The Non-Ohmic Proton Leak—25 Years On

1997 ◽  
Vol 17 (3) ◽  
pp. 251-257 ◽  
Author(s):  
David G. Nicholls
Keyword(s):  
Ion Transport ◽  
Respiratory Chain ◽  
Atp Synthesis ◽  
Proton Leak ◽  
Protonmotive Force ◽  
Proton Conductance ◽  
Inner Membrane ◽  
Ohm’S Law ◽  
Mitochondrial Inner Membrane ◽  
Proton Current

The proton conductance of the mitochondrial inner membrane can be quantified by applying Ohm's law to the experimentally determined protonmotive force and the proton current flowing around the proton circuit in the absence of ATP synthesis or ion transport. This last parameter is derived from the rate of State 4 respiration multiplied by the H+/O stoichiometry for the substrate. When the activity of the dehydrogenase supplying electrons to the respiratory chain is progressively increased the proton conductance increases rapidly when the protonmotive force is greater than 220 mV. The consequences of this non-ohmic relationship are discussed.

Targeting Dinitrophenol to Mitochondria: Limitations to the Development of a Self-limiting Mitochondrial Protonophore

2006 ◽  
Vol 26 (3) ◽  
pp. 231-243 ◽  
Author(s):  
Frances H. Blaikie ◽  
Stephanie E. Brown ◽  
Linda M. Samuelsson ◽  
Martin D. Brand ◽  
Robin A. J. Smith ◽  
...  
Keyword(s):  
Fatty Acids ◽  
Membrane Potential ◽  
Oxidative Phosphorylation ◽  
Atp Synthesis ◽  
Proton Leak ◽  
Protonmotive Force ◽  
Inner Membrane ◽  
Atp Production ◽  
Mitochondrial Inner Membrane ◽  
Predominant Component

The protonmotive force (Δp) across the mitochondrial inner membrane drives ATP synthesis. In addition, the energy stored in Δp can be dissipated by proton leak through the inner membrane, contributing to basal metabolic rate and thermogenesis. Increasing mitochondrial proton leak pharmacologically should decrease the efficiency of oxidative phosphorylation and counteract obesity by enabling fatty acids to be oxidised with decreased ATP production. While protonophores such as 2,4-dinitrophenol (DNP) increase mitochondrial proton leak and have been used to treat obesity, a slight increase in DNP concentration above the therapeutically effective dose disrupts mitochondrial function and leads to toxicity. Therefore we set out to develop a less toxic protonophore that would increase proton leak significantly at high Δp but not at low Δp. Our design concept for a potential self-limiting protonophore was to couple the DNP moiety to the lipophilic triphenylphosphonium (TPP) cation and this was achieved by the preparation of 3-(3,5-dinitro-4-hydroxyphenyl)propyltriphenylphosphonium methanesulfonate (MitoDNP). TPP cations accumulate within mitochondria driven by the membrane potential (Δψ), the predominant component of Δp. Our hypothesis was that MitoDNP would accumulate in mitochondria at high Δψ where it would act as a protonophore, but that at lower Δψ the accumulation and uncoupling would be far less. We found that MitoDNP was extensively taken into mitochondria driven by Δψ. However MitoDNP did not uncouple mitochondria as judged by its inability to either increase respiration rate or decrease Δψ. Therefore MitoDNP did not act as a protonophore, probably because the efflux of deprotonated MitoDNP was inhibited.

scholarly journals Experimental discrimination between proton leak and redox slip during mitochondrial electron transport

1994 ◽  
Vol 297 (1) ◽  
pp. 27-29 ◽  
Author(s):  
M D Brand ◽  
L F Chien ◽  
P Diolez
Keyword(s):  
Oxygen Consumption ◽  
Proton Leak ◽  
Protonmotive Force ◽  
Proton Pumps ◽  
Proton Conductance ◽  
Turnover Rates ◽  
High Turnover ◽  
Mitochondrial Inner Membrane ◽  
Rate Of Oxygen Consumption ◽  
The Relationship

