Temperature-dependent changes in respiration rates and redox poise of the ubiquinone pool in protoplasts and isolated mitochondria of potato leaves

In many environments, leaves experience large diurnal variations in temperature. Such short‐term changes in temperature are likely to have important implications for respiratory metabolism in leaves. Here, we used intact leaf, protoplasts and isolated mitochondria to determine the impact of short‐term changes in temperature on respiration rates (R), adenylate concentrations and the redox poise of the ubiquinone (UQ) pool in mitochondria of potato leaves. The Q₁₀ (i.e. proportional change in R for each 10°C rise in temperature) of respiration was 1.8, both for intact leaves and protoplasts. In protoplasts, the redox poise of the extracted UQ pool (UQ_R/UQ_T) increased from 0.33 at 22°C, to 0.76 at 15°C. Further decreases in temperature (from 15 to 5°C) resulted in UQ_R/UQ_T decreasing to 0.40. Adenylate ratios in protoplasts were also temperature dependent. At high adenosine 5′‐triphosphate (ATP) adenosine 5′‐diphosphate (ADP) ratios (i.e. low ADP concentrations), UQ_R/UQ_T values were low, suggesting that adenylates restricted flux via the UQ‐reducing pathways more than they restricted flux via pathways that oxidized UQH₂. To assess whether high rates of alternative oxidase (AOX) activity could have uncoupled respiratory flux (and thus UQ_R/UQ_T) from adenylate restriction of the cytochrome (Cyt) pathway, we constructed kinetic curves of O₂ uptake (via the two pathways) vs UQ_R/UQ_T in isolated mitochondria, measured at two temperatures (15 and 25°C); measurements were made for mitochondria operating under state 3 (i.e. +ADP) and state 4 (i.e. −ADP) conditions. In contrast to the Cyt pathway, flux via the AOX was temperature insensitive, with maximal rates of AOX activity representing 21–57% of total O₂ uptake in isolated mitochondria. We conclude that temperature‐dependent variations in UQ_R/UQ_T are largely dependent on temperature‐dependent changes in adenylate ratios, and that flux via the AOX could in some circumstances help reduce maximal UQ values.     

Low temperature effects on ubiquinone content, respiration rates and lipid peroxidation levels of etiolated seedlings of two differentially chilling-sensitive species

Ubiquinone functions primarily in the electron transport chain of the mitochondria of plants and animals. Secondary roles in plant tissue, such as antioxidant activity, have also been proposed. The effect of low temperature exposure on etiolated seedling embryonic axes of two differentially chilling-sensitive species, mung bean ( Vigna radiata L.) (chilling-sensitive) and pea ( Pisum sativum L. cv. Lincoln) (chilling-tolerant) with respect to respiration rate, lipid peroxidation and ubiquinone content was examined. Whole seedlings (embryonic axis and cotyledon) of both species were exposed to control temperatures (20°C) (6 days) or an acclimatory low temperature treatment of 10°C (3 days) followed by exposure at 5°C (3 days). Measurements were initiated 3 days after seedlings had reached 50% germination (D0). Prior to measurements the cotyledons were removed and only the embryonic axis was used in these experiments. Ubiquinol (UQH₂), ubiquinone (UQ) and total ubiquinone (UQ_tot) content decreased in mung bean in response to the temperature treatment and UQH₂ and UQ_tot remained stable in the more chilling-tolerant pea. The reduction of the total Q-pool was approximately 85–92%, suggesting a high degree of saturation of the respiration pathways. Respiration declined and the RQ ratio increased in both species in response to low temperature. Cytochrome c oxidase (COX) (EC 1.9.3.1) activity was higher in pea than in mung bean but decreased during low temperature exposure in both species. Considering that levels of MDA (lipid peroxidation) did not increase in either species in response to chilling, decreased levels of UQH₂ and UQ observed in chilling-sensitive mung bean may indicate that these compounds were damaged prior to other membrane lipids during low temperature treatment and rendered undetectable.     

