Transketolase and

MORE NADPH THAN RmOSE-5-P IS NEEDED BY THE CELL Large amounts of N/VDPH can be supplied for biosynthesis without concomitant production of ribose-5-P, if ribose-5-P produced in the pentose phosphate pathway is recycled to produce glycolytic intermediates. As shown in Figure 23.39, this alternative involves a complex interplay between the transketolase and transaldolase reac-... 

Figure 10.37 Kinetic resolution by transketolase, and nonequilibrium C—C bond formation by decomposition of hydroxypyruvate.

Figure 18 Reaction mechanism of transketolase and the stereochemical course.

Subsequent studies196 on crystalline transketolase have revealed the presence of a contaminating enzyme termed pentulose 5-phosphate waldenase (or epimerase) the presence of which had led to the erroneous conclusion that d-erythro-pentulose 5-phosphate was the substrate for transketolase. d-erythro-Pentulose 5-phosphate is virtually unattacked by transketolase prepared from spinach or liver. In subsequent discussions of experiments involving the use of transketolase, in this article, the enzymic reactions must be viewed as the result of action of transketolase and pentulose 5-phosphate waldenase (epimerase). 

The above transketolase and transaldolase reactions were found inadequate to explain the metabolism of D-ribose 5-phosphate, because of the non-accumulation of tetrose phosphate, the 75 % yield of hexose phosphate, and the results of experiments with C14 (the distribution of which differed markedly from the values predicted for such a sequence). 24(b) Thus, with D-ribose-l-C14, using rat-liver enzymes, any hexose formed should have equal radioactivity at Cl and C3, whereas, actually, 74% appeared at Cl. Furthermore, D-ribose-2,3-Cl42 should have given material having equal labels at C2 and C4 in the resultant hexose, whereas, in fact, it had 50% of the activity at C4, C3 was nearly as active as C2, and Cl had little activity. Similar results were obtained with pea-leaf and -root preparations.24 The following reactions, for which there is enzymic evidence,170(b) were proposed, in addition to those involving D-aftro-heptulose, to account for these results.24(b) (o) 200... 

Transketolase and transaldolase appear to play a significant part in photosynthesis.86 Photosynthesizing plants have been exposed to C14C>2 for short periods (up to 15 seconds) and the radioactive products have been... 

In contrast to transketolase and the DHAP-dependent aldolases, deoxyribose aldolase (DERA) catalyzes the aldol reaction with the simple aldehyde, acetaldehyde. In vivo it catalyzes the formation of 2-deoxyribose-5-phosphate, the building block of DNA, from acetaldehyde and D-glyceraldehyde-3-phosphate, but in vitro it can catalyze the aldol reaction of acetaldehyde with other non-phosphorylated aldehydes. The example shown in Scheme 6.28 involves a tandem aldol reaction... 

Thiamine (vitamin Bi) is phosphorylated by ATP to thiamine pyrophosphate. This is a coenzyme for, among others, alpha-ketoglutarate dehydrogenase, transketolase and pyruvate dehydrogenase. Thiamine pyrophosphate is involved in fatty acid... 

TALDO deficiency can be confirmed in lymphoblasts, fibroblasts and in erythrocytes. These cells are incubated with ribose-5-phosphate, after which formation of transketolase and TALDO products are analysed by gas chromatography with nitrogen phosphorous detection by liquid chromatography tandem mass spectrometry [8, 11]. A similar enzyme assay is available for RPI [2]. Confirmation of the gene defect can be performed by sequence analysis. Disease-causing mutations have been detected in all TALDO-deficient patients and in the RPI-deficient patient. 

FIGURE 14-22 Nonoxidative reactions of the pentose phosphate pathway, (a) These reactions convert pentose phosphates to hexose phosphates, allowing the oxidative reactions (see Fig. 14-21) to continue. The enzymes transketolase and transaldolase are specific to this pathway the other enzymes also serve in the glycolytic or gluconeogenic pathways, (b) A schematic diagram showing the pathway... 

FIGURE 14-26 Carbanion intermediates stabilized by covalent interactions with transketolase and transaldolase, (a) The ring of TPP stabilizes the two-carbon carbanion carried by transketolase see Fig. 14-13 for the chemistry of TPP action, (b) In the transaldolase reaction, the protonated Schiff base formed between the e-amino group of a Lys side chain and the substrate stabilizes a three-carbon carbanion. 

Stromal enzymes, including transketolase and transaldolase, rearrange the carbon skeletons of triose phosphates, generating intermediates of three, four, five, six, and seven carbons and eventually yielding pentose phosphates. 

The ingenious sugar rearrangement system uses two enzymes, transketolase and transaldolase. 

Transfer of C2 and C3 units in reactions catalysed by transketolase and transaldolase respectively modify the chain length of the sugar. 

The transketolase and transaldolase reactions are reversible and so allow either the conversion of ribose 5-phosphate into glycolytic intermediates when it is not needed for other cellular reactions, or the generation of ribose 5-phosphate from glycolytic intermediates when more is required. The rate of the pentose phosphate pathway is controlled by NADP+ regulation of the first step, catalyzed by glucose 6-phosphate dehydrogenase. 

If at any time only a little ribose 5-phosphate is required for nucleic acid synthesis and other synthetic reactions, it will tend to accumulate and is then converted to fructose 6-phosphate and glyceraldehyde 3-phosphate by the enzymes transketolase and transaldolase. These two products are intermediates of glycolysis. Therefore, these reactions provide a link between the pentose phosphate pathway and glycolysis. The outline reactions are shown below. 

NADPH, fructose 6-phosphate and glyceraldehyde 3-phosphate can be taken from glycolysis and converted into ribose 5-phosphate by reversal of the transketolase and transaldolase reactions. 

Because both transketolase and fructose-1,6-bisphosphate (FBP) aldolase provide ketoses of D-threo configuration on carbons 3 and 4, they are... 

Recent Syntheses Involving Transketolase and Fructose-1,6-hrsphosphate Aldolase... 

Transketolase and FruA can yield the same ketose if the aldehyde acceptor of FruA (especially an a-hydroxylated aldehyde as shown in Figure 18.1) has one carbon less than the aldehyde acceptor used for transketolase. Hence for a particular synthesis, the choice of the enzyme will depend strongly on the availability of the acceptor substrate. 

Recent Syntheses Involving Transketolase and Fnictose-1,6-bisphosphate Aldolase 29J... 

In the first step, we showed by analytical studies that compound 28 was a donor substrate for transketolase in the presence of D-ribose-5-phosphate as acceptor substrate and that in the second step, the hydroxylated aldehyde released 29 led to the P-elimination of protected L-tyrosine. We showed that the free L-tyrosine can thus be obtained by enzymatic deprotection of N-acetyl-L-tyrosine ethyl ester using acylase and subtilisine. In this conditions, it should be possible to carry out this assay in vivo in the presence of host cells overexpressing transketolase and auxotrophic for L-tyrosin. This strategy should offer the first stereospecific selection test of transketolase mutants. The principle of this assay may be extended to other enzymes that can release aldehydes P-substituted by L-tyrosine. 

Pekovich SR, Martin PR and Singleton CK (1998) Thiamine deficiency decreases steady-state transketolase and pyruvate dehydrogenase but not alpha-ketoglutarate dehydrogenase mRNA levels in three human cell types. Journal of Nutrition 128, 683-7. 

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