heme a have been observed in multiple studies and these shifts have the potential to shed light on the proton pumping intermediates. Previously we found that heme a had two spectral components in the α-band during redox titrations in living RAW 264.7 mouse macrophage cells, the classical 605 nm form and a blue-shifted 602 nm form. To confirm these spectral changes were not an artifact due to the complex milieu of the living cell, redox titrations were performed in the isolated detergent-solubilized bovine enzyme from both the Soret- and α-band using precise multiwavelength spectroscopy. This data verified the presence of the 602 nm form in the α-band, revealed a similar shift of heme a in the Soret-band and ruled out the reversal of calcium binding as the origin of the blue shift. The 602 nm form was found to be stabilized at high pH or by binding of azide, which is known to blue shift the α-band of heme a. Azide also stabilized the 602 nm form in the living cells. It is concluded there is a form of cytochrome oxidase in which heme a undergoes a blue shift to a 602 nm form and that redox titrations can be successfully performed in living cells where the oxidase operates in its authentic environment and in the presence of a proton motive force.
    Keywords:  Azide; Cytochrome oxidase; Heme spectroscopy; Midpoint potential; Proton pumping; Redox titration
    DOI:  https://doi.org/10.1016/j.bbabio.2018.04.009
  2. Redox Biol. 2018 Mar 22. pii: S2213-2317(18)30147-2. [Epub ahead of print]17 207-212
    García-Roche M, Casal A, Carriquiry M, Radi R, Quijano C, Cassina A.
      The aim of this work was to develop a cryopreservation method of small liver biopsies for in situ mitochondrial function assessment. Herein we describe a detailed protocol for tissue collection, cryopreservation, high-resolution respirometry using complex I and II substrates, calculation and interpretation of respiratory parameters. Liver biopsies from cow and rat were sequentially frozen in a medium containing dimethylsulfoxide as cryoprotectant and stored for up to 3 months at -80 °C. Oxygen consumption rate studies of fresh and cryopreserved samples revealed that most respiratory parameters remained unchanged. Additionally, outer mitochondrial membrane integrity was assessed adding cytochrome c, proving that our cryopreservation method does not harm mitochondrial structure. In sum, we present a reliable way to cryopreserve small liver biopsies without affecting mitochondrial function. Our protocol will enable the transport and storage of samples, extending and facilitating mitochondrial function analysis of liver biopsies.
    Keywords:  Biopsy; Cryopreservation; High-resolution respirometry; Mitochondria; Mitochondrial function; Oxygen consumption rate
    DOI:  https://doi.org/10.1016/j.redox.2018.03.008
  3. Biochim Biophys Acta. 2018 Apr 25. pii: S0005-2728(18)30082-3. [Epub ahead of print]
    Dreinert A, Wolf A, Mentzel T, Meunier B, Fehr M.
      Ametoctradin is an agricultural fungicide that selectively inhibits the cytochrome bc1 complex of oomycetes. Previous spectrophotometric studies using the purified cytochrome bc1 complex from Pythium sp. showed that Ametoctradin binds to the Qo-site of the enzyme. However, as modeling studies suggested a binding mode like that of the substrate ubiquinol, the possibility for a dual Qo- and Qi-site binding mode was left open. In this work, binding studies and enzyme assays with mitochondrial membrane preparations from Pythium sp. and an S. cerevisiae strain with a modified Qi-site were used to investigate further the binding mode of Ametoctradin. The results obtained argue that the compound could bind to both the Qo- and Qi-sites of the cytochrome bc1 complex and that its position or binding pose in the Qi-site differs from that of Cyazofamid and Amisulbrom, the two Qi-site-targeting, anti-oomycetes compounds. Furthermore, the data support the argument that Ametoctradin prefers binding to the reduced cytochrome bc1 complex. Thus, Ametoctradin has an unusual binding mode and further studies with this compound may offer the opportunity to better understand the catalytic cycle of the cytochrome bc1 complex.
    Keywords:  Ametoctradin; Amisulbrom; Cyazofamid; Cytochrome bc(1) complex; Initium; Oomycetes; Respiration inhibitor; Respiratory complex III
    DOI:  https://doi.org/10.1016/j.bbabio.2018.04.008
  4. Redox Biol. 2018 Mar 24. pii: S2213-2317(18)30218-0. [Epub ahead of print]17 200-206
    Erdogan AJ, Ali M, Habich M, Salscheider SL, Schu L, Petrungaro C, Thomas LW, Ashcroft M, Leichert LI, Roma LP, Riemer J.
      Disulfide formation in the mitochondrial intermembrane space is an essential process catalyzed by a disulfide relay machinery. In mammalian cells, the key enzyme in this machinery is the oxidoreductase CHCHD4/Mia40. Here, we determined the in vivo CHCHD4 redox state, which is the major determinant of its cellular activity. We found that under basal conditions, endogenous CHCHD4 redox state in cultured cells and mouse tissues was predominantly oxidized, however, degrees of oxidation in different tissues varied from 70% to 90% oxidized. To test whether differences in the ratio between CHCHD4 and ALR might explain tissue-specific differences in the CHCHD4 redox state, we determined the molar ratio of both proteins in different mouse tissues. Surprisingly, ALR is superstoichiometric over CHCHD4 in most tissues. However, the levels of CHCHD4 and the ratio of ALR over CHCHD4 appear to correlate only weakly with the redox state, and although ALR is present in superstoichiometric amounts, it does not lead to fully oxidized CHCHD4.
    Keywords:  CHCHD4/ALR/disulfide/redox/mitochondria
    DOI:  https://doi.org/10.1016/j.redox.2018.03.014
  5. Neurochem Int. 2018 Apr 25. pii: S0197-0186(18)30090-1. [Epub ahead of print]
    Zilocchi M, Finzi G, Lualdi M, Sessa F, Fasano M, Alberio T.
      Mitochondrial impairment is one of the most important hallmarks of Parkinson's disease (PD) pathogenesis. In this work, we wanted to verify the molecular basis of altered mitochondrial dynamics and disposal in Substantia nigra specimens of sporadic PD patients, by the comparison with two cellular models of PD. Indeed, SH-SY5Y cells were treated with either dopamine or 1-methyl-4-phenylpyridinium (MPP+) in order to highlight the effect of altered dopamine homeostasis and of complex I inhibition, respectively. As a result, we found that fusion impairment of the inner mitochondrial membrane is a common feature of both PD human samples and cellular models. However, the effects of dopamine and MPP+ treatments resulted to be different in terms of the mitochondrial damage induced. Opposite changes in the levels of two mitochondrial protein markers (voltage-dependent anion channels (VDACs) and cytochrome c oxidase subunit 5β (COX5β)) were observed. In this case, dopamine treatment better recapitulated the molecular picture of patients' samples. Moreover, the accumulation of PTEN-induced putative kinase 1 (PINK1), a mitophagy marker, was not observed in both PD patients samples and cellular models. Eventually, in transmission electron microscopy images, small electron dense deposits were observed in mitochondria of PD subjects, which are uniquely reproduced in dopamine-treated cells. In conclusion, our study suggests that the mitochondrial molecular landscape of Substantia nigra specimens of PD patients can be mirrored by the impaired dopamine homeostasis cellular model, thus supporting the hypothesis that alterations in this process could be a crucial pathogenetic event in PD.
    Keywords:  Dopamine; MPP(+); Mitochondria; Parkinson's disease; Substantia nigra
    DOI:  https://doi.org/10.1016/j.neuint.2018.04.013