Transketolase catalyzes the transfer of a ketol from a ketose (xylulose 5-phosphate, fructose 6-phosphate or sedoheptulose 7-phosphate) to an aldose (ribose 5-phosphate, erythrose 4-phosphate or glyceraldehyde 3-phosphate) and is a key enzyme in the pentose phosphate shunt (4, 5). Transketolase, together with transaldolase, creates a reversible link between two main metabolic pathways, the pentose phosphate pathway and glycolysis, which allows the cell to adapt its NADPH production, and ribose-5-phosphate production to meet its immediate needs (4). In Saccharomyces cerevisiae, TKL1 encodes the major isoform and TKL2 encodes a minor isoform (6). Double null mutants for tkl1 and tkl2 are viable, but auxotrophic for aromatic amino acids (6).

Transketolase from S. cerevisiae, is a homodimer, and is dependent on thiamine diphosphate as a cofactor and on divalent cations for activity (1, 4, 7, 8, 9, 10). Each subunit is folded into three consecutive alpha/beta-domains (4). The coenzyme-binding site is located in a deep cleft at the interface between the subunits, and residues of both subunits interact with the thiamine diphosphate cofactor (4, 11, 12, 13) to stabilize the holoenyme. Binding of Ca2+, or to a lesser extent of Mg2+, also stabilize the holoenzyme (13). In vivo, transketolase activity may be negatively regulated by RNA (14). Each is also capable of nonspecifically catalyzing the reactions in the non-oxidative branch of the pentose phosphate pathway (5).

The Tkl1p enzyme is well characterized. Specific amino acids have shown to be important for specific activities, such as stability of the Tkl1p transketolase holoenzyme (15), coenzyme binding (10, 15), substrate specificity and enzymatic activity (10, 11, 16, 17, 18). In addition to catalyzing the common two-substrate transketolase reaction, Tkl1p has also specifically been shown to catalyze a one-substrate reaction utilizing only xylulose 5-phosphate to produce glyceraldehyde 3-phosphate and erythrulose (19).

Null tkl1 mutants are viable and display normal growth on rich media, but display different growth defects on minimal media depending on the strain background (1, 6, 11, 15, 16, 20). Tkl1p is required for efficient use of fermentable carbon sources and for the biosynthesis of aromatic amino acids (1). Tkl1p is also indirectly involved in the response to reactive oxygen species (ROS) through its involvement in determining intracellular NADPH levels (21). Overexpression of TKL1 reduces growth on fermentable carbon sources, such as glucose and raffinose, and on gluconeogenic carbon sources, such as pyruvate, ethanol, and glycerol. This is similar to the phenotype of strains disrupted for TKL1 and suggests that a proper balance between glycolysis and the pentose phosphate pathway is important for efficient use of fermentable carbon sources (1).

Tkl1p is of industrial interest for the fermentation of xylose to ethanol (2). Xylose is the predominant sugar found in biomass such as agricultural wastes, wood, municipal solid wastes, and wastes from pulp and paper industries, and possibly could serve as a low-cost and abundant raw material for fuel ethanol production (22). Tkl1p expression is increased in mutant strains that display increased fitness during growth on xylose relative to the parental strain, which was engineered to utilize xylose (23, 24). These and other data suggest that TKL1 may be a good target for improving xylose fermentation in S. cerevisiae (2, 22, 23, 24, 25).

Tkl1p has similarity to S. cerevisiae Tkl2p, Escherichia coli transketolase, Rhodobacter sphaeroides transketolase, Streptococcus pneumoniae recP, Hansenula polymorpha dihydroxyacetone synthase, Kluyveromyces lactis TKL1 (which complements the phenotype of the tkl1 tkl2 double null mutant, (26)), Pichia stipitis TKT, rabbit liver transketolase, rat TKT, mouse TKT, and human TKT (1, 6, 9, 26). Tkl1p is also related to E. coli pyruvate dehydrogenase E1 subunit, which is another vitamin B1-dependent enzyme (1).

Last updated: 2005-12-19