Enzymes and reduction

Interactions between tannins and proteins have been extensively studied (Hager-man 1989 Haslam and Lilley 1988 Haslam et al. 1992), owing to their role in haze formation, astringency perception, and nutritional and anti-nutritional effects resulting from inhibition of various enzymes and reduction of dietary protein digestion. Other effects include reduced adsorption of /3-casein at the air-liquid interface in the presence of epigallocatechin gallate with potential consequences on foam properties (Sausse et al. 2003). 

Disulfides. As shown in Figure 4, the and h-chains of insulin are connected by two disulfide bridges and there is an intrachain cycHc disulfide link on the -chain (see Insulin and other antidiabetic drugs). Vasopressin [9034-50-8] and oxytocin [50-56-6] also contain disulfide links (48). Oxidation of thiols to disulfides and reduction of the latter back to thiols are quite common and important in biological systems, eg, cysteine to cystine or reduced Hpoic acid to oxidized Hpoic acid. Many enzymes depend on free SH groups for activation—deactivation reactions. The oxidation—reduction of glutathione (Glu-Cys-Gly) depends on the sulfhydryl group from cysteine. 

The abihty of iron to exist in two stable oxidation states, ie, the ferrous, Fe ", and ferric, Fe ", states in aqueous solutions, is important to the role of iron as a biocatalyst (79) (see Iron compounds). Although the cytochromes of the electron-transport chain contain porphyrins like hemoglobin and myoglobin, the iron ions therein are involved in oxidation—reduction reactions (78). Catalase is a tetramer containing four atoms of iron peroxidase is a monomer having one atom of iron. The iron in these enzymes also undergoes oxidation and reduction (80). 

Ammonia reacts with the ketone carbonyl group to give an iinine (C=NH), which is then reduced to the anine function of the a-anino acid. Both iinine fonnation and reduction are enzyme-catalyzed. The reduced fonn of nicotinamide adenine diphosphonu-cleotide (NADPH) is a coenzyme and acts as a reducing agent. The step in which the iinine is reduced is the one in which the chirality center is introduced and gives only L-glutfflnic acid. 

The next three steps—reduction of the /3-carbonyl group to form a /3-alcohol, followed by dehydration and reduction to saturate the chain (Figure 25.7) — look very similar to the fatty acid degradation pathway in reverse. However, there are two crucial differences between fatty acid biosynthesis and fatty acid oxidation (besides the fact that different enzymes are involved) First, the alcohol formed in the first step has the D configuration rather than the L form seen in catabolism, and, second, the reducing coenzyme is NADPH, although NAD and FAD are the oxidants in the catabolic pathway. 

The citric acid cycle, a nine-step process, also diverts chemical energy to the production of ATP and the reduction of NAD and FAD. In each step of the citric acid cycle (also known as the Krebs cycle) a glucose metabolite is oxidized while one of the carrier molecules, NAD or FAD, is reduced. Enzymes, nature s chemical catalysts, do a remarkable job of coupling the oxidation and reduction reactions so that energy is transferred with great efficiency. 

Enzymes are classified into six categories depending on the kind of reaction they catalyze, as shown in Table 26.2. Oxidoreductases catalyze oxidations and reductions hansferases catalyze the transfer of a group from one substrate to another hydrolases catalyze hydrolysis reactions of esters, amides, and related substrates lyases catalyze the elimination or addition of a small molecule such as H2O from or to a substrate isomerases catalyze isomerizalions and ligases catalyze the bonding together of two molecules, often coupled with the hydrolysis... 

Assuming that the enzymatic reaction is highly enantioselective, then even after only four cycles the enantiomeric excess will have reached 93.4% whereas after seven catalytic cycles the enantiomeric excess is >99% (Figure 5.3). This type of deracemization is really a stereoinversion process in that the reactive enantiomer undergoes stereoinversion during the process. One of the challenges of developing this type of process is to find conditions under which the enzyme catalyst and chemical reactant can coexist, particularly in the case of redox chemistry in which the coexistence of an oxidant and reductant in the same reaction vessel is difficult to achieve. For this... 

A representative set of a- and -keto esters was also tested as substrates (total 11) for each purified fusion protein (Figure 8.13b,c) [9bj. The stereoselectivities of -keto ester reductions depended both on the identity of the enzyme and the substrate stmcture, and some reductases yielded both l- and o-alcohols with high stereoselectivities. While a-keto esters were generally reduced with lower enantioselec-tivities, it was possible to identify pairs of yeast reductases that delivered both alcohol antipodes in optically pure form. These results demonstrate the power of genomic fusion protein libraries to identify appropriate biocatalysts rapidly and expedite process development. 

Another example of dynamic kinetic resolution is the reduction of a sulfur-substituted ketone. Thus, yeast reduction of (R/S)-2-(4-methoxyphenyl)-l, 5-benzothiazepin-3,4(2H, 5H)-dione gave only (2S, 3S)-alcohol as a product out of four possible isomers as shown in Figure 8.39c [29kj. Only (S)-ketone was recognized by the enzyme as a substrate and reduction of the ketone proceeded... 

It is probable that the negative charge induced by these three electrons on FeMoco is compensated by protonation to form metal hydrides. In model hydride complexes two hydride ions can readily form an 17-bonded H2 molecule that becomes labilized on addition of the third proton and can then dissociate, leaving a site at which N2 can bind (104). This biomimetic chemistry satisfyingly rationalizes the observed obligatory evolution of one H2 molecule for every N2 molecule reduced by the enzyme, and also the observation that H2 is a competitive inhibitor of N2 reduction by the enzyme. The bound N2 molecule could then be further reduced by a further series of electron and proton additions as shown in Fig. 9. The chemistry of such transformations has been extensively studied with model complexes (15, 105). 

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