Transketolase, synthesis

A number of mechanistically distinct enzymes can likewise be employed for the synthesis of product structures identical to those accessible from aldolase catalysis. Such alternative cofactor-dependent enzymes (e.g. transketolase) are emerging as useful catalysts in organic synthesis. As these operations often extend and/or... 

Flgure10.23 Sialyl Lewis -related selectin inhibitorandfluorogenicscreening compound for transketolase prepared using enzymatic aldolization, and multienzymatic oxidation-aldolization strategy for the synthesis of bicyclic higher carbon sugars. 

Figure 10.39 Synthesis of a novel N-hydroxypyrrolidine and a fluorogenic screening substrate for transaldolases based on stereospecific transketolase catalysis.

Ribulose 5-phosphate is the substrate for two enzymes. Ribulose 5-phosphate 3-epimerase alters the configuration about carbon 3, forming another ketopentose, xylulose 5-phosphate. Ribose 5-phosphate ketoisom-erase converts ribulose 5-phosphate to the corresponding aldopentose, ribose 5-phosphate, which is the precursor of the ribose required for nucleotide and nucleic acid synthesis. Transketolase transfers the two-carbon... 

Sprenger, G.A. et al.. Identification of a thiamin-dependent synthase in Escherichia coli required for the formation of the 1-deoxy-D-xylulose 5-phosphate precursor to isoprenoids, thiamin, and pyridoxol, Proc. Natl. Acad Sci. USA 94, 12857, 1997. Lange, B.M. et al., A family of transketolases that directs isoprenoid biosynthesis via a mevalonate-independent pathway, Proc. Natl. Acad Sci. USA 95, 2100, 1998. Lois, L.M. et al., Cloning and characterization of a gene from Escherichia coli encoding a transketolase-like enzyme that catalyzes the synthesis of D-1- deoxyxylulose 5-phosphate, a common precursor for isoprenoid, thiamin, and pyridoxol biosynthesis, Proc. Natl. Acad. Sci. USA 95, 2105, 1998. 

Transketolase (TKase) [EC 2.2.1.1] essentially catalyzes the transfer of C-2 unit from D-xylulose-5-phosphate to ribose-5-phosphate to give D-sedoheptulose-7-phosphate, via a thiazolium intermediate as shown in Fig. 16. An important discovery was that hydroxypyruvate works as the donor substrate and the reaction proceeds irreversibly via a loss of carbon dioxide (Fig. 17). In this chapter, we put emphasis on the synthesis with hydroxypyruvate, as it is the typical TPP-mediated decarboxylation reaction of a-keto acid. ... 

The hypE proteins are 302-376 residues long and appear to consist of three domains. Domain 1 shows sequence identity to a domain from phosphoribosyl-aminoimida-zole synthetase which is involved in the fifth step in de novo purine biosynthesis and to a domain in thiamine phosphate kinase which is involved in the synthesis of the cofactor thiamine diphosphate (TDP). TDP is required by enzymes which cleave the bond adjacent to carbonyl groups, e.g. phosphoketolase, transketolase or pyruvate decarboxylase. Domain 2 also shows identity to a domain found in thiamine phosphate kinase. Domain 3 appears to be unique to the HypF proteins. 

Although the purified enzyme is much less active than transketolase (3 U mg versus 80-100 U mg ), it could be utilized as a tool in a one-pot multi-enzyme asymmetric synthesis yielding about 700 mg of the barium salt of DXP [21]. 

The ready availability of the transketolase (TK E.C. 2.2.1.1) from E. coli within the research collaboration in G. A. Sprenger s group suggested the joint development of an improved synthesis of D-xylulose 5-phosphate 19, which was expensive but required routinely for activity measurements [27]. In vivo, transketolase catalyzes the stereospecific transfer of a hydroxyacetyl nucleophile between various sugar phosphates in the presence of a thiamine diphosphate cofactor and divalent cations, and the C2 donor component 19 offers superior kinetic constants. For synthetic purposes, the enzyme is generally attractive for its high asymmetric induction at the newly formed chiral center and high kinetic enantioselectivity for 2-hydroxyaldehydes, as well as its broad substrate tolerance for aldehyde acceptors [28]. 

