Intermembrane transport

The movement of lipids within the cell can be divided into two different general classes of transport intramembrane transport, which entails the transbilayer movement of the lipid molecule and intermembrane transport, which is the movement of lipid molecules from one distinct membrane to another. In some cases, the transmembrane movement of a phospholipid is coupled to a process that also removes the lipid from the membrane in which it was resident, and these events have the character of the lipid being vectorially pumped across and out of the membrane. Extensive reviews of these processes have been published [6,7,9-12]. 

Fig. 4. Genetic and biochemical elements of transbilayer and intermembrane transport of Ra-LPS. Ra-LPS is synthesized on the cytoplasmic face of the inner membrane of gram-negative bacteria. MsbA transports the Ra-LPS to the outer aspect of the inner membrane in an ATP-dependent reaction. The Ra-LPS is transferred to LptA, a subunit of a multipartite ATPase. The LptX is proposed to physically couple ATP hydrolysis by LptB to the transport function of LptA. LptA transfers the LPS to RlpB, which subsequently transfers the molecule to Imp. The Imp completes the transfer of the LPS to the outer aspect of the outer membrane.

From a theoretical perspective a number of processes could contribute to the intermembrane transport of lipids. These include (i) monomer solubility and diffusion, (ii) soluble carriers such as lipid transfer proteins, (iii) carrier vesicles, (iv) membrane apposition and lipid transfer, and (v) membrane fusion processes. Lipids such as fatty acids, lysophos-phatidic acid, and CDP-DG might have sufficient water solubility to allow for some monomeric transport, but most other lipids are likely to require one of the other potential mechanisms due to their extremely low solubility. 

Cytochrome c, like UQ is a mobile electron carrier. It associates loosely with the inner mitochondrial membrane (in the intermembrane space on the cytosolic side of the inner membrane) to acquire electrons from the Fe-S-cyt C aggregate of Complex 111, and then it migrates along the membrane surface in the reduced state, carrying electrons to cytochrome c oxidase, the fourth complex of the electron transport chain. 

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. 

In resting muscle the high concentration of ADP does not decrease the proton gradient effectively and the high membrane potential slows electron transport. ADP, formed when ATP is hydrolyzed by myosin ATPase during contraction, may stimulate electron transport. However, the concentration of ATP (largely as its Mg salt) is buffered by its readily reversible formation from creatine phosphate catalyzed in the intermembrane space, and in other cell compartments, by the various isoenzymes of creatine kinase (reviewed by Walliman et al., 1992). 

Figure 12.2 Copper chaperone function, (a) Copper homeostasis in Enterococcus hirae is affected by the proteins encoded by the cop operon. CopA, Cu1+-import ATPase CopB, Cu1+-export ATPase CopY, Cu1+-responsive repressor copZ, chaperone for Cu1+ delivery to CopY. (b) The CTR family of proteins transports copper into yeast cells. Atxlp delivers copper to the CPx-type ATPases located in the post Golgi apparatus for the maturation of Fet3p. (c) Coxl7p delivers copper to the mitochondrial intermembrane space for incorporation into cytochrome c oxidase (CCO). (d) hCTR, a human homologue of CTR, mediates copper-ion uptake into human cells. CCS delivers copper to cytoplasmic Cu/Zn superoxide dismutase (SOD1). Abbreviations IMM, inner mitochondrial membrane OMM, outer mitochondrial membrane PM, plasma membrane PGV, post Golgi vessel. Reprinted from Harrison et al., 2000. Copyright (2000), with permission from Elsevier Science.

FIGURE 31-7 Mitochondrial carriers. Ions and small molecules enter the intermembrane space, since the outer mitochondrial membrane is not a significant permeability barrier. However, the inner mitochondrial membrane is impermeable to ions except those for which there are specific carriers. Most of the carriers are reversible, as indicated by two-headed arrows. Compounds transported in one direction are indicated in red. The ATP/ADP translocase and the aspartate-glutamate carrier are both electrophoretic their transport is driven in the direction of the mitochondrial membrane potential, as indicated by red arrows. Glutamine is carried into the matrix by an electroneutral carrier. The unimpaired functioning of mitochondrial carriers is essential for normal metabolism. (Adapted with permission from reference [70].)... 

