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Differential Selectivity of the Escherichia coli Cell Membrane Shifts the Equilibrium for the Enzyme-Catalyzed Isomerization of Galactose to Tagatose

Departments of Bioscience and Biotechnology,¹ Chemical Engineering, Konkuk University, Seoul 143-701,² Division of Applied Life Science, EB-NCRC and PMBBRC, Gyeongsang National University, Jinju 660-701, South Korea³

Received 28 November 2007/ Accepted 1 February 2008

An Escherichia coli galactose kinase gene knockout (galK) strain, which contains the L-arabinose isomerase gene (araA) to isomerize D-galactose to D-tagatose, showed a high conversion yield of tagatose compared with the original galK strain because galactose was not metabolized by endogenous galactose kinase. In whole cells of the galK strain, the isomerase-catalyzed reaction exhibited an equilibrium shift toward tagatose, producing a tagatose fraction of 68% at 37°C, whereas the purified L-arabinose isomerase gave a tagatose equilibrium fraction of 36%. These equilibrium fractions are close to those predicted from the measured equilibrium constants of the isomerization reaction catalyzed in whole cells and by the purified enzyme. The equilibrium shift in these cells resulted from the higher uptake and lower release rates for galactose, which is a common sugar substrate, than for tagatose, which is a rare sugar product. A mglB mutant had decreased uptake rates for galactose and tagatose, indicating that a methylgalactoside transport system, MglABC, is the primary contributing transporter for the sugars. In the present study, whole-cell conversion using differential selectivity of the cell membrane was proposed as a method for shifting the equilibrium in sugar isomerization reactions.

Published ahead of print on 8 February 2008.