The respiratory chain

We are interested now in the fate of the reduced coenzymes or, more precisely, of the hydrogen with which the coenzymes are charged. This hydrogen is introduced into a chain of redox systems. Electron transport chains of this kind are not new to us. We already know of them in connection with the primary processes of photosynthesis. Here again, just as in a few transport steps in that case, we must visualize 2 H as 2 H -h 2 e—. 

The electron transport chain into which the hydrogen of the reduced coenzymes of the citric acid cycle is channelled is called the respiratory chain. In this chain hydrogen or electrons are conducted downhill from redox systems of high to redox systems of low electron pressure. The energy set free in the process is utilized for the formation of ATP. At the end of the chain hydrogen is oxidized to water. 

What are the components of the respiratory chain At the outset it must be said that the respiratory chain in animals, bacteria, and many lower plants is different from that in higher plants. Some of the details are still unexplained. Nonetheless the following redox systems are components of the respiratory chain of higher plants  

3 Cytochromes b (Cytochrome b complex, typical of higher plants) 

2 Cytochromes c (C549 and C547. The naming of individual cytochromes of the a-, b-, c- groups is based on one of the characteristic absorption maxima that appear in the reduced state.) 

Some organic co-factors and metal centers in proteins act as electron transfer agents in a number of biological processes we need to be able to predict which species is reduced or oxidized in a redox reaction. 

We have seen that a cell reaction has 1C 1 if 0 and that 0 corresponds to reduction at the right-hand electrode. We have also seen that may be written as the difference of the standard potentials of the redox couples in the right and left electrodes (eqn 5.17, = r - El). A reaction corresponding to 

A couple with a low standard potential has a thermodynamic tendency to reduce a couple with a high standard potential. 

More briefly low reduces high and, equivalently, high oxidizes low. The same arguments apply to the biological standard values of the potentials. 

Consider the iron-containing protein ferredoxin, which participates in plant photosynthesis (Section 5.11), and cytochrome c, which participates in the last steps of respiration (Section 5.10). It follows from Table 5.2 that 

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ATP results from the movement of approximately three protons from the cytosol into the matrix through Fg. Altogether this means that approximately four protons are transported into the matrix per ATP synthesized. Thus, approximately one-fourth of the energy derived from the respiratory chain (electron transport and oxidative phosphorylation) is expended as the electrochemical energy devoted to mitochondrial ATP-ADP transport. 

Wefss, H., Friedrich, T, Hofliaus, G., and Preis, D., 1991. The respiratory-chain NADH dehydrogena.se (Complex I) of mitochondria. European Journal of Biochemistry 197 563—576. 

The thiazolidinediones have also been reported to act as inhibitors of the respiratory chain at high concentrations, and this appears to account for their ability to activate AMGPK in cultured cells. However, the primary target of the thiazolidinediones appears to be the peroxisome proliferator-activated receptor-y ( PPAR-y), a member of the nuclear receptor superfamily expressed in adipocytes. One of the major effects of stimulation of PPAR-y in adipocytes is the release ofthe... 

Atovaquone, a hydroxynaphthoquinone, selectively inhibits the respiratory chain of protozoan mitochondria at the cytochrome bcl complex (complex III) by mimicking the natural substrate, ubiquinone. Inhibition of cytochrome bcl disrupts the mitochondrial electron transfer chain and leads to a breakdown of the mitochondrial membrane potential. Atovaquone is effective against all parasite stages in humans, including the liver stages. 

Mitochondrial permeability transition involves the opening of a larger channel in the inner mitochondrial membrane leading to free radical generation, release of calcium into the cytosol and caspase activation. These alterations in mitochondrial permeability lead eventually to disruption of the respiratory chain and dqDletion of ATP. This in turn leads to release of soluble intramito-chondrial membrane proteins such as cytochrome C and apoptosis-inducing factor, which results in apoptosis. 

NAD+ and NADP+ are coenzymes of dehydrogenases. NADH and NADPH are intermediate carriers of both hydrogen and electrons. Most NAD-dependent enzymes are located in the mitochondria and deliver H2 to the respiratory chain whereas NADP-dependent enzymes take part in cytosolic syntheses (reductive biosyntheses). 