To explain how H+ transfer occurred across the membrane Mitchell suggested the protons were translocated by redox loops with different reducing equivalents in their two arms. The first loop would be associated with flavoprotein/non-heme iron interaction and the second, more controversially, with CoQ. Redox loops required an ordered arrangement of the components of the electron transport system across the inner mitochondrial membrane, which was substantiated from immunochemical studies with submitochondrial particles. Cytochrome c, for example, was located at the intermembranal face of the inner membrane and cytochrome oxidase was transmembranal. The alternative to redox loops, proton pumping, is now known to be a property of cytochrome oxidase. 

The proton-motive Q-cycle model, put forward by Mitchell (references 80 and 81) and by Trumpower and co-workers, is invoked in the following manner (1) One electron is transferred from ubiquinol (ubiquinol oxidized to ubisemi-quinone see Figure 7.27) to the Rieske [2Fe-2S] center at the Qo site, the site nearest the intermembrane space or p side (2) this electron can leave the bci complex via an attached cytochrome c or be transferred to cytochrome Ci (3) the reactive ubisemiquinone reduces the low-potential heme bL located closer to the membrane s intermembrane (p) side (4) reduced heme bL quickly transfers an electron to high-potential heme bn near the membrane s matrix side and (5) ubiquinone or ubisemiquinone oxidizes the reduced bn at the Qi site nearest the matrix or n side. Proton translocation results from the deprotonation of ubiquinol at the Qo site and protonation of ubisemiquinone at the Qi site. Ubiquinol generated at the Qi site is reoxidized at the Qo site (see Figure 7.27). Additional protons are transported across the membrane from the matrix (see Figure 7.26 illustrating a similar process for cytochrome b(6)f). The overall reaction can be written... 

What do I mean by a proton concentration gradient Simply, there is a higher concentration of protons in the space between the inner and outer membranes of the mitochondrion than in the mitochondrial interior. The gradient is formed from the energy released in the transfer of electrons down the electron transport chain. Put another way, the released energy is employed to pump protons across the inner mitochondrial membrane into the intermembrane space. 

The ATP, which is transported out of the mitochondrion, immediately phosphorylates creatine, catalysed by the creatine kinase in the intermembrane space. 

Transport of protons from the intermembrane space into the matrix across the inner membrane of the mitochondria occurs via the ATP synthase complex (Fo-Ei), which generates ATP. Somewhat surprisingly, it was discovered that proteins that transport protons back into the matrix, but without generating ATP, do, in fact, exist and are... 

Proton transport via complexes I, III, and IV takes place vectorially from the matrix into the intermembrane space. When electrons are being transported through the respiratory chain, the concentration in this space increases—i. e., the pH value there is reduced by about one pH unit. For each H2O molecule formed, around 10 H ions are pumped into the intermembrane space. If the inner membrane is intact, then generally only ATP synthase (see p. 142) can allow protons to flow back into the matrix. This is the basis for the coupling of electron transport to ATP synthesis, which is important for regulation purposes (see p. 144). 

Both mitochondrial membranes are very rich in proteins. Porins (see p. 214) in the outer membrane allow small molecules (< 10 kDa) to be exchanged between the cytoplasm and the intermembrane space. By contrast, the inner mitochondrial membrane is completely impermeable even to small molecules (with the exception of O2, CO2, and H2O). Numerous transporters in the inner membrane ensure the import and export of important metabolites (see p. 212). The inner membrane also transports respiratory chain complexes, ATP synthase, and other enzymes. The matrix is also rich in enzymes (see B). 

The inner membrane itself plays an important part in oxidative phosphorylation. As it is impermeable to protons, the respiratory chain—which pumps protons from the matrix into the intermembrane space via complexes 1, 111, and IV—establishes a proton gradient across the inner membrane, in which the chemical energy released during NADH oxidation is conserved (see p. 126). ATP synthase then uses the energy stored in the gradient to form ATP from ADP and inorganic phosphate. Several of the transport systems are also dependent on the H"" gradient. 

A. As electrons pass through complexes I, III, and IV (but not complex II), protons are transported across the inner mitochondrial membrane from the matrix to the intermembrane space, creating a pH gradient that represents a form of stored energy. 