By measuring the relationship between protonmotive force and the increment in oxygen consumption by mitochondria treated with submaximal amounts of uncoupler, we have experimentally tested four different models of imperfect coupling of oxidative phosphorylation. The results show that the increased rate of oxygen consumption at high protonmotive force is explained entirely by the dependence on protonmotive force of the passive proton leak conductance of the mitochondrial inner membrane. There is no measurable contribution from redox-slip reactions in the proton pumps caused by high protonmotive force. Neither is there any contribution from increased proton conductance of the membrane or increased redox slip in the respiratory chain caused by high turnover rates of the complexes.

scholarly journals YME1L controls the accumulation of respiratory chain subunits and is required for apoptotic resistance, cristae morphogenesis, and cell proliferation

2012 ◽  
Vol 23 (6) ◽  
pp. 1010-1023 ◽  
Author(s):  
Lukas Stiburek ◽  
Jana Cesnekova ◽  
Olga Kostkova ◽  
Daniela Fornuskova ◽  
Kamila Vinsova ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Membrane Protein ◽  
Respiratory Chain ◽  
Integral Membrane Protein ◽  
Intermembrane Space ◽  
Wild Type ◽  
Inner Membrane ◽  
Mitochondrial Inner Membrane ◽  
Human Orthologue ◽  
Excessive Accumulation

Mitochondrial ATPases associated with diverse cellular activities (AAA) proteases are involved in the quality control and processing of inner-membrane proteins. Here we investigate the cellular activities of YME1L, the human orthologue of the Yme1 subunit of the yeast i‑AAA complex, using stable short hairpin RNA knockdown and expression experiments. Human YME1L is shown to be an integral membrane protein that exposes its carboxy-terminus to the intermembrane space and exists in several complexes of 600–1100 kDa. The stable knockdown of YME1L in human embryonic kidney 293 cells led to impaired cell proliferation and apoptotic resistance, altered cristae morphology, diminished rotenone-sensitive respiration, and increased susceptibility to mitochondrial membrane protein carbonylation. Depletion of YME1L led to excessive accumulation of nonassembled respiratory chain subunits (Ndufb6, ND1, and Cox4) in the inner membrane. This was due to a lack of YME1L proteolytic activity, since the excessive accumulation of subunits was reversed by overexpression of wild-type YME1L but not a proteolytically inactive YME1L variant. Similarly, the expression of wild-type YME1L restored the lamellar cristae morphology of YME1L-deficient mitochondria. Our results demonstrate the importance of mitochondrial inner-membrane proteostasis to both mitochondrial and cellular function and integrity and reveal a novel role for YME1L in the proteolytic regulation of respiratory chain biogenesis.

scholarly journals The translocator maintenance protein Tam41 is required for mitochondrial cardiolipin biosynthesis

2008 ◽  
Vol 183 (7) ◽  
pp. 1213-1221 ◽  
Author(s):  
Stephan Kutik ◽  
Michael Rissler ◽  
Xue Li Guan ◽  
Bernard Guiard ◽  
Guanghou Shui ◽  
...  
Keyword(s):  
Membrane Potential ◽  
Respiratory Chain ◽  
Matrix Protein ◽  
Mitochondrial Protein ◽  
Pleiotropic Effects ◽  
Mitochondrial Matrix ◽  
Carrier Proteins ◽  
Inner Membrane ◽  
Mitochondrial Inner Membrane

The mitochondrial inner membrane contains different translocator systems for the import of presequence-carrying proteins and carrier proteins. The translocator assembly and maintenance protein 41 (Tam41/mitochondrial matrix protein 37) was identified as a new member of the mitochondrial protein translocator systems by its role in maintaining the integrity and activity of the presequence translocase of the inner membrane (TIM23 complex). Here we demonstrate that the assembly of proteins imported by the carrier translocase, TIM22 complex, is even more strongly affected by the lack of Tam41. Moreover, respiratory chain supercomplexes and the inner membrane potential are impaired by lack of Tam41. The phenotype of Tam41-deficient mitochondria thus resembles that of mitochondria lacking cardiolipin. Indeed, we found that Tam41 is required for the biosynthesis of the dimeric phospholipid cardiolipin. The pleiotropic effects of the translocator maintenance protein on preprotein import and respiratory chain can be attributed to its role in biosynthesis of mitochondrial cardiolipin.

