**6. Mitochondrial inner membrane carriers**

metabolism effects. Aldehydes derivation during lipid peroxidation, VDAC closure is probably a common feature leading to liver pathologies as was pointed out on almost indistinguishable histopathological manifestations in alcoholic liver disease, nonalcoholic fatty liver disease, nonalcoholic steatohepatitis, and toxicant-associated steatohepatitis. Ethanol is metabolized predominantly by the liver in two-step oxidation: first to acetaldehyde (AcAld) by catalytic action of alcohol dehydrogenase (ALD) followed by oxidation to acetate by aldehyde dehydrogenase (ALDH). The first step undergoes in cytosol and peroxisomes by effect of cytochrome P4502E1 and catalase. Although the oxidation is prevalent over ALD, the consequence of cytochrome P450 metabolism is overwhelmingly negative due to the formation of ROS, e.g., hydroxyethyl radicals. Of the 19 known mammalian ALDH genes, mitochondrial ALDH2 with high affinity for AcAld (Km < 1 μM) is the most important for AcAld oxidation (and detoxification) to acetate [73]. In both reactions, by oxidation of 1 mole of ethanol, 2 moles of NADH are formed, further requiring oxidation in the respiratory chain. As little as 2.5 h after a single bolus dose of ethanol ingestion a swift increase of alcohol metabolism (SIAM), an adaptive increase of hepatic ethanol metabolism occurs [74]. Mitochondrial respiration causing NADH oxidation nearly doubles, but it does not lead to increased ATP generation. To the contrary, hepatic ATP decreases, glycolysis is stimulated, and glycogen stores are depleted. Furthermore, mitochondrial β-oxidation of fatty acids becomes inhibited, promoting fat accumulation within hepatocytes (steatosis) [75]. This is probably the result of decrease in MOM permeability most likely by VDAC closure, promoting selective oxidation of AcAld, since VDAC closure blocks mitochondrial ATP release, respiratory substrates uptake, and uptake of fatty acids for β-oxidation [67]. Adrenergic hormones release free fatty acids from adipose tissue, which serve as substrates for

long-chain fatty acid peroxisomal β-oxidation. The ensuing peroxisomal H<sup>2</sup>

sented in diet capable to prevent steatosis development.

redox translocator in MIM have been identified [80, 81].

**5.1. Translocase of the MOM**

258 Mitochondrial Diseases

can promote catalase-dependent alcohol metabolism [67]. AcAld is toxic to mitochondria and aggravates oxidative stress by binding to GSH and promoting GSH leakage [76]. Moreover, as mentioned before, ethanol metabolism and also NADH overproduction cause formation of ROS, lipid peroxidation, onset of the mitochondrial permeability transition, and apoptosis [77]. However, as has been showed, short- and medium-chain fatty acids can cross mitochondrial membrane freely using carnitine shuttle or other transport system [78] and therefore are pre-

The endosymbiotic relationship of α-proteobacteria and archaic eukaryotic cell results in massive loss and transfer of coding sequences from mtDNA to the nucleus and only less than 1% is retained in today's mtDNA. Thus, most mitochondrial proteins (1000–1500) undergo cytosol translation and are subsequently transferred to mitochondria, requiring membrane complexes of protein translocators, translocases, or translocons. They include TOM and TIM for large conductance channels with almost identical properties [79]. In addition, other mitochondria protein translocators like TOB/SAM complex in MOM and Mia40/Tim401-Erv1

The general entry gate for mitochondrial proteins is thought to be TOM40 complex in MOM consisting of core sequence Tom40, Tom22, Tom7, Tom6, Tom5, peripheral associated receptors Tom20, Tom70, and a minor component Tom71. Among them, only Tom40, Tom22, and

O2

formation then

The inner mitochondrial membrane is relatively low permeable to ions in order to minimize energy dissipation formed on complexes through generation of electrochemical proton gradient, in its direct link with ADP phosphorylation. Random flow of charged metabolites via MIM would lead to a reduction in the membrane potential and ATP formation [1]. The relative impermeability of the MIM is the basis of chemiosmotic hypothesis proposed by Mitchell. As discussed in O'Rourke [90], Mitchell recognized three modes of ion transport. Symporters cotransport multiple ions (or an ion and a metabolite) in the same direction across the membrane often utilizing the asymmetric electrochemical ion gradient to drive the transport in a thermodynamically favorable direction, as for example mitochondrial Pi /H+ carrier. Antiporters exchange ions on different sides of the membrane. Antiporters can be electroneutral (the Na+ /H+ antiporter of the mitochondrial or plasma membrane) or electrogenic. For electrogenic transporters, ion flux is driven by both the electrochemical gradients of the transported ions and the membrane potential. For uniporters, the transport rates are in the range of 104 –106 ions s−1, based on ions flowing down their electrochemical gradient.