Variety-specific response of wheat (<Emphasis Type="Italic">Triticum aestivum</Emphasis> L.) leaf mitochondria to drought stress

The main objective of the present work was to examine leaf respiratory responses to dehydration and subsequent recovery in three varieties of winter wheat (Triticum aestivum L.) known to differ in their level of drought tolerance. Under dehydration, both total respiration and salicylhydroxamic acid (SHAM)-resistant cytochrome (Cyt) pathway respiration by leaf segments decreased significantly compared with well-watered plants. This decrease was more pronounced in the drought-sensitive Sadovo and Prelom genotypes. In contrast, the KCN-resistant SHAM-sensitive alternative (Alt) pathway became increasingly engaged, and accounted for about 80% of the total respiration. In the drought-tolerant Katya variety, increased contribution of the Alt pathway was accompanied by a slight decrease in Cyt pathway activity. Respiration of isolated leaf mitochondria also showed a variety-specific drought response. Mitochondria from drought-sensitive genotypes had low oxidative phosphorylation efficiency after dehydration and rewatering, whereas the drought-tolerant Katya mitochondria showed higher phosphorylation rates. Morphometric analysis of leaf ultrastructure revealed that mitochondria occupied approximately 7% of the cell area in control plants. Under dehydration, in the drought-sensitive varieties this area was reduced to about 2.0%, whereas in Katya it was around 6.0%. The results are discussed in terms of possible mechanisms underlying variety-specific mitochondrial responses to dehydration.     

Interactions Between the Cytochrome Pathway and the Alternative Oxidase in Isolated Acanthamoeba castellanii Mitochondria

The steady-state activity of the two quinol-oxidizing pathways of Acanthamoeba castellanii mitochondria, the phosphorylating cytochrome pathway (i.e. the benzohydroxamate(BHAM)-resistant respiration in state 3) and the alternative oxidase (i.e. the KCN-resistant respiration), is shown to be fixed by ubiquinone (Q) pool redox state independently of the reducing substrate (succinate or exogenous reduced nicotinamide adenine dinucleotide (NADH)), indicating that the active Q pool is homogenous. For both pathways, activity increases with the Q reduction level (up to 80%). However, the cytochrome pathway respiration partially inhibited (about 50%) by myxothiazol decreases when the Q reduction level increases above 80%. The decrease can be explained by the Q cycle mechanism of complex III. It is also shown that BHAM has an influence on the relationship between the rate of ADP phosphorylation and the Q reduction level when alternative oxidase is active, and that KCN has an influence on the relationship between the alternative oxidase activity and the Q reduction level. These unexpected effects of BHAM and KCN observed at a given Q reduction level are likely due to functional connections between the two pathways activities or to protein–protein interaction.     

Measurements of in Vivo Ubiquinone Reduction Levels in Plant Cells

A method is described for the determination of in vivo ubiquinone (UQ) reduction levels in nongreen tissues by extraction and subsequent detection of ubiquinone-10 and ubiquinol-10 with high-performance liquid chromatography. In Petunia hybrida cell suspensions UQ reduction remained at a stable level of about 60%, despite the changing conditions during the batch culture (from excess sugar to starvation) and the concomitant variations in respiration. Also, in the presence of uncoupler, which causes a large increase in respiration via both the cytochrome pathway and the alternative pathway, UQ reduction levels stayed at 60%. In mitochondria isolated from these cells, activity of the alternative pathway was only observed at UQ reduction levels higher than 80%. It is proposed that in vivo the relationship between UQ reduction and the activity of the alternative oxidase is modulated by mechanisms such as thiol modifications and accumulation of organic acids. Accordingly, pyruvate concentration in P. hybrida cells increased in the presence of uncoupler.     

