Alcoholysis and reverse hydrolysis reactions in organic one-phase system with a hyperthermophilic beta-glycosidase

Alcoholysis and reverse hydrolysis reactions were performed enzymatically i n one-phase water-saturated 1-heptanol systems. Lactose or glucose was used as substrate to produce heptyl-beta-galactoside and/or heptyl-beta-glucosi de, respectively. When alcoholysis of lactose was performed at 37 degrees C with beta-galactosidase from Escherichia coli, the initial rate was 14 nmo l/mL min, and the limiting factors were the poor solubility of the substrat e in 1-heptanol and low thermal stability of the enzyme. When a hyperthermo philic beta-glycosidase was used at 90 degrees C, the rate was 3.14-fold hi gher; in this case a higher concentration of soluble lactose in the water-s aturated heptanol was available to the enzyme due to the higher temperature . The hyperthermophilic beta-glycosidase was also able to use glucose and g alactose as substrates to achieve the reverse hydrolysis reaction. As a con sequence, when lactose was used as substrate, heptyl-beta-galactoside was f ormed by alcoholysis, while the released glucose moiety was used in a secon dary reverse hydrolysis reaction to produce heptyl-beta-glucoside. Both rea ctions followed Michaelis-Menten kinetics behavior. Neither lactose nor hep tyl glycosides were hydrolyzed by this enzyme in water-saturated heptanol. However, the conversion was limited by a strong product inhibition and the formation of oligosaccharides, especially at high substrate concentrations, reducing the final glycoside yield. (C) 2000 John Wiley & Sons, Inc.