Thiamin requirements

A number of lyases are known which, unlike the aldolases, require thiamine pyrophosphate as a cofactor in the transfer of acyl anion equivalents, but mechanistically act via enolate-type additions. The commercially available transketolase (EC 2.2.1.1) stems from the pentose phosphate pathway where it catalyzes the transfer of a hydroxyacetyl fragment from a ketose phosphate to an aldehyde phosphate. For synthetic purposes, the donor component can be replaced by hydroxypyruvate, which forms the reactive intermediate by an irreversible, spontaneous decarboxylation. 

Now this reaction is effectively a repeat of the pyruvate acetyl-CoA oxidative decarboxylation we saw at the beginning of the Krebs cycle. It similarly requires thiamine diphosphate, lipoic acid, coenzyme A and NAD+. A further feature in common with that reaction is that 2-oxoglutarate dehydrogenase is also an enzyme complex comprised of three separate enzyme activities. 2-Oxoglutarate is thus transformed into succinyl-CoA, with the loss of... 

Pyruvate dehydrogenase (lipoamide) [EC 1.2.4.1], which requires thiamin pyrophosphate, catalyzes the reaction of pyruvate with lipoamide to produce 5-acetyldihydroli-poamide and carbon dioxide. It is a component of the pyruvate dehydrogenase complex (which also includes dihydrolipoamide dehydrogenase [EC 1.8.1.4] and dihy-drolipoamide acetyltransferase [EC 2.3.1.12]). Pyruvate dehydrogenase (cytochrome) [EC 1.2.2.2] catalyzes the... 

This enzyme [EC 1.2.3.3], which requires thiamin pyrophosphate and FAD, catalyzes the reaction of pyruvate with orthophosphate, dioxygen, and water to produce acetyl phosphate, carbon dioxide, and hydrogen peroxide. 

The project encompassed the comparative characterization of pyruvate decarboxylase from Z. mohilis (PDC) and benzoylformate decarboxylase from P. putida (BED) as well as their optimization for bioorganic synthesis. Both enzymes require thiamine diphosphate (ThDP) and magnesium ions as cofactors. Apart from the decarboxylation of 2-ketoacids, which is the main physiological reaction of these 2-ketoacid decarboxylases, both enzymes show a carboligase site reaction leading to chiral 2-hydroxy ketones (Scheme 2.2.3.1). A well-known example is... 

The first examples of mechanism must be divided into two principal classes the chemistry of enzymes that require coenzymes, and that of enzymes without cofactors. The first class includes the enzymes of amino-acid metabolism that use pyridoxal phosphate, the oxidation-reduction enzymes that require nicotinamide adenine dinucleotides for activity, and enzymes that require thiamin or biotin. The second class includes the serine esterases and peptidases, some enzymes of sugar metabolism, enzymes that function by way of enamines as intermediates, and ribonuclease. An understanding of the mechanisms for all of these was well underway, although not completed, before 1963. 

The conversion of pyruvate to ethanol occurs by the two reactions summarized in Figure 8.24. The decarboxylation of pyruvate by pyruvate decarboxylase occurs in yeast and certain microorganisms, but not in humans. The enzyme requires thiamine pyrophosphate as a coenzyme, and catalyzes a reaction similar to that described for pyruvate dehydrogenase (see p. 108). 

Coenzymes The pyruvate dehydrogenase complex contains five coenzymes that act as carriers or oxidants for the intermediates of the reactions shown in Figure 9.3. Ei requires thiamine pyrophosphate, Ep requires lipoic acid and coenzyme A, and E3 requires FAD and NAD+. [Note Deficiencies of thiamine or niacin can cause serious central nervous system problems. This is because brain cells are unable to produce sufficient ATP (via the TCA cycle) for proper function if pyruvate dehydrogenase is inactive.]... 

In most organisms undergoing aerobic metabolism, pyruvate is oxidized to acetyl-CoA in a complex process involving its decarboxylation (Eq. 10-6). This oxidative decarboxylation, like the decarboxylation of pyruvate to acetaldehyde, requires thiamin diphosphate. In addition, an array of other catalysts participate in the process (see Fig. 15-15). Among these are the electron carrier flavin adenine diphosphate (FAD), which is derived from the vitamin riboflavin. Like NAD+, this... 