Beta-carotene redox reactions in photosystem II: electron transfer pathway

A carotenoid (Car), a chlorophyll (Chl(Z)), and cytochrome b(559) (Cyt b(559)) are able to donate electrons with a low quantum yield to the photooxidized chlorophyll, P680(+), when photosystem II (PSII) is illuminated at low temperatures. Three pathways for electron transfer from Cyt b(559) to P680(+) are considered: (a) the "linear pathway" in which Cyt b(559) donates via Chl(Z) to Car, (b) the "branched pathway" in which Cyt b(559) donates via Car and where Chl(Z) is also able to donate to Car, and (c) the "parallel pathway" where Cyt b(559) donates to P680 without intermediate electron carriers and electron donation from Chl(Z) and Car occurs by a competing pathway. Experiments were performed using EPR and spectrophotometry in an attempt to distinguish among these pathways, and the following observations were made. (1) Using PSII with an intact Mn cluster in which Cyt b(559) was preoxidized, Car oxidation was dominant upon illumination at < or =20 K, while electron donation from Chl dominated at >120 K. (2) When Cyt b(559) was prereduced, its light-induced oxidation occurred at < or =20 K in what appeared to be all of the centers and without the formation of a detectable Car(+) intermediate. The small and variable quantity of Car(+) photoinduced in these experiments can be attributed to the residual centers in which Cyt b(559) remained oxidized prior to illumination. (3) The relative rates for irreversible electron donation from Cyt b(559) and Car were determined indirectly at 20 K by monitoring the flash-induced loss of charge separation (i.e., the accumulation of Cyt b(559)(+)Q(A)(-) or Car(+)Q(A)(-)). Similar yields per flash were observed (13% for Cyt b(559) and 8% for Car), indicating similar donation rates. The slightly lower yield with Car as a donor is attributed at least in part to slow charge recombination occurring from the Car(+)Q(A)(-) radical pair in a fraction of centers. (4) Light-induced oxidation of Cyt b(559) and Car at 20 K was monitored directly by EPR, and the rates were found to be indistinguishable. The parallel pathway predicts that when both Cyt b(559) and Car are prereduced, the relative amounts of Cyt b(559)(+) and Car(+) produced upon illumination at 20 K should depend directly on their relative electron donation rates. The measured similarity in the donation rates thus predicts comparable yields of oxidation for both donors. However, what is observed experimentally is that Cyt b(559) oxidation occurs almost exclusively, and this argues strongly against the parallel pathway. The lack of Car(+) as a detectable intermediate is attributed to rapid electron transfer from Cyt b(559) to Car(+). The trapping of Car(+) at low temperature when Cyt b(559) is preoxidized but its absence when Cyt b(559) is prereduced is taken as an argument against the simple linear pathway. Overall, the data reported here and previously favor the branched pathway over the linear pathway, while the parallel pathway is thought to be unlikely. Structural considerations provide further arguments in favor of the branched model.     

Turnover of ubiquinone-0 at the acceptor side of photosynthetic reaction center

The steady-state operation of photosynthetic reaction center from Rhodobacter sphaeroides was investigated by measuring the rate of cytochrome photo-oxidation under intensive continuous illumination (808 nm, 5 W cm(-2)). The native quinone UQ(10) in Q(B) binding site of the reaction center was substituted by tailless UQ(0) and the binding parameters and the turnover rate of the UQ(0) was studied to test the recently discovered light-intensity dependent acceptor side effect (Gerencsér and Maróti 2006). The binding parameters of UQ(0) (k (on) = 2.1 x 10(5) M(-1) s(-1) and k (off) = 100 s(-1)) were characteristic to the RC exposed to high light-intensity. The dissociation constant (K (D) = 480 muM) determined under high light intensity is 2-3 times larger than that determined from flash-experiments. The light-intensity dependent acceleration of cytochrome turnover measured on reaction center of inhibited proton binding was independent of the type of the quinone and was sensitive only to the size ("pressure") of the quinone pool. The dissociation constants of different types of semiquinones show similarly high (several orders of magnitude) increase in the modified conformation of the Q(B) binding pocket due to high intensity of illumination. This result indicates the exclusive role of the quinone headgroup in the binding of semiquinone to different conformations of the protein.     

Dibromothymoquinone inhibition of the cyanide-sensitive and -insensitive respiration in MI-1 mutant of neurospora.

Effects of dibromothymoquinone (DBMIB) on the cyanide-sensitive and -insensitive pathways of respiration in mitochondria isolated from wild type and mi-1 mutant of Neurospora crassa have been investigated. It is shown that DBMIB inhibits the overall respiration in both strains in a similar manner. However, separate measurements of the DBMIB -induced inhibition of the KCN- and salicylhydroxamic acid (SHAM)-sensitive oxidation pathways in mi-1 pointed to some differences in the pattern and the degree of inhibition of these particular pathways, as reflected by a difference in the DBMIB concentration required for half-maximal inhibition and by the finding that the KCN-sensitive pathway is resistant to low concentrations of DBMIB. These results are consistent with a regulatory function of ubiquinone (UQ) in the cyanide-insensitive pathway in addition to its known carrier function in the cyanide -sensitive pathway of oxidation.     