Fig. 7.3.1 The heme synthesis pathway starts in the mitochondrion. The next four steps proceed in the cytosol. Coproporphyrinogen oxidase is in the intermembrane space of the mitochondrion, and the last two enzymes reside at the mitochondral matrix side of the inner membrane. The product heme represses the first and rate-limiting enzyme -aminolevulinic acid (5-ALA) synthase at transcription, during the translation step, and by its transport into the mitochondrion... 

In addition, both ATM1 and Ervl, which are required for the maturation of extra-mitochondrial (cytosolic) proteins and that are part of the mt export machinery, have been identified in the C. parvum genome (reviewed in Barbera et al. 2006). The former is an IM protein of the ABC transporter family that faces the mt matrix (Decottignies and Goffeau 1997), and the second is an intermembrane sulhydryl oxidase whose function in cytosolic FeS protein biosynthesis is still unknown (reviewed in Barbera et al. 2006). When combined with glutathione, which may help stabilize proteins being exported from the mitochondrion into the cytosol (reviewed in Tachezy and Dolezal... 

The basic mechanism underlying the toxicity of salicylate is the uncoupling of oxidative phosphorylation. For oxidative phosphorylation to take place, there is a requirement of a charge difference between the intermembrane space and the matrix of the mitochondria (Fig. 7.60). This is achieved when electrons move down the chain of multienzyme complexes and electron carriers (the electron transport chain), causing protons to move from the mitochondrial matrix to the intermembrane space. Consequently, a pH difference builds up, which is converted into an electrical potential across the membrane of approximately 200 mV over 8 nm. 

FIGURE17-6 Fatty acid entry into mitochondria via the acyl-carnitine/ carnitine transporter. After fatty acyl-carnitine is formed at the outer membrane or in the intermembrane space, it moves into the matrix by facilitated diffusion through the transporter in the inner membrane. In the matrix, the acyl group istransferred to mitochondrial coenzyme... 

A, freeing carnitine to return to the intermembrane space through the same transporter. Acyltransferase I is inhibited by malonyl-CoA, the first intermediate in fatty acid synthesis (see Fig. 21-1). This inhibition prevents the simultaneous synthesis and degradation of fatty acids... 

In the third and final step of the carnitine shuttle, the fatty acyl group is enzymatically transferred from carnitine to intramitochondrial coenzyme A by carnitine acyltransferase II. This isozyme, located on the inner face of the inner mitochondrial membrane, regenerates fatty acyl-CoA and releases it, along with free carnitine, into the matrix (Fig. 17-6). Carnitine reenters the intermembrane space via the acyl-camitine/car-nitine transporter. 

Complex III Ubiquinone to Cytochrome c The next respiratory complex, Complex III, also called cytochrome focx complex or ubiquinone icytochrome c oxidoreductase, couples the transfer of electrons from ubiquinol (QH2) to cytochrome c with the vectorial transport of protons from the matrix to the intermembrane space. The determination of the complete structure of this huge complex (Fig. 19-11) and of Complex IV (below) by x-ray crystallography, achieved between 1995 and 1998, were landmarks in the study of mitochondrial electron transfer, providing the structural framework to integrate the many biochemical observations on the functions of the respiratory complexes. 

The adenine nucleotide translocase, integral to the inner membrane, binds ADP3 - in the intermembrane space and transports it into the matrix in exchange for an ATP4 molecule simultaneously transported outward (see Fig. 13-1 for the ionic forms of ATP and ADP). Because this antiporter moves four negative charges out for every three moved in, its activity is favored by the... 

Proton pump Electron transport is coupled to the phosphorylation of ADP by the transport of protons (H+) across the inner mitochon drial membrane from the matrix to the intermembrane space. This process creates across the inner mitochondrial membrane an electrical gradient (with more positive charges on the outside of the membrane than on the inside) and a pH gradient (the outside of the... 

The inner mitochondrial membrane is impermeable to most charged or hydrophilic substances. However, it contains numerous transport proteins that permit passage of specific molecules from the cytosol (or more correctly, the intermembrane space) to the mitochondrial matrix. 

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