Transketolase is one of several enzymes that catalyze reactions of intermediates with a negative charge on what was initially a carbonyl carbon atom. All such enzymes require thiamine pyrophosphate (TPP) as a cofactor (chapter 10). The transketolase reaction is initiated by addition of the thiamine pyrophosphate anion to the carbonyl of a ketose phosphate, for example xylulose-5-phosphate (fig. 12.33). The adduct next undergoes an aldol-like cleavage. Carbons 1 and 2 are retained on the enzyme in the form of the glycol-aldehyde derivative of TPP. This intermediate condenses with the carbonyl of another aldolase. If the reactants are xylulose-5-phosphate and ribose-5-phosphate, the products are glyceraldehyde-3-phosphate and the seven-carbon ketose, sedoheptulose-7-phosphate (see fig. 12.33). 

The conversion of pyruvate to acetyl-CoA. The reactions are catalyzed by the enzymes of the pyruvate dehydrogenase complex. This complex has three enzymes pyruvate decarboxylase, dihydrolipoyl transacetylase, and dihydrolipoyl dehydrogenase. In addition, five coenzymes are required thiamine pyrophosphate, lipoic acid, CoASH, FAD, and NAD+. Lipoic acid is covalently attached to... 

Transketolase from common yeast (Saccharomyces cerevisiae) is commercially available, but it is possible to work with a partially purified enzyme, isolated with little expense from spinach leaves.54 Transketolase catalyzes the transfer of a hydroxyacetyl group, reversibly from a ketose phosphate, or irreversibly from hydroxypyruvate to an acceptor aldose, phosphorylated or not.55 It requires thiamine pyrophosphate as a coenzyme, but only in catalytic amounts. In all the cases listed in Table V, the new chiral center, C-3 of the ketose, has the l-glycero configuration. 

The immobilized enzyme requires thiamine pyrophosphate, calcium ions, and FAD for efficient activity (92,235,236). Concentrations as low as 1 pM have been determined (92). 

Transketolase, which requires thiamine pyrophosphate, transfers two-carbon units (Figure 5-33). 

Glyceraldehyde 3-phosphate in a reaction requiring thiamine pyrophosphate... 

D. Both transaldolase and transketolase produce glyceraldehyde 3-phosphate, but only transketolase requires thiamine pyrophosphate. 

Degradation of all three branched-chain amino acids begins with a transamination followed by an oxidative decarboxylation catalyzed by the branched-chain a-keto acid dehydrogenase complex. This enzyme, like a-ketoglutarate dehydrogenase, requires thiamine pyrophosphate, lipoic acid, coenzyme A, FAD, and NAD+ (Figure 7-11). 

D. The branched-chain amino acids (valine, isoleucine, and leucine) are transaminated and then oxidatively decarboxylated by an enzyme that requires thiamine, lipoic add, coenzyme A, FAD, and NAD. 

A. The transketolase of the pentose phosphate pathway requires thiamine pyrophosphate. 

I d like to give an example of how this works. When the B vitamin thiamin is destroyed by boiling or overcooking, it is not available to work as a coenzyme with four different enzymes that require thiamin before they can break down carbohydrates to produce energy. What happens then The usual series of steps to make the energy is broken, and intermediate proteins from the disrupted process build up to toxic levels in the cells. Other enzymes and macrophages are diverted from other tasks to detoxify the overload of useless proteins. 

Transfer of a two-carbon unit from a 2-keto sugar to the carbonyl carbon (Ci) of an aldose by a transketolase, which requires thiamine pyrophosphate and magnesium as cofactors. A covalent enzyme-substrate intermediate is formed similar to the one that occurs during the pyruvate dehydrogenase reaction (Chapter 13). 

In the course of evolution, nature has devised a multitude of enzymes which are capable of stereoselectively forming C-C bonds in vivo. Depending on the kind of substrate employed, nucleophilic acylation reactions are accomplished in nature by means of different lyases such as transketolases and pyruvate decarboxylases, which all require thiamine (1) as coenzyme [1,2,3,4,5,6]. For the synthetic organic chemist, asymmetric catalytic C-C bond formation reactions, in... 

The decarboxylation of pymvic acid to yield carbon dioxide and a two-carbon fragment is catalyzed by a class of enzymes that require thiamin pyrophosphate (TPP) and a divalent metal ion. We will restrict our attention to the simplest member of the class, pyruvate decarboxylase 132, 133), which catalyzes reaction 18). 

The process is initiated by the entry of pyruvate into the mitochondrial matrix, where the enzyme pyruvate dehydrogenase will irreversibly convert pyruvate to acetyl CoA. This enzyme is a complex of a large number of subunits of independent but cooperating individual enzymic activities. (1) The first subunit (pyruvate decarboxylase) causes the decarboxylation of pyruvate and requires thiamine pyrophosphate. A twocarbon thiamine pyrophosphate adduct is formed and then transferred from this subunit to a second subunit containing oxidized lipoic acid (i.e., S-S). (2) The twocarbon adduct is transferred to form an HS and... 

---