Abstract
Initial rate and product inhibition studies using pH-stat unbuffered biotransformation mixtures at 25°C and pH 7.0 have been performed on overexpressed transketolase (EC 2.2.1.1.) from a self-cloned transformant of Escherichia coli. The Michaelis and inhibition constants for the cosubstrates, hydroxypyruvate and glycolaldehyde, and the inhibition constant of the l-erythrulose product were determined. The kinetic data are consistent with a Ping Pong Bi Bi mechanism where the ketol donor (hydroxypyruvate) is first bound to the enzyme and is followed by release of the first product CO 2, followed by the binding of the aldehyde acceptor (glycolaldehyde) and subsequent release of the second product, l-erythrulose. The enzyme was shown to be free from excess substrate inhibition up to 40 mmol l −1 for hydroxypyruvate and 100 mmol l −1 for glycolaldehyde. In addition, inhibition by the l-erythrulose product in conformity with the predicted mechanism was found to be competitive.
Published Version
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