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  2. Abstract

When human erythrocyte apo-transketolase is mixed with cofactors and substrates, the progress curve exhibits a lag phase. Elimination of the lag phase requires the presence of saturating concentrations of cofactors, thiamin diphosphate and Mg2+. The most simple explanation of the observed hysteretic transition is that the slow binding of a Mg2+-thiamin-diphosphate species precedes slow isomerisation of the enzyme to the active form. Although the hysteretic transition involves more than one process, it does not involve the dissociation–association of enzyme subunits. The best estimate of the apparent Km, 1.59 ± 0.23 μM, for the binding of Mg2+-thiamin diphosphate to transketolase was obtained in the presence of a high non-inhibitory concentration of magnesium and varied concentrations of thiamin diphosphate. Thus the reconstitution of the human enzyme differs from the yeast enzyme, which undergoes a rate-limiting dimerisation during reconstitution.


thiamin diphosphate


International unit (IU) 1 IU is the amount of enzyme that produces 1 μmol product/min under optimal conditions


Human transketolase (EC


triose-phosphate isomerase (EC


glycerol-3-phosphate dehydrogenase (EC


  1. Top of page
  2. Abstract
  • 1
    Datta, A. & Racker, E. (1961) Mechanisms of action of transketolase. I. Properties of the crystalline yeast enzyme, J. Biol. Chem. 236, 617623.
  • 2
    Tomita, I., Saitou, S. & Ishikawa, M. (1979) Purification and properties of transketolase from pig liver. I. An attempt to resolve the enzyme into apoenzyme and cofactors, J. Nutr. Sci. Vitaminol. 25, 175184.
  • 3
    Tate, J., Kaczmarek, M. & Nixon, P. (1984) Stimulation of erythrocyte transketolase by added thiamin diphosphate is pH dependent, Clin. Chim. Acta 137, 8186.
  • 4
    Jung, E., Takeuchi, T., Nishino, K. & Itokawa, Y. (1988) Studies on the nature of thiamin pyrophosphate binding and dependency on divalent cations of transketolase from human erythrocytes, Int. J. Biochem. 20, 12551259.
  • 5
    Heinrich, P., Steffen, H., Janser, P. & Wiss, O. (1972) Studies on the reconstitution of apotransketolase with thiamin pyrophosphate and analogues of the co-enzyme, Eur. J. Biochem. 30, 533541.
  • 6
    Egan, R. & Sable, H. (1981) Transketolase kinetics. The slow reconstitution of the holoenzyme is due to rate-limiting dimerization of the subunits, J. Biol. Chem. 256, 48774883.
  • 7
    Blass, J., Piacentini, S., Boldizsar, E. & Baker, A. (1982) Kinetic studies of mouse brain transketolase, J. Neurochem. 39, 729733.
  • 8
    Smeets, E., Muller, H. & De Wael, J. (1971) A NADH-dependent transketolase assay in erythrocyte haemolysates, Clin. Chim. Acta 33, 379386.
  • 9
    Tate, J. & Nixon, P. (1987) Measurement of the Michaelis constant for human erythrocyte transketolase and thiamin diphosphate, Anal. Biochem. 160, 7887.
  • 10
    Duggleby, R. (1984) A program for non-linear least squares regression analysis, Comput. Biol. Med. 14, 447452.
  • 11
    Neet, K. & Ainsley, G. (1980) Hysteretic enzymes, Methods Enzymol. 64, 192226.
  • 12
    Cavalieri, S., Neet, K. & Sable, H. (1975) Enzymes of pentose biosynthesis. The quaternary structure and reacting form of transketolase from bakers yeast, Arch. Biochem. Biophys. 171, 527532.
  • 13
    Booth, C., Nixon, P. & Winzor, D. (1992) Gel chromatographic evaluation of the binding constant for the interaction of thiamin diphosphate with magnesium ion, J. Chromatogr. 609, 8387.
  • 14
    Blass, J. & Gibson, G. (1977) Abnormality of a thiamin requiring enzyme in patients with Wernicke-Korsakoff syndrome, N. Engl. J. Med. 297, 13671370.
  • 15
    Warnock, L. & Prudhomme, C. (1982) The isolation and preliminary characterization of apotransketolase from human erythrocytes, Biochem. Biophys. Res. Commun. 106, 719723.
  • 16
    Nixon, P., Kaczmarek, M., Tate, J., Kerr, R. & Price, J. (1984) An erythrocyte transketolase isoenzyme pattern associated with the Wernicke-Korsakoff syndrome, Eur. J. Clin. Invest. 14, 278281.
  • 17
    Grudzinski, A., Waltham, M., Ioannoni, B., Price, J. & Nixon, P. (1989) Interactions with haemoglobin: a source of error in measurements of transketolase activity in haemolysates, Clin. Chim. Acta 180, 265276.