Mitochondria have their own DNA (mtDNA) and genetic continuity. This DNA only encodes 13 peptide subunits synthesized in the matrix that are components of complexes I, III, IV, and V of the respiratory chain. Most mitochondrial proteins are synthesized on cytoplasmic ribosomes and imported by specific mechanisms to their specific locations in the mitochondrion (see below). 

I These dehydrogenases are lim-j ited to the respiratory chain at 7 the level of complex III by ETF (5) dehydrogenase (6) and ubiqui-... 

The mechanism of ATP synthesis discussed here assumes that protons extruded during electron transport are in the bulk phase surrounding the inner mitochondrial membrane (intermembrane and extramitochondrial spaces). An alternative view is that there are local proton circuits within or close to the respiratory chain and complex V, and that these protons may not be in free equilibrium with the bulk phase (Williams, 1978), although this has not been supported experimentally (for references see Nicholls and Ferguson, 1992). The chemiosmotic mechanism is both elegant and simple and explains all the known facts about ATP synthesis and its dependence on the structural integrity of the mitochondria, although the details may appear complex. This mechanism will now be discussed in more detail. 

Another pathway is the L-glycerol 3-phosphate shuttle (Figure 11). Cytosolic dihydroxyacetone phosphate is reduced by NADFl to s.n-glycerol 3-phosphate, catalyzed by s,n-glycerol 3-phosphate dehydrogenase, and this is then oxidized by s,n-glycerol 3-phosphate ubiquinone oxidoreductase to dihydroxyacetone phosphate, which is a flavoprotein on the outer surface of the inner membrane. By this route electrons enter the respiratory chain.from cytosolic NADH at the level of complex III. Less well defined is the possibility that cytosolic NADH is oxidized by cytochrome bs reductase in the outer mitochondrial membrane and that electrons are transferred via cytochrome b5 in the endoplasmic reticulum to the respiratory chain at the level of cytochrome c (Fischer et al., 1985). 

Normally, these reactive species are destroyed by protective enzymes, such as superoxide dismutase in mitochondria and cytosol and catalase in peroxisomes, but if a tissue has been anoxic the respiratory chain is very reduced and reoxygenation allows dangerous amounts to be formed. Muscle also contains significant quantities of the dipeptide, camosine ((J-alanylhistidine) (10—25 mM). The functions of camosine are obscure although it has been suggested to be an effective antioxidant (Pavlov et al., 1993). 

Mitochondrial DNA is transcribed as a polycistronic RNA which is subsequently cleaved to generate the various mature mRNA, tRNA, and rRNA (Clayton, 1984). The 13 proteins encoded by mtDNA are all components of the respiratory chain and are seven subunits of complex I, one subunit of complex III, three subunits of complex IV, and two subunits of complex V. 

It is conventional to discuss the stoichiometry for proton extrusion as HV2e ratios, although there are two-, one-, and four-electron reductions at different stages in the respiratory chain. Most textbooks still assert that the flow of two electrons... 

Brand, M.D. Murphy, M.P. (1987). Control of electron flux through the respiratory chain in mitochondria and cells. Biol. Rev. 62, 141-193. 

Sumegi, B., Porpaczy, Z., Alkonyi, 1. (1991). Kinetic advantage of the interaction between the fatty acid P-oxidation enzymes and the complexes of the respiratory chain. Biochim. Biophys. Acta 1081, 121-128. 