scholarly journals Oxygen-pulse curves in rat liver mitochondrial suspensions. Some observations and deductions

1979 ◽  
Vol 180 (1) ◽  
pp. 161-174 ◽  
Author(s):  
G P Archbold ◽  
C L Farrington ◽  
S A Lappin ◽  
A M McKay ◽  
F H Malpress
Keyword(s):  
Rat Liver ◽  
Atp Synthesis ◽  
Liver Mitochondria ◽  
Rat Liver Mitochondria ◽  
Protonmotive Force ◽  
Mitochondrial Inner Membrane ◽  
Fixed Charges ◽  
Charge Concentration ◽  
C 50 ◽  
Proton Movement

1. The inference, implicit in the chemiosmotic hypothesis, that protons move into the bulk phase during ATP synthesis was investigated. 2. Incubation of rat liver mitochondria in the presence of the cation exchanger CM-Sephadex C-50 caused alkalinization in the medium, though total ATP synthesis remained unchanged. The addition of N-ethylmaleimide prevented the alkalinization, but there was still no indication of protons passing into the medium. The expected proton movement [Mitchell & Moyle (1967) Biochem. J. 105, 1147–1162] was readily detected when as an equivalent acid pulse. 3. Analysis of delta H+ decay curves after O2 pulses (3 micrograms-atoms of O/g of protein) indicated the presence of fast and slow components of decay, with first-order rate constants (k) of 0.24s-1 and 0.032s-1. The fast decay was finite and was eliminated in the presence of N-ethylmaleimide. 4. These observations are interpreted as evidence for the development of unmasking of fixed charges on the outer surface of the mitochondrial inner membrane during energization and for the existence of proton-retentive electrical fields (rho-zones) on this surface. The charge concentration is calculated as about 1 charge/10nm2. 5. A cycle of changes in a single fixed-charge molecule is proposed which mediates both Ca2+ uptake and the first step in the utilization of the rho-zone protonmotive force, delta p rho.

scholarly journals Control of respiration and ATP synthesis in mammalian mitochondria and cells

1992 ◽  
Vol 284 (1) ◽  
pp. 1-13 ◽  
Author(s):  
G C Brown
Keyword(s):  
Energy Metabolism ◽  
Oxidative Phosphorylation ◽  
Respiratory Chain ◽  
Atp Synthase ◽  
Energy Charge ◽  
Atp Synthesis ◽  
Basic Mechanism ◽  
Proton Leak ◽  
Control Of Respiration ◽  
Phosphorylation Potential

We have seen that there is no simple answer to the question ‘what controls respiration?’ The answer varies with (a) the size of the system examined (mitochondria, cell or organ), (b) the conditions (rate of ATP use, level of hormonal stimulation), and (c) the particular organ examined. Of the various theories of control of respiration outlined in the introduction the ideas of Chance & Williams (1955, 1956) give the basic mechanism of how respiration is regulated. Increased ATP usage can cause increased respiration and ATP synthesis by mass action in all the main tissues. Superimposed on this basic mechanism is calcium control of matrix dehydrogenases (at least in heart and liver), and possibly also of the respiratory chain (at least in liver) and ATP synthase (at least in heart). In many tissues calcium also stimulates ATP usage directly; thus calcium may stimulate energy metabolism at (at least) four possible sites, the importance of each regulation varying with tissue. Regulation of multiple sites may occur (from a teleological point of view) because: (a) energy metabolism is branched and thus proportionate regulation of branches is required in order to maintain constant fluxes to branches (e.g. to proton leak or different ATP uses); and/or (b) control over fluxes is shared by a number of reactions, so that large increases in flux requires stimulation at multiple sites because each site has relatively little control. Control may be distributed throughout energy metabolism, possibly due to the necessity of minimizing cell protein levels (see Brown, 1991). The idea that energy metabolism is regulated by energy charge (as proposed by Atkinson, 1968, 1977) is misleading in mammals. Neither mitochondrial ATP synthesis nor cellular ATP usage is a unique function of energy charge as AMP is not a significant regulator (see for example Erecinska et al., 1977). The near-equilibrium hypothesis of Klingenberg (1961) and Erecinska & Wilson (1982) is partially correct in that oxidative phosphorylation is often close to equilibrium (apart from cytochrome oxidase) and as a consequence respiration and ATP synthesis are mainly regulated by (a) the phosphorylation potential, and (b) the NADH/NAD+ ratio. However, oxidative phosphorylation is not always close to equilibrium, at least in isolated mitochondria, and relative proximity to equilibrium does not prevent the respiratory chain, the proton leak, the ATP synthase and ANC having significant control over the fluxes. Thus in some conditions respiration rate correlates better with [ADP] than with phosphorylation potential, and may be relatively insensitive to mitochondrial NADH/NAD+ ratio.(ABSTRACT TRUNCATED AT 400 WORDS)