Respiration in postharvest sugarbeet roots is not limited by respiratory capacity or adenylates

Control of respiration has largely been studied with growing and/or photosynthetic tissues or organs, but has rarely been examined in harvested and stored plant products. As nongrowing, heterotrophic organs that are reliant on respiration to provide all of their metabolic needs, harvested plant products differ dramatically in their metabolism and respiratory needs from growing and photosynthetically active plant organs, and it cannot be assumed that the same mechanism controls respiration in both actively growing and harvested plant organs. To elucidate mechanisms of respiratory control for a harvested and stored plant product, sugarbeet (Beta vulgaris L.) root respiration was characterized with respect to respiratory capacity, adenylate levels and cellular energy status in roots whose respiration was altered by wounding or cold treatment (1 degrees C) and in response to potential effectors of respiration. Respiration rate was induced by wounding in roots stored at 10 degrees C and by cold temperature in roots stored at 1 degrees C for 11-13d. Alterations in respiration rate due to wounding or storage temperature were unrelated to changes in total respiratory capacity, the capacities of the cytochrome c oxidase (COX) or alternative oxidase (AOX) pathways, adenylate concentrations or cellular energy status, measured by the ATP:ADP ratio. In root tissue, respiration was induced by exogenous NADH indicating that respiratory capacity was capable of oxidizing additional electrons fed into the electron transport chain via an external NADH dehydrogenase. Respiration was not induced by addition of ADP or a respiratory uncoupler. These results suggest that respiration rate in stored sugarbeet roots is not limited by respiratory capacity, ADP availability or cellular energy status. Since respiration in plants can be regulated by substrate availability, respiratory capacity or energy status, it is likely that a substrate, other than ADP, limits respiration in stored sugarbeet roots.     

Seasonal variation of leaf respiration and the alternative pathway in field-grown Populus × canadensis

The temperature response of plant respiration varies between species and can acclimate to changing temperatures. Mitochondrial respiration in plants has two terminal oxidases: the cytochrome c oxidase (COX) and the cyanide-resistant alternative oxidase (AOX). In Populus × canadensis var. italica, a deciduous tree species, we investigated the temperature response of leaf respiration via the alternative and cytochrome pathways, as well as seasonal changes in these pathways, using the oxygen isotope fractionation technique. The electron partitioning through the alternative pathway (τ(a) ) increased from 0 to 30-40% with measurement temperatures from 6 to 30°C at all times measured throughout the growing season. τ(a) at the growth temperature (the average temperature during 3 days prior to sampling) increased from 12 to 29% from spring until late summer and decreased thereafter. Total respiration declined throughout the growing season by 50%, concomitantly with decreases in both AOX (64%) and COX (32%) protein abundances. Our results provide new insight into the natural variability of AOX protein abundances and alternative respiration electron partitioning over immediate and seasonal timescales.     

Tight ATP and ADP binding in the noncatalytic sites of Escherichia coli F1-ATPase is not affected by mutation of bulky residues in the 'glycine-rich loop'

It is shown that ATP dissociates very slowly (koff less than 6.4 x 10(5) s-1, t1/2 greater than 3 h) from the three noncatalytic sites of E. coli F1-ATPase and that ADP dissociates from these three sites in a homogeneous fashion with koff = 1.5 x 10(-4) s-1 (t1/2 = 1.35 h). Mutagenesis of alpha-subunit residues R171 and Q172 in the 'glycine-rich loop' (Homology A) consensus region of the noncatalytic sites was carried out to test the hypothesis that unusually bulky residues at these positions are responsible wholly or partly for the observed tight binding of adenine nucleotides. The mutations alpha Q172G or alpha R171S,Q172G had no effects on ATP or ADP binding to or rates of dissociation from F1 noncatalytic sites. KdATP and KdADP of isolated alpha-subunit were weakened by approximately 1 order of magnitude in both mutants. The results suggest that neither residue alpha R171 nor alpha Q172 interacts directly with bound nucleotide, and show that the presence of bulky residues per se in the glycine-rich loop region of F1-alpha-subunit is not responsible for tight binding in the noncatalytic sites.     

