Hydroxylation enzyme catalysis

The effects of the feed ratio in the lipase CA-catalyzed polymerization of adipic acid and 1,6-hexanediol were examined by using NMR and MALDI-TOF mass spectroscopies. NMR analysis showed that the hydroxyl terminated product was preferentially formed at the early stage of the polymerization in the stoichiometric substrates. As the reaction proceeded, the carboxyl-terminated product was mainly formed. Even in the use of an excess of the dicarboxylic acid monomer, the hydroxy-terminated polymer was predominantly formed at the early reaction stage, which is a specific polymerization behavior due to the unique enzyme catalysis. 

In the field of enzyme catalysis, heme-proteins such as cytochrome P450, for example, exhibit both types of 0-0 bond cleavages in organic hydroperoxides and peroxy acids (178). Heterolytic cleavage of HOOH/ROOH yields H20 or the corresponding alcohol, ROH and a ferryl-oxo intermediate (Scheme 4). Homolytic 0-0 bond cleavage results in the formation of a hydroxyl (HO ) or an alkoxyl (RO ) radical and an iron-bound hydroxyl radical. 

The NAD+-dependent alcohol dehydrogenase from horse liver contains one catalytically essential zinc ion at each of its two active sites. An essential feature of the enzymic catalysis appears to involve direct coordination of the enzyme-bound zinc by the carbonyl and hydroxyl groups of the aldehyde and alcohol substrates. Polarization of the carbonyl group by the metal ion should assist nucleophilic attack by hydride ion. A number of studies have confirmed this view. Zinc(II) catalyzes the reduction of l,10-phenanthroline-2-carbaldehyde by lV-propyl-l,4-dihy-dronicotinamide in acetonitrile,526 and provides an interesting model reaction for alcohol dehydrogenase (Scheme 45). The model reaction proceeds by direct hydrogen transfer and is absolutely dependent on the presence of zinc(II). The zinc(II) ion also catalyzes the reduction of 2- and 4-pyridinecarbaldehyde by Et4N BH4-.526 The zinc complex of the 2-aldehyde is reduced at least 7 x 105 times faster than the free aldehyde, whereas the zinc complex of the 4-aldehyde is reduced only 102 times faster than the free aldehyde. A direct interaction of zinc(II) with the carbonyl function is clearly required for marked catalytic effects to be observed. 

Since a number of the studies we shall review were concerned with the effect of synthetic chain molecules or micelles on the hydrolysis rate of nitrophenyl acetate and similar esters, it will be useful to consider briefly some characteristics of the enzymic catalysis of this process. A particularly detailed study has been carried out on the enzyme chymo-trypsin (14) and a great deal of evidence shows that the catalytic site of this enzyme contains a serine residue with an unusually reactive hydroxyl group. Denoting the chymotrypsin by Ch—OH, the interaction with the ester involves first acyl transfer to the enzyme and this is followed by acyl enzyme hydrolysis to regenerate Ch—OH ... 

In the first reaction, intramolecular attack by the hydroxyl on the carboxyl carbonyl proceeds 5 X 108 times faster than the corresponding inter-molecular process (2). In the second reaction, the rate increases 107-fold upon switching the solvent from methanol to dimethylformamide (3). Obviously, the huge rate increases in these organic systems do not necessarily prove that similar effects are at work in enzymes. But to be suspicious is quite natural, and many people, too numerous to mention, have pointed out the possible relationship between enzyme catalysis and intramolecularity or solvation effects. 

What has been described as biomimetic control of chemical selectivity is already possible. When the steroid 3a-cholestanol is esterified with 4-iodophenylacetic acid and treated with chlorine in the dark the iododichloride can generate free radicals which attack the C-17 of the steroid. The shorter ester derived from benzoic acid attacks C-9 (Figure 6.33). In another context metal porphyrin complexes have been devised which can hydroxylate hydrocarbons (Figure 6.34). Many more ideas of this kind are likely to follow a better understanding of the mechanisms of enzyme catalysis. 

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