scholarly journals Effects of magnesium and nucleotides on the proton conductance of rat skeletal-muscle mitochondria

2000 ◽  
Vol 348 (1) ◽  
pp. 209-213 ◽  
Author(s):  
Susana CADENAS ◽  
Martin D. BRAND
Keyword(s):  
Skeletal Muscle ◽  
Atp Synthase ◽  
Atp Synthesis ◽  
Proton Conductance ◽  
Rat Skeletal Muscle ◽  
Maximal Inhibition ◽  
Muscle Mitochondria ◽  
Mitochondrial Inner Membrane ◽  
Skeletal Muscle Mitochondria ◽  
The Matrix

During oxidative phosphorylation most of the protons pumped out to the cytosol across the mitochondrial inner membrane return to the matrix through the ATP synthase, driving ATP synthesis. However, some of them leak back to the matrix through a proton-conductance pathway in the membrane. When the ATP synthase is inhibited with oligomycin and ATP is not being synthesized, all of the respiration is used to drive the proton leak. We report here that Mg2+ inhibits the proton conductance in rat skeletal-muscle mitochondria. Addition of Mg2+ inhibited both oligomycin-inhibited respiration and the proton conductance, while removal of Mg2+ using EDTA activated these processes. The proton conductance was inhibited by more than 80% as free Mg2+ was raised from 25 nM to 220 μM. Half-maximal inhibition occurred at about 1 μM free Mg2+, which is close to the contaminating free Mg2+ concentration in our incubations in the absence of added magnesium chelators. ATP, GTP, CTP, TTP or UTP at a concentration of 1 mM increased the oligomycin-inhibited respiration rate by about 50%. However, these NTP effects were abolished by addition of 2 mM Mg2+ and any NTP-stimulated proton conductance was explained completely by chelation of endogenous free Mg2+. The corresponding nucleoside diphosphates (ADP, GDP, CDP, TDP or UDP) at 1 mM had no effect on oligomycin-inhibited respiration. We conclude that proton conductance in rat skeletal-muscle mitochondria is very sensitive to free Mg2+ concentration but is insensitive to NTPs or NDPs at 1 mM.

The Regulation and Physiology of Mitochondrial Proton Leak

Physiology ◽  
2011 ◽  
Vol 26 (3) ◽  
pp. 192-205 ◽  
Author(s):  
Ajit S. Divakaruni ◽  
Martin D. Brand
Keyword(s):  
Coupling Efficiency ◽  
Atp Synthesis ◽  
Uncoupling Proteins ◽  
Proton Gradient ◽  
Physiological Significance ◽  
Proton Leak ◽  
Inner Membrane ◽  
Related Disease ◽  
Electrochemical Proton Gradient ◽  
Age Related

Mitochondria couple respiration to ATP synthesis through an electrochemical proton gradient. Proton leak across the inner membrane allows adjustment of the coupling efficiency. The aim of this review is threefold: 1) introduce the unfamiliar reader to proton leak and its physiological significance, 2) review the role and regulation of uncoupling proteins, and 3) outline the prospects of proton leak as an avenue to treat obesity, diabetes, and age-related disease.

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