Location and mobility of ubiquinones of different chain lengths in artificial membrane vesicles

Ubiquinone (UQn with n = 2, 3, or 10 isoprenoid groups) was incorporated into small, sonicated vesicles made of dipalmitoylphosphatidylcholine (DPPC) or dimyristoylphosphatidylcholine (DMPC). (1) The accessibility of oxidized UQ in DPPC or DMPC vesicles to the reductant sodium borohydride (NaBH4), measured by UV spectroscopy, was UQ2 greater than UQ3 greater than UQ10 (DPPC) and UQ2 greater than UQ3 approximately UQ10 (DMPC). (2) Catalysis of the reduction of entrapped ferricyanide by exogenous NaBH4 was more effective with UQ2 than UQ10 but was slower with all quinones than reduction by added dithionite. (3) The methoxy protons of UQ2 and UQ3 in DPPC and DMPC vesicles exhibited a single NMR resonance centered at approximately 3.95 ppm, whereas the methoxy groups of UQ10 gave rise to two separate proton resonances, at 3.93 ppm and a more narrow resonance at 3.78 ppm. The UQ10 population characterized by the 3.78 ppm resonance was present at a higher concentration in DPPC than in DMPC vesicles and was relatively insensitive to reduction by NaBH4. (4) UQ10 perturbed the melting temperature (Tm) of DPPC vesicles to a smaller extent (delta Tm = -1 degrees C) than did UQ2 and UQ3 (delta Tm = -3 to -4 degrees C). The combined UV and NMR data imply the following: The UQ10 pool characterized by the 3.78 ppm peak corresponds to a more mobile UQ10 fraction that is not reduced by NaBH4 in 2-3 min and is thought to be localized close to the center of the DPPC bilayer since it has little effect on the DPPC Tm.(ABSTRACT TRUNCATED AT 250 WORDS)     

Use of phosphocreatine kinetics to determine the influence of creatine on muscle mitochondrial respiration: an in vivo 31P-MRS study of oral creatine ingestion.

Recent human isolated muscle fiber studies suggest that phosphocreatine (PCr) and creatine (Cr) concentrations play a role in the regulation of mitochondrial respiration rate. To determine whether similar regulatory mechanisms are present in vivo, this study examined the relationship between skeletal muscle mitochondrial respiration rate and end-exercise PCr, Cr, PCr-to-Cr ratio (PCr/Cr), ADP, and pH by using (31)P-magnetic resonance spectroscopy in 16 men and women (36.9 +/- 4.6 yr). The initial PCr resynthesis rate and time constant (T(c)) were used as indicators of mitochondrial respiration after brief (10-12 s) and exhaustive (1-4 min) dynamic knee extension exercise performed in placebo and creatine-supplemented conditions. The results show that the initial PCr resynthesis rate has a strong relationship with end-exercise PCr, Cr, and PCr/Cr (r > 0.80, P < 0.001), a moderate relationship with end-exercise ADP (r = 0.77, P < 0.001), and no relationship with end-exercise pH (r = -0.14, P = 0.34). The PCr T(c) was not as strongly related to PCr, Cr, PCr/Cr, and ADP (r < 0.77, P < 0.001-0.18) and was significantly influenced by end-exercise pH (r = -0.43, P < 0.01). These findings suggest that end-exercise PCr and Cr should be taken into consideration when PCr recovery kinetics is used as an indicator of mitochondrial respiration and that the initial PCr resynthesis rate is a more reliable indicator of mitochondrial respiration compared with the PCr T(c).     

Oxidative Activity of Mitochondria Isolated from Plant Tissues Sensitive and Resistant to Chilling Injury

Arrhenius plots of the respiration rates of mitochondria isolated from chilling sensitive plant tissues (tomato and cucumber fruit, and sweet potato roots) showed a linear decrease from 25 C to about 9 to 12 C (with Q(10) values of 1.3 to 1.6), at which point there was a marked deviation with an increased slope as temperatures were reduced to 1.5 C (Q(10) of 2.2 to 6.3). The log of the respiration rate of mitochondria from chilling resistant tissues (cauliflower buds, potato tubers, and beet roots) showed a linear decrease over the entire temperature range from 25 to 1.5 C with Q(10) values of 1.7 to 1.8. Phosphorylative efficiency of mitochondria from all the tissues, as measured by ADP:O and respiratory control ratios, was not influenced by temperatures from 25 to 1.5 C. These results indicate that an immediate response of sensitive plant tissues to temperatures in the chilling range (0 to 10 C) is to depress mitochondrial respiration to an extent greater than that predicted from Q(10) values measured above 10 C. The results are also consistent with the hypothesis that a phase change occurs in the mitochondrial membrane as the result of a physical effect of temperature on some membrane component such as membrane lipids.     

