Substrate shuttles

Oxidation of Extramitochondrial NADH Is Mediated by Substrate Shuttles... 

Under aerobic conditions, the hydrogen atoms of NtUDH are oxidised within the mitochondrion pyruvate is also oxidised in the mitochondrion (Figure 9.15). However, NADH cannot be transported across the inner mitochondrial membrane, and neither can the hydrogen atoms themselves. This problem is overcome by means of a substrate shuttle. In principle, this involves a reaction between NADH and an oxidised substrate to produce a reduced product in the cytosol, followed by transport of the reduced product into the mitochondrion, where it is oxidised to produce hydrogen atoms or electrons, for entry into the electron transfer chain. Finally, the oxidised compound is transported back into the cytosol. The principle of the shuttle is shown in Figure 9.16. 

The final reactions to be considered in the metabolism of ethanol in the liver are those involved in reoxidation of cytosolic NADH and in the reduction of NADP. The latter is achieved by the pentose phosphate pathway which has a high capacity in the liver (Chapter 6). The cytosolic NADH is reoxidised mainly by the mitochondrial electron transfer system, which means that substrate shuttles must be used to transport the hydrogen atoms into the mitochondria. The malate/aspartate is the main shuttle involved. Under some conditions, the rate of transfer of hydrogen atoms by the shuttle is less than the rate of NADH generation so that the redox state in the cytosolic compartment of the liver becomes highly reduced and the concentration of NAD severely decreased. This limits the rate of ethanol oxidation by alcohol dehydrogenase. 

The majority of synthetic reactions in mammalian cells takes place in the cytosol. The intramitochondrial localization of transhydrogenase excludes a direct participation in these anabolic processes. Substrate shuttle mechanisms (176, 177) are required to allow for the interaction between intra- and extramitochondrial nicotinamide nucleotide-dependent reactions. In the first instance transhydrogenase can be regarded to be functionally related to intramitochondrial NADP-linked reactions. A number of studies on isolated mitochondria have elaborated these relationships in some detail, in particular with regard to mitochondrial monooxygenation reactions and to the metabolism of glutamate and isocitrate. 

Figure 21 Characterization of aminoacyl transferase CmaE in coronamic acid biosynthesis pathway, (a) Within the biosynthetic pathway, CmaE carries out substrate shuttling from theCmaAT domain to the CmaDT domain, (b) CmaE substrate tolerance was characterized by MALDI-TOF MS (observed and calculated mass shift of CmaD in the table). In addition, evidence of reversible aminoacyl transfer by CmaE was detected.

Jenni, S., Leibundgut, M., Boehringer, D., Frick, C., Mikolasek, B., Ban, N. 2007. Structure of fungal fatty acid synthase and implications for iterative substrate shuttling. Science 316 254-261. 

Jenni, S., M. Leibundgut, D. Boehringer, C. Frick, B. Mikolasek, B., and N. Ban. Structure of Fungal Fatty Acid Synthase and Implications for Iterative Substrate Shuttling. Science 316, 254—261 (2007). [An in-depth look at the reaction sites for a multiple-subunit enzyme]. 

The mitochondrial inner membrane is impermeable to NAD, and therefore the NADH produced in the cytosol in glycolysis cannot enter the mitochondria for reoxidation. In order to transfer the reducing equivalents from cytosolic NADH into the mitochondria, two substrate shuttles are used ... 

In red blood cells, which lack mitochondria, reoxidation of NADH formed in glycolysis cannot be by way of the substrate shuttles discussed above (section 5.4.1.1) and the electron transport chain. 

In bulk coating processes, bulk materials are joined to the substrate either by a surface melt process or by attachment of the soHd material. An example of the latter is the appHcation of heat-resistant tiles of sHica-type material to the aluminum alloy skin of a space shuttle vehicle, enabling the vehicle to withstand the reentry heat. 

