High control coefficient of transketolase in the nonoxidative pentose phosphate pathway of human erythrocytes: NMR, antibody, and computer simulation studies

Abstract

The degree of control exerted by transketolase over metabolite flux in the nonoxidative pentose phosphate pathway in human erythrocytes was investigated using transketolase antiserum to modulate the activity of that enzyme. 31P NMR enabled the simultaneous measurement of the levels of pentose phosphate pathway metabolites following incubation of hemolysates with ribose 5-phosphate. The variations in metabolic flux which occurred as the transketolase activity of hemolysate samples was altered indicated that a high degree of control was exerted by transketolase. Investigations using transaldolase-depleted hemolysates showed that transaldolase exhibits a lesser degree of control over pathway flux. Experimental data were compared with simulations generated by a computer model encompassing the reactions of the classical nonoxidative pentose phosphate pathway. The sensitivity coefficients (also called "control strengths" or "flux-control coefficients") calculated from the computer simulations were 0.74 and 0.03 for transketolase and transaldolase, respectively.

Orchestration of Thiamin Biosynthesis and Central Metabolism by Combined Action of the Thiamin Pyrophosphate Riboswitch and the Circadian Clock in Arabidopsis

“…Since the thiamin-requiring enzymes are also key control steps of the TCA cycle and the PPP (Berthon et al, 1992;Wang et al, 1997aWang et al, , 1997bHenkes et al, 2001;Araújo et al, 2008), we determined the relative Glc oxidation rate on the basis of 14 CO 2 evolution of plants harboring a defective riboswitch. It was observed that riboswitch deficiency and the consequent activation of thiamin biosynthesis also caused the augmentation of the metabolic fluxes through the TCA cycle and the PPP.…”

“…It is an essential coenzyme for the enzyme complexes pyruvate dehydrogenase (PDH) and 2-oxoglutarate dehydrogenase (2-OGDH) of the tricarboxylic acid (TCA) cycle and for the a-ketose transketolase (TK) homodimer of the pentose phosphate pathway (PPP; Frank et al, 2007). Intriguingly, these enzyme complexes appear to be the controlling steps of the pathways they are involved in (Berthon et al, 1992;Wang et al, 1997aWang et al, , 1997bHenkes et al, 2001;Araújo et al, 2008), and TPP is additionally limiting for their assembly (Singleton and Martin, 2001;Tovar-Méndez et al, 2003;Bunik and Fernie, 2009). Altogether, the above facts emphasize the importance of TPP for the proper functioning of the TCA cycle and the PPP and consequently for general energy production in all cells.…”

The H2O2 Stimulon in Saccharomyces cerevisiae

“…Then, pentose phosphates are interconverted to glyceraldehyde 3-phosphate or fructose 6-phosphate by transketolases (Tkl1p and Tkl2p) and transaldolase (Tal1p). Glyceraldehyde 3-phosphate can enter glycolysis and fructose 6-phosphate is converted to glucose 6-phosphate (49). However, repression of glycolysis (see below) and purine biosynthesis pathway suggest that most of the pentose phosphates are recycled to the hexose phosphate pool for NADPH production.…”

Properties and functions of the thiamin diphosphate dependent enzyme transketolase

“…It has been suggested for S. cerevisiae that TK and TA control the¯ux of metabolites through the non-oxidative limb of the PPP [133,144]. NMR studies on human hemolysates have demonstrated that TK has a high¯ux-control coecient, indicating that this enzyme exhibits a controlling function on the PPP in erythrocytes [5]. Although this pathway is ubiquitous there is evidence that its metabolic activity in higher organisms is tissue-dependent, e.g.…”