川西亚高山针叶林土壤呼吸速率与不同土层温度的关系

采用密闭气室红外CO2分析法（IRGA）,在野外连续定位测定了川西亚高山冷杉原始林的土壤呼吸,并对其不同土层（0、5、10、15和20cm）的温度进行了同步测定．在此基础上,分析了土壤呼吸的日、季节动态变化,及其与不同土层温度的关系和土壤呼吸Q10值变化．结果表明：冷杉原始林土壤呼吸呈现明显的日变化和季节变化．土壤呼吸的日最高值出现在12：00-14：00,最低值出现在8：00—10：00,与土壤表面温度的日变化一致;土壤呼吸的季节变化表明：7—8月的土壤呼吸高于9-11月,与不同土层温度季节变化规律一致;土壤呼吸与不同土层温度呈显著的指数增长关系,土壤呼吸速率与土壤15cm深处温度的相关性明显高于其它土层;利用Q10模型计算0～20cm各土层的Q10值分别为2．36、4．75、4．90、6．27和5．46,表明海拔高、温度低的环境条件下,土壤呼吸的Q10值偏高．     

High efficiency versus maximal performance — The cause of oxidative stress in eukaryotes: A hypothesis

Degenerative diseases are in part based on elevated production of ROS (reactive oxygen species) in mitochondria, mainly during stress and excessive work under stress (strenuous exercise). The production of ROS increases with increasing mitochondrial membrane potential (ΔΨ_m). A mechanism is described which is suggested to keep ΔΨ_m at low values under normal conditions thus preventing ROS formation, but is switched off under stress and excessive work to maximize the rate of ATP synthesis, accompanied by decreased efficiency. Low ΔΨ_m and low ROS production are suggested to occur by inhibition of respiration at high [ATP]/[ADP] ratios. The nucleotides interact with phosphorylated cytochrome c oxidase (COX), representing the step with the highest flux-control coefficient of mitochondrial respiration. At stress and excessive work neural signals are suggested to dephosphorylate the enzyme and abolish the control of COX activity (respiration) by the [ATP]/[ADP] ratio with consequent increase of ΔΨ_m and ROS production. The control of COX by the [ATP]/[ADP] ratio, in addition, is proposed to increase the efficiency of ATP production via a third proton pumping pathway, identified in eukaryotic but not in prokaryotic COX. We conclude that ‘oxidative stress’ occurs when the control of COX activity by the [ATP]/[ADP] ratio is switched off via neural signals.     

西双版纳热带季节雨林与橡胶林土壤呼吸

季节雨林和橡胶(Hevea brasiliensis)林是西双版纳热带森林生态系统中原始林和大面积种植人工林的两种代表类型。热带季节雨林层次结构复杂,多样性丰富,而橡胶林结构简单,乔木层只有橡胶树1种。应用碱吸收法,研究了这两种植被类型土壤呼吸速率、地下5 cm土壤温度、气温和土壤含水率的季节变化规律,以及土壤呼吸速率与地下5 cm土壤温度、气温和土壤含水率的关系。结果表明:1)季节雨林和橡胶林土壤呼吸速率、土壤温度、气温和土壤含水率都有明显的季节变化,而且两种林型的变化趋势基本一致;2)季节雨林和橡胶林土壤呼吸速率与地下5 cm土壤温度和气温之间具有显著的指数相关关系,显著水平达1%,与地下5 cm温度的相关性(r²分别为0.87和0.82)明显高于与气温的相关性(r²分别是0.80和0.72);3)季节雨林和橡胶林土壤呼吸速率与土壤含水率具有显著的线性相关(r²分别是0.73和0.63),显著水平达1%;4)橡胶林的土壤呼吸速率明显高于季节雨林,这与两种林型的结构有关;5)季节雨林和橡胶林土壤呼吸的Q₁₀分别为2.16和2.18,比文献报道的热带土壤的Q₁₀(1.96)稍高。     

