Differential selectivity of the Escherichia coli cell membrane shifts the equilibrium for the enzyme-catalyzed isomerization of galactose to tagatose.
An Escherichia coli galactose kinase gene knockout (DeltagalK) 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 DeltagalK strain, the isomerase-catalyzed reaction exhibited an equilibrium shift toward tagatose, producing a tagatose fraction of 68% at 37 degrees 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 DeltamglB 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.
Kim
Jin-Ha
JH
Department of Bioscience and Biotechnology, Konkuk University, 1 Hwayang-dong, Gwangjin-gu, Seoul 143-701, South Korea. deokkun@konkuk.ac.kr
Lim
Byung-Chul
BC
Yeom
Soo-Jin
SJ
Kim
Yeong-Su
YS
Kim
Hye-Jung
HJ
Lee
Jung-Kul
JK
Lee
Sook-Hee
SH
Kim
Seon-Won
SW
Oh
Deok-Kun
DK
D019747:Aldose-Ketose Isomerases / Q000302:isolation & purification / Q000378:metabolism*; D002462:Cell Membrane / Q000378:metabolism*; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics / Q000254:growth & development / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000235:genetics / Q000378:metabolism*; D005686:Galactokinase / Q000235:genetics; D005690:Galactose / Q000378:metabolism*; D017353:Gene Deletion; D006601:Hexoses / Q000378:metabolism*; D007700:Kinetics
D029968:Escherichia coli Proteins; D006601:Hexoses; D005686:Galactokinase; D019747:Aldose-Ketose Isomerases; C019715:L-arabinose isomerase; C030192:tagatose; D005690:Galactose