Mitochondria are unique organelles in that they contain their own DNA (mtDNA), which, in addition to ribosomal RN A (rRNA) and transfer RN A (tRNA)-coding sequences, also encodes 13 polypeptides which are components of complexes I, III, IV, and V (Anderson et al., 1981). This fact has important implications for both the genetics and the etiology of the respiratory chain disorders. Since mtDNA is maternally-inherited, a defect of a respiratory complex due to a mtDNA deletion would be expected to show a pattern of maternal transmission. However the situation is complicated by the fact that the majority of the polypeptide subunits of complexes I, III, IV, and V, and all subunits of complex II, are encoded by nuclear DNA. A defect in a nuclear-coded subunit of one of the respiratory complexes would be expected to show classic Mendelian inheritance. A further complication exists in that it is now established that some respiratory chain disorders result from defects of communication between nuclear and mitochondrial genomes (Zeviani et al., 1989). Since many mitochondrial proteins are synthesized in the cytosol and require a sophisticated system of posttranslational processing for transport and assembly, it is apparent that a diversity of genetic errors is to be expected. 

This complex consists of four subunits, all of which are encoded on nuclear DNA, synthesized on cytosolic ribosomes, and transported into mitochondria. The succinate dehydrogenase (SDH) component of the complex oxidizes succinate to fumarate with transfer of electrons via its prosthetic group, FAD, to ubiquinone. It is unique in that it participates both in the respiratory chain and in the tricarboxylic acid (TC A) cycle. Defects of complex II are rare and only about 10 cases have been reported to date. Clinical syndromes include myopathy, but the major presenting features are often encephalopathy, with seizures and psychomotor retardation. Succinate oxidation is severely impaired (Figure 11). 

Reduced nicotinamide-adenine dinucleotide (NADH) plays a vital role in the reduction of oxygen in the respiratory chain [139]. The biological activity of NADH and oxidized nicotinamideadenine dinucleotide (NAD ) is based on the ability of the nicotinamide group to undergo reversible oxidation-reduction reactions, where a hydride equivalent transfers between a pyridine nucleus in the coenzymes and a substrate (Scheme 29a). The prototype of the reaction is formulated by a simple process where a hydride equivalent transfers from an allylic position to an unsaturated bond (Scheme 29b). No bonds form between the n bonds where electrons delocalize or where the frontier orbitals localize. The simplified formula can be compared with the ene reaction of propene (Scheme 29c), where a bond forms between the n bonds. 

As components in the respiratory chain of electron transport from substrate to oxygen (Figure 12-3). 

Generally, NAD-linked dehydrogenases catalyze ox-idoreduction reactions in the oxidative pathways of metabolism, particularly in glycolysis, in the citric acid cycle, and in the respiratory chain of mitochondria. NADP-linked dehydrogenases are found characteristically in reductive syntheses, as in the extramitochon-drial pathway of fatty acid synthesis and steroid synthesis—and also in the pentose phosphate pathway. 

The cytochromes are iron-containing hemoproteins in which the iron atom oscillates between Fe + and Fe + during oxidation and reduction. Except for cytochrome oxidase (previously described), they are classified as dehydrogenases. In the respiratory chain, they are involved as carriers of electrons from flavoproteins on the one hand to cytochrome oxidase on the other (Figure 12-4). Several identifiable cytochromes occur in the respiratory chain, ie, cytochromes b, Cp c, a, and (cytochrome oxidase). Cytochromes are also found in other locations, eg, the endoplasmic reticulum (cytochromes P450 and h, and in plant cells, bacteria, and yeasts. 

Mitochondria have an outer membrane that is permeable to most metabohtes, an inner membrane that is selectively permeable, and a matrix within (Figure 12-1). The outer membrane is characterized by the presence of various enzymes, including acyl-CoA synthetase and glycerolphosphate acyltransferase. Adenylyl kinase and creatine kinase are found in the intermembrane space. The phospholipid cardiolipin is concentrated in the inner membrane together with the enzymes of the respiratory chain. 

THE RESPIRATORY CHAIN COLLECTS OXIDIZES REDUCING EQUIVALENTS... 

Most of the energy liberated during the oxidation of carbohydrate, fatty acids, and amino acids is made available within mitochondria as reducing equivalents (—H or electrons) (Figure 12-2). Mitochondria contain the respiratory chain, which collects and transports reducing equivalents directing them to their final reaction with oxygen to form water, the machinery for... 

Components of the Respiratory Chain Are Arranged in Order of Increasing Redox Potential... 

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