Development of an ultra-high-temperature process for the enzymatic hydrolysis of lactose: II. Oligosaccharide formation by two thermostable β-glycosidases

✍ Scribed by Inge Petzelbauer; Reinhard Zeleny; Andreas Reiter; Klaus D. Kulbe; Bernd Nidetzky

• Publisher: John Wiley and Sons
• Year: 2000
• Tongue: English
• Weight: 188 KB
• Volume: 69
• Category: Article
• ISSN: 0006-3592
• DOI: 10.1002/(sici)1097-0290(20000720)69:2<140::aid-bit3>3.0.co;2-r

✦ Synopsis

During lactose conversion at 70°C, when catalyzed by ␤-glycosidases from the archea Sulfolobus solfataricus (Ss␤Gly) and Pyrococcus furiosus (CelB), galactosyl transfer to acceptors other than water competes efficiently with complete hydrolysis of substrate. This process leads to transient formation of a range of new products, mainly disaccharides and trisaccharides, and shows a marked dependence on initial substrate concentration and lactose conversion. Oligosaccharides have been analyzed quantitatively by using capillary electrophoresis and high performance anion-exchange chromatography. At 270 g/L initial lactose, they accumulate at a maximum concentration of 86 g/L at 80% lactose conversion. With both enzymes, the molar ratio of trisaccharides to disaccharides is maximal at an early stage of reaction and decreases directly proportional to increasing substrate conversion. Overall, CelB produces about 6% more hydrolysis byproducts than Ss␤Gly. However, the product spectrum of Ss␤Gly is richer in trisaccharides, and this agrees with results obtained from the steadystate kinetics analyses of galactosyl transfer catalyzed by Ss␤Gly and CelB. The major transgalactosylation products of Ss␤Gly and CelB have been identified. They are ␤-D-Galp-(1→3)-Glc and ␤-D-Galp-(1→6)-Glc, and ␤-D-Galp-(1→3)-lactose and ␤-D-Galp-(1→6)-lactose, and their formation and degradation have been shown to be dependent upon lactose conversion. Both enzymes accumulate ␤(1→6)-linked glycosides, particularly allolactose, at a late stage of reaction. Because a high oligosaccharide concentration prevails until about 80% lactose conversion, thermostable ␤-glycosidases are efficient for oligosaccharide production from lactose. Therefore, they prove to be stable and versatile catalysts for lactose utilization.