Scheme 2.2.5.S Multi-enzymatic route for the stereoselective synthesis of D-xylulose 5-phosphate 19 from commercial D-fructose 1,6-bisphosphate (abbreviations TPI, triose phosphate isomerase TK, transketolase).

An example of an a-ketol formation that does not involve decarboxylation is provided by the reaction catalyzed by transketolase, an enzyme that plays an essential role in the pentose phosphate pathway and in photosynthesis (equation 21) (B-77MI11001). The mechanism of the reaction of equation (21) is similar to that of acetolactate synthesis (equation 20). The addition of (39) to the carbonyl group of (44) is followed by aldol cleavage to give a TPP-stabilized carbanion (analogous to (41)). The condensation of this carbanionic intermediate with the second substrate, followed by the elimination of (39), accounts for the observed products (B-7IMIHOO1). 

A reaction that is related to that of transketolase but is likely to function via acetyl-TDP is phosphoketolase, whose action is required in the energy metabolism of some bacteria (Eq. 14-23). A product of phosphoketolase is acetyl phosphate, whose cleavage can be coupled to synthesis of ATP. Phosphoketolase presumably catalyzes an a cleavage to the thiamin-containing enamine shown in Fig. 14-3. A possible mechanism of formation of acetyl phosphate is elimination of HzO from this enamine, tautomerization to 2-acetylthiamin, and reaction of the latter with inorganic phosphate. 

Transketolase 733,736 Translation of genetic information, 5. See also Protein synthesis definition of 5 nick 257 regulation of 536 Translocation... 

Transketolase reactions leading via the pentose or hexose monophosphate shunt pathway of glucose oxidation to the eventual production of pentoses for RNA/DNA synthesis and NADPH for the biosynthesis of fatty acids... 

Mostly the baker s yeast transketolase has been used so far to prepare several valuable ketose sugars and derivatives 141 (cf. Sect. 7). Recently, the transketolase was utilized in the key stereogenic transformation of racemic 2-hydroxybutyraldehyde 142 into the homochiral synthon 5,6-dideoxy-D-t/ireo-hexulose 110 for the chemoenzymatic synthesis of (+ )-exo-brevicomin 107 [314], Transketolase has also been applied for the in-situ generation of Ery4P (35) from Fru6P (38) in a multi-enzymatic synthesis of 34 (Scheme 5). 

Transketolases (TK) aie widely used in I >ry m ic synthesis to extend the chain of an acceptor aldose by two carbon units. 

Enantiopure, bifunctional acyloins (a-hydroxy ketones) are versatile intermediates in natural product synthesis (also see Sect. 2.3, Fig. 11). In nature, the formation of a-hydroxy ketones is efficiently catalyzed by thiamine diphosphate-dependent enzymes transketolases, decarboxylases, and other lyases, such as BALs. A great portfolio of biotransformations, especially with benzaldehyde derivatives as starting materials, were realized [204]. 

LM Lois, N Campos, SR Putra, K Danielsen, M Rohmer, A Boronat. Cloning and characterization of a gene from Escherichia coli encoding a transketolase-like enzyme that catalyzes the synthesis of D-l-deoxyxylulose 5-phosphate, a common precursor for isoprenoid, thiamin, and pyridoxol biosynthesis. Proc Natl Acad Sci (USA) 95 2105-2110, 1998. 

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. 

The sugar metabolism is a source of many enzymes, the transketolase (TK) being one of them. TK transfers an a-hydroxy carbonyl fragment from D-xylu-lose-5-phosphate onto D-ribose-5-phosphate, forming D-sedoheptulose-7-phos-phate and D-glyceraldehyde-3-phosphate (Scheme 5.14). Since this reaction is an equilibrium reaction and starting materials and products are of similar stability, it is not very versatile for organic synthesis. Fortunately TK also accepts pyruvate instead of xylulose. Under these modified circumstances carbon dioxide... 

Scheme 5.40. Preparative scale production of DAHP by multi-enzyme synthesis. Ei = hexoki-nase, E2 = pyruvate kinase, E3 = transketolase + D-ribose 5-P, E4 = DHAP synthetase. P, = inorganic phosphate, P = PC>32. ...

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