Vitamin B6 Enzyme Models

Copyright 2005 WILEY-VCH Verlag GmbH Co. KGaA, Weinheim ISBN 3-527-31165-3 

A basic group of the enzyme catalyzes the proton transfer characteristics of transaminations. Bruice found that imidazole buffer can catalyze transamination reactions in model systems [6], We conjectured that transamination rates should be improved by attaching a basic side arm to pyridoxamine. Thus we synthesized a series of simple pyridoxamine derivatives carrying basic groups at the end of flexible chains of various lengths [7-9]. We measured the transamination rates of these pyridoxamine derivatives in the presence of Zn(OAc)2 at pH 4.00 in methanol. 

C4 carbon, the dimethylamino group then transferred the proton to the a-carbon of the developing amino acid. Thus this preference for increased chain length seems to argue for such a dual catalytic role of the amine. As expected from this, compound 6 with yet an additional methylene group, which is no longer needed to permit the a-carbon to be reached, is now less active. 

Thus we designed and synthesized a bicyclic pyridoxamine derivative carrying an oriented catalytic side arm (16) [11], Rates for conversion of the ketimine Schiff base into the aldimine, formed with 26 (below) and a-ketovaleric acid, indolepyruvic acid, or pyruvic acid, were enhanced 20-30 times relative to those carried out in the presence of the corresponding pyridoxamine derivatives without the catalytic side arm. With a-ketovaleric acid, 16 underwent transamination to afford D-norvaline with 90% ee. The formation of tryptophan and alanine from indolepyruvic acid and pyruvic acid, respectively, showed a similar preference. A control compound (17), with a propylthio group at the same stereochemical position as the aminothiol side arm in 16, produced a 1.5 1 excess of L-norvaline, in contrast to the large preference for D-amino acids with 16. Therefore, extremely preferential protonation seems to take place on the si face when the catalytic side arm is present as in 16. 

A further method to induce chirality in the pyridoxamine-mediated transamination reactions was developed by Kuzuhara et al. [13]. They synthesized optically resolved pyridinophanes (21, 22) having a nonbranched ansa chain between the 2 - and 5 -positions of pyridoxamine. With the five-carbon chain in 21 and 22, the two isomers do not interconvert readily. In the presence of zinc(n) in organic solvents such as methanol, tert-butanol, acetonitrile, and nitromethane, they observed stereoselective transamination between pyridinophanes and keto acids. The highest ee%s are 95 % for d-and L-leucine by reaction of the corresponding a-keto acid with (S)- and (R)- 22, respectively. On the basis of kinetic analysis of the transamination reactions, Kuzuhara et al. originally proposed a mechanism for the asymmetric induction through kinetically controlled stereoselective protonation to the carboanion attached to an octahedral Zn(n) chelate intermediate. However, they subsequently raised some questions about this proposal [14]. 

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Vitamin B6 enzyme models that can catalyze five types of reactions - transamination, racemization, decarboxylation, P-elimination and replacement, and aldolase-type reactions - have been reviewed. There are also five approaches to construct the vitamin B6 enzyme models (i) vitamin B6 augmented with basic or chiral auxiliary functional groups (ii) vitamin B6 having an artificial binding site (iii) vitamin B6-surfactant systems (iv) vitamin B6-polypeptide systems (v) polymeric and dendrimeric vitamin B6 systems. These model systems show rate enhancement and some selectivity in vitamin B6-dependent reactions, but they are still primitive compared with the real enzymes. We expect to see improved reaction rates and selectivities in future generations of vitamin B6 enzyme models. An additional goal, which has not received ade-... 

Liu L, Breslow R. Vitamin B6 enzyme models. In Artificial Enzymes. Breslow R, ed. 2005. Wiley-VCH. Weinheim, Germany, pp. 37-62. 

Pyridoxal phosphate is the coenzyme for the enzymic processes of transamination, racemization and decarboxylation of amino-acids, and for several other processes, such as the dehydration of serine and the synthesis of tryptophan that involve amino-acids (Braunstein, 1960). Pyridoxal itself is one of the three active forms of vitamin B6 (Rosenberg, 1945), and its biochemistry was established by 1939, in considerable part by the work of A. E. Braunstein and coworkers in Moscow (Braunstein and Kritzmann, 1947a,b,c Konikova et al 1947). Further, the requirement for the coenzyme by many of the enzymes of amino-acid metabolism had been confirmed by 1945. In addition, at that time, E. E. Snell demonstrated a model reaction (1) for transamination between pyridoxal [1] and glutamic acid, work which certainly carried with it the implication of mechanism (Snell, 1945). 

An important feature of the enzymatic systems is the presence of a binding site. Thus it is attractive to learn how to construct vitamin B6-dependent enzyme models that can provide a substrate binding site and perform molecular recognition. The first example was a catalyst (23) in which pyridoxamine was linked to the primary face of [1-cyclo-dextrin (P-CD) through a sulfur atom [15]. Catalyst 23 could transform a-keto acids into a-amino acids, as pyridoxamine does, but with selectivity. That is, phenylpyruvic acid... 

Modeling vitamin B6 metabolism is further complicated by the fact that the activity of the kinase, oxidase, and phosphatase enzymes varies between organs and species. A very simplified diagram of vitamin B6 metabolism is shown in Fig. 2. In the intestine any phosphoiylated forms are hydrolyzed. The free vitamers are readily taken up by diffusion into the intestinal wall where significant phosphorylation (Middleton, 1979) and other metabolism (Middleton, 1985) occurs. In mice small doses (up to 14 nmol) of pyridoxine (Sakurai et aL, 1988) and pyridoxamine (Sakurai et oL, 1992) were converted almost completely to pyridoxal before being released into the portal circulation. While it is dear that the intestinal microflora produce vitamin B6,... 

Vitamin B plays important roles in cell metabolism. Vitamin B actually consists of eight chemically distinct biologically active agents that function as coenzymes. Pyridoxine, pyridoxal, and pyridoxamine or pyridoxine hydrochloride can all be called vitamin B6 as they are all converted to the active form. Pyridoxine is involved in the metabolism of amino acids and lipids. A vitamin B6-dependent enzyme model... 

FIGURE 12.3 Pridoxamine-PCDs as vitamin B6-dependent enzyme model. 

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