FIGURE 25.1 The citrate-malate-pyruvate shuttle provides cytosolic acetate units and reducing equivalents (electrons) for fatty acid synthesis. The shuttle collects carbon substrates, primarily from glycolysis but also from fatty acid oxidation and amino acid catabolism. Most of the reducing equivalents are glycolytic in origin. Pathways that provide carbon for fatty acid synthesis are shown in blue pathways that supply electrons for fatty acid synthesis are shown in red. 

Mediated electrolyses make use of electron transfer mediators PjQ that shuttle electrons between electrodes and substrates S, avoiding adverse effects encountered with the direct heterogeneous reaction of substrates at electrode surfaces (Scheme 6). In recent years this mode of electrochemical synthesis has been widely studio and it is becoming increasingly well understood. A review is given in vol 1 of the present electrochemistry series... 

Coenzymes serve as recyclable shuttles—or group transfer reagents—that transport many substrates from their point of generation to their point of utilization. Association with the coenzyme also stabilizes substrates such as hydrogen atoms or hydride ions that are unstable in the aqueous environment of the cell. Other chemical moieties transported by coenzymes include methyl groups (folates), acyl groups (coenzyme A), and oligosaccharides (dolichol). 

The microbial degradation of contaminants under anaerobic conditions using humic acids as electron acceptors has been demonstrated. These included the oxidations (a) chloroethene and 1,2-dichloroethene to CO2 that was confirmed using C-labeled substrates (Bradley et al. 1998) and (b) toluene to CO2 with AQDS or humic acid as electron acceptors (Cervantes et al. 2001). The transformation of l,3,5-trinitro-l,3,5-triazine was accomplished using Geobacter metallireducens and humic material with AQDS as electron shuttle (Kwon and Finneran 2006). 

The high catalytic activity of enzymes has a number of sources. Every enzyme has a particular active site configured so as to secure intimate contact with the substrate molecule (a strictly defined mutual orientation in space, a coordination of the electronic states, etc.). This results in the formation of highly reactive substrate-enzyme complexes. The influence of tfie individual enzymes also rests on the fact that they act as electron shuttles between adjacent redox systems. In biological systems one often sees multienzyme systems for chains of consecutive steps. These systems are usually built into the membranes, which secures geometric proximity of any two neighboring active sites and transfer of the product of one step to the enzyme catalyzing the next step. 

Ferric iron can act as an electron acceptor under the anaerobic conditions the azo dye is in. Like sulfate, it was found that addition of ferric iron to the reactor stimulates the azo dye reduction. Indeed, the reactions are dealing with the redox couple Fe (III)/Fe (II), which can act as an electron shuttle for transferring electrons from electron donor to the electron accepting azo dye. Meanwhile, reactions of both reduction of Fe (III) to Fe (II) and oxidation of Fe (II) to Fe (III) facilitate the electron transport from the substrate to azo dye, thus acting as an extracellular redox mediator [31]. 

As the proton shuttles from the 04 to N5, there is a barrier of about 8.8 kcal/mol at which point the 04-H04 distance is 1.4 A. Once this barrier is crossed, the system becomes more stabilized. Following proton transfer the energy is reduced by -6.9 kcal/mol. This series of findings support the possibility that an energetically feasible mechanism can be proposed for proton transfer from 04 to N5 via an intermediate water molecule. But as has been discussed earlier, there was no solvent molecule in direct contact with either 04 or N5 of the substrate during these simulations. 

The astrocyte-neuron lactate shuttle hypothesis is controversial. In recent years the possibility that lactate, formed within the brain and released by astrocytes, is an important neuronal substrate both for energy and incorporation into neurotransmitters has been the subject of many studies and considerable controversy. There is evidence that suggests transient release of lactate in human brain on stimulation [48,8,88], Little is known about the highly active metabolism that takes place in the many elaborate, lamellar distal processes of astrocytes dispersed through the neuropil and interacting with an estimated >100,000 synapses [82, and references therein]. However, it is well established that astrocytes do respond to neuronal activity [89], For example, in the isolated mouse optic nerve preparation, upon stimulation, astrocytic glycogen... 

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