The mutation in gene-encoded DNC (chromosomal localization 17q25.1) is known to be associated with Amish microcephaly. Amish microcephaly has only been observed in Old Order Amish community in Pennsylvania, U.S.A, with a high prevalence of about 1:500. The disease is characterized by severe congenital microcephaly, elevated levels of α-ketoglutarate in urine, and premature death. The only non-CNS physical anomaly is moderate micrognathia. Patients manifest no orientation to sight or sound and no fine or gross motor development and have metabolic acidosis enhanced by episodic viral illnesses, and in some cases patients have mild hepatomegaly and difficulty maintaining normal body temperature and develop increasing irritability [97]. Study on SLC25A19 knock-out mice has shown that metabolic

Nuclear Encoded Mitochondrial Proteins in Metabolite Transport and Oxidation Pathway…

http://dx.doi.org/10.5772/intechopen.72937

261

Four ANT isoforms are encoded in human genome on the chromosome X. ANT1–3 are structurally similar and proteins are about 90% identical, and ANT4 only shares 66–68% consistency in the amino acid composition with other isoforms. Isoforms are specifically expressed in different types of cells and tissues. ANT1 (SLC25A4) is expressed in the skeletal muscle, brain, and heart. ANT2 (SLC25A5) is expressed in the liver and proliferating cells and is overexpressed in various types of cancer cell lines. ANT3 (SLC25A6) is ubiquitous in all tissues, and ANT4 (SLC25A31) is specific to the testis and germ cells [105]. The translocase is highly selective of the adenine nucleotide and provides a continuous shift of ADP to the mitochondria required to maintain oxidative phosphorylation and membrane potential. ANT is also

Impaired translocase activity affects the energy metabolism of the cell by decreasing mitochondrial ATP synthesis and increasing mitochondrial membrane potential [108], thus contributing to the promotion of apoptosis. The rate-limiting factor of apoptosis is mtPTP formation, which is actually increased permeabilization of the mitochondrial membrane for all the solvents up to 1.5 kDa. It is a nonspecific pore, where ANT, VDAC, cyclophilin D, hexokinase, creatine kinase, and peripheral benzodiazepine receptor are effective but not as direct components or core structures. Moreover, there is an evidence for apoptosis regulators of the Bcl-2 family, Bak and Bax, requirement for mtPTP-dependent MOM permeabilization [109]. PTP opening is linked to mitochondrial dysfunction because its occurrence leads to the set of consequences that will arise, as mitochondrial depolarization, cessation of ATP synthesis, Ca2+ release, pyridine nucleotide depletion, inhibition of respiration and matrix swelling, MOM rupture, and release of pro-apoptotic proteins such as cytochrome c, endonuclease G, and AIF [110, 111]. Detrimental effects are seen for long-lasting mtPTP opening, while shortterm effects are involved in physiological regulation of Ca2+ and ROS homeostasis [112, 113]. Cancer cells are able to survive suppression of mitochondrial oxidative phosphorylation under hypoxic conditions through higher rate of glycolysis; however, it depends on ATP uptake especially for mitochondrial potential generation and Ca2+ exchange [114]. The expression of ANT isoforms is related to the adaptation of metabolic properties of cancer cells. ANT2 is overexpressed in various types of human cancer cells and in several hormone-dependent cancers [115, 116]. It was found that ANT2 proves properties allowing the import of ATP into mitochondria (in coexpression with hexokinase II and a subunit of mitochondrial F0F1-ATPase, ATPsynβ),

abnormalities in humans are due to absent TPC activity [104].

implicated in leakage of protons and inducible proton leakage [106, 107].

*6.1.2. Deoxynucleotide carrier*

Mitchell and Moyle [91] reported that anions, including Pi , succinate, and malonate, accelerated the rate of decay of the pH gradient induced by a pulse of oxygen. This suggested the presence of anion transport systems coupled to proton movement, leading to the identification of the anion/metabolite-coupled cotransporter family. Inner membrane anion uniporters have been less well studied, but in the 1980s, an inner membrane anion channel was postulated to account for anion-selective mitochondrial swelling responses [92]. Moreover, some mitochondrial membrane proteins (e.g., mitochondrial uncoupling protein) were identified to display anion channel activity [90]. Based on the research, seven metabolite-specific mitochondrial transporters or carriers were proposed. Studies of amino acid sequence composition showed that the carriers form a well-defined family (in humans known as the solute carrier 25 family (SLC25)), with the one defining feature, a tripartite structure of three homologous sequence repeats of about 100 amino acid residues each, which was first noted in the published sequence of the bovine ADP/ATP carrier [93]. A signature motif containing P-X-[D/E]-X-X-[R/K] sequence is conserved in all members and in all three sequence repeats [94]. According to typical sequence repeats and signature motif, eukaryotic mitochondria were found to contain 35–55 different carriers when compared to genomic DNA database [95]. The human genome encodes 48 members of the SLC25 family, of which 30 are identified [96]. The isoforms of carrier members are encoded by different genes, and only the phosphate carrier has two alternatively spliced isoforms [97].