Mitochondrial energy metabolism is regulated via nuclear-coded subunits of cytochrome c oxidase

A new mechanism on regulation of mitochondrial energy metabolism is proposed on the basis of reversible control of respiration by the intramitochondrial ATP/ADP ratio and slip of proton pumping (decreased H+/e- stoichiometry) in cytochrome c oxidase (COX) at high proton motive force delta p. cAMP-dependent phosphorylation of COX switches on and Ca2+-dependent dephosphorylation switches off the allosteric ATP-inhibition of COX (nucleotides bind to subunit IV). Control of respiration via phosphorylated COX by the ATP/ADP ratio keeps delta p (mainly delta psi(m)) low. Hormone induced Ca2+-dependent dephosphorylation results in loss of ATP-inhibition, increase of respiration and delta p with consequent slip in proton pumping. Slip in COX increases the free energy of reaction, resulting in increased rates of respiration, thermogenesis and ATP-synthesis. Increased delta psi(m) stimulates production of reactive oxygen species (ROS), mutations of mitochondrial DNA and accelerates aging. Slip of proton pumping without dephosphorylation and increase of delta p is found permanently in the liver-type isozyme of COX (subunit VIaL) and at high intramitochondrial ATP/ADP ratios in the heart-type isozyme (subunit VIaH). High substrate pressure (sigmoidal v/s kinetics), palmitate and 3,5-diiodothyronine (binding to subunit Va) increase also delta p, ROS production and slip but without dephosphorylation of COX.     

Analysis of Respiratory Chain Regulation in Roots of Soybean Seedlings

Changes in the respiratory rate and the contribution of the cytochrome (Cyt) c oxidase and alternative oxidase (COX and AOX, respectively) were investigated in soybean (Glycine max L. cv Stevens) root seedlings using the ¹⁸O-discrimination method. In 4-d-old roots respiration proceeded almost entirely via COX, but by d 17 more than 50% of the flux occurred via AOX. During this period the capacity of COX, the theoretical yield of ATP synthesis, and the root relative growth rate all decreased substantially. In extracts from whole roots of different ages, the ubiquinone pool was maintained at 50% to 60% reduction, whereas pyruvate content fluctuated without a consistent trend. In whole-root immunoblots, AOX protein was largely in the reduced, active form at 7 and 17 d but was partially oxidized at 4 d. In isolated mitochondria, Cyt pathway and succinate dehydrogenase capacities and COX I protein abundance decreased with root age, whereas both AOX capacity and protein abundance remained unchanged. The amount of mitochondrial protein on a dry-mass basis did not vary significantly with root age. It is concluded that decreases in whole-root respiration during growth of soybean seedlings can be largely explained by decreases in maximal rates of electron transport via COX. Flux via AOX is increased so that the ubiquinone pool is maintained in a moderately reduced state.     

Muscle ubiquinone in healthy physically active males

Thirty-five (35) healthy physically active males had muscle biopsies taken from their vastus lateralis muscle to analyze for ubiquinone (vitamin Q, UQ), oxidative (muscle fiber types expressed as %ST and citrate synthase activity, CS) and fermentative (lactate dehydrogenase, LD) profiles. Graded cycle ergometer exercise to determine the intensities corresponding to onset of blood lactate accumulation set to 4.0 mmol × 1^–1 (W_OBLA) and symptom limited exercise (maximal, W_SL) were also undertaken. Eleven (11) subjects had also recently participated in a marathon race. UQ was positively related to CS (r = 0.67, p < 0.001) and %ST (r = 0.60, p < 0.001) but not to LD. UQ was also positively related to exercise capacity and/or marathon performance (e.g. W_OBLA × kg^–1 BW, r = 0.70, p < 0.001). It was suggested that muscle UQ allocation in man was related to variables describing molecular oxygen availability, respiratory activity and oxidative energy releasing processes but not to fermentation activity. UQ allocation to ST fibers/CS activity was suggested to be due to the double role of UQ: 1) as a mitochondrial coenzyme (CoQ₁₀) and 2) as a nonspecific antioxidant.