Thiamines, reactions with base

Addition of thiamine. The conversion of pyruvate to acetyl CoA begins by reaction of pyruvate with thiamine pyrophosphate, a derivative of vitamin B, The hydrogen on the heterocyclic thiamine pyrophosphate is weakly acidic and can be removed by reaction with base to yield a nucleophilic ylide much like the phosphorus ylides u.sed in Wittig reactions /Section 19.12>. This nucleophilic yJide adds to the ketone carbonyl group of pyruvate to yield a tetrahedral intermediate. 

Studies on thiamine (vitamin Bi) catalyzed formation of acyloins from aliphatic aldehydes and on thiamine or thiamine diphosphate catalyzed decarboxylation of pyruvate have established the mechanism for the catalytic activity of 1,3-thiazolium salts in carbonyl condensation reactions. In the presence of bases, quaternary thiazolium salts are transformed into the ylide structure (2), the ylide being able to exert a cat ytic effect resembling that of the cyanide ion in the benzoin condensation (Scheme 2). Like cyanide, the zwitterion (2), formed by the reaction of thiazolium salts with base, is nucleophilic and reacts at the carbonyl group of aldehy s. The resultant intermediate can undergo base-catalyzed proton... 

Thiamine, or vitamin Bi, is a water-soluble compound which is rapidly broken down by moist heat in neutral or alkaline solutions into its constituent pyrimidine and thiazole rings. The ready destructability of thiamine is important in human nutrition, since much may be lost in the preparation of food. Some of the biochemical methods used in evaluating thiamine nutrition are based on reactions with the thiazole and pyrimidine portions of the thiamine molecule. The thiochrome method is widely used in assaying biological materials for thiamine, while determination of the urinary excretion of pyramine (a pyrimidine-like compound) has been used to assist in assessment of nutritional status. 

Thiamine is among the least stable of the vitamins. The reaction catalysed by thiaminase 1, which is, for example, present in raw fish, lies in the cleavage of the thiamine molecule by an exchange reaction with nitrogen bases (such as nicotinic acid) or thiols (such as cysteine). Thiaminase 11 (found in some microorganisms) catalyses thiamine hydrolysis with the formation of the same products that form by non-enzymatic hydrolysis (Figure 5.10). 

Transketolase requires the cofactor thiamine pyrophosphate (TPP), which stabilizes a two-carbon car-banion in this reaction (Fig. 14—26a), just as it does in the pyruvate decarboxylase reaction (Fig. 14-13). Transaldolase uses a Lys side chain to form a Schiff base with the carbonyl group of its substrate, a ketose,... 

Below the structures of the adducts in Eq. 14-20 are those of a 2-oxo acid and a (3-ketol with arrows indicating the electron flow in decarboxylation and in the aldol cleavage. The similarities to the thiamin-dependent cleavage reaction are especially striking if one remembers that in some aldolases and decarboxylases the substrate carbonyl group is first converted to an N-proto-nated Schiff base before the bond cleavage. 

Nucleophilic catalysis is a specific example of covalent catalysis the substrate is transiently modified by formation of a covalent bond with the catalyst to give a reactive intermediate. There are also many examples of electrophilic catalysis by covalent modification. It will be seen later that in the reactions of pyridoxal phosphate, Schiff base formation, and thiamine pyrophosphate, electrons are stabilized by delocalization. 

Based on the conventional analysis of the mechanism of decarboxylation of thiamin-derived intermediates, there is no role for a catalyst in the carbon-carbon bond-breaking step of this reaction. The thiazolium nitrogen is at its maximum electron deficiency with no available coordination sites. Ultimately, there is no place for a proton or other cation to position itself in order to promote the reaction by stabilizing a transition state that resembles the product of the reaction. Since there is no role for an acid, base, or metal to accelerate the decarboxylation of these intermediates by stabilizing the transition state for C-C bond-breaking, the means by which this could be achieved became a source of interest and speculation. 

This looks like a simple reaction based on very small molecules. But look again. It Is a very strange reaction indeed. The molecule of CO2 clearly comes from the carboxyl group of pyruvate, but how is the C-C bond cleaved, and how does acetyl CoA Join on If you try to draw a mechanism you will see that there must be more to this reaction than meets the eye. The extra features are two new cofactors, thiamine pyrophosphate and lipoic acid, and the reaction takes place in several stages with some interesting chemistry involved. 

Part of the dark reactions of photosynthesis is interconversion of sugars with an enzyme called transketolase using thiamine pyrophosphate, TPP, as a catalyst (Section 8.12.8). Provide a reasonable mechanism for this enzymatic reaction. In addition to water, there are weak general acids and general bases present in the active site at pH 7. 

An understanding of mechanisms of thiamin pyrophosphate-dependent processes must begin with the classic work of Breslow 105, 134), who showed that the hydrogen at C-2 of thiamin pyrophosphate can be removed by bases and that the resulting anion is highly reactive. The at this site is 18 135). In an enzyme active site, this p a value may be considerably lower, as the value decreases with decreasing medium polarity. Reaction of the anion with a variety of carbonyl compounds (e.g., acetaldehyde, pyruvate) gives rise to characterizable adducts 132). 

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