Abstract
BackgroundLactose intolerance is a common health concern causing gastrointestinal symptoms and avoidance of dairy products by afflicted individuals. Since milk is a primary source of calcium and vitamin D, lactose intolerant individuals often obtain insufficient amounts of these nutrients which may lead to adverse health outcomes. Production of lactose-free milk can provide a solution to this problem, although it requires use of lactase from microbial sources and increases potential for contamination. Use of thermostable lactase enzymes can overcome this issue by functioning under pasteurization conditions.ResultsA thermostable β-glucosidase gene from Pyrococcus furiosus was cloned in frame with the Saccharomyces cerecisiae a-factor secretory signal and expressed in Pichia pastoris strain X-33. The recombinant enzyme was purified by a one-step method of weak anion exchange chromatography. The optimum temperature and pH for this β-glucosidase activity was 100°C and pH 6.0, respectively. The enzyme activity was not significantly inhibited by Ca2+. We tested the additive amount, hydrolysis time, and the influence of glucose on the enzyme during pasteurization and found that the enzyme possessed a high level of lactose hydrolysis in milk that was not obviously influenced by glucose.ConclusionsThe thermostablity of this recombinant β-glucosidase, combined with its neutral pH activity and favorable temperature activity optima, suggest that this enzyme is an ideal candidate for the hydrolysis of lactose in milk, and it would be suitable for application in low-lactose milk production during pasteurization.
Highlights
Lactose intolerance is a common health concern causing gastrointestinal symptoms and avoidance of dairy products by afflicted individuals
In vivo, lactase (β-D-galactosidase) is an enzyme secreted by intestinal villi that hydrolyses the disaccharide lactose into glucose and galactose [1,2] and is essential for the digestion of bovine milk [3] which contains an average of 4.8% lactose
Since lactose is a disaccharide composed of glucose and galactose, it can be hydrolyzed into these monosaccharides using either a β-glucosidase or β-galactosidase
Summary
Lactose intolerance is a common health concern causing gastrointestinal symptoms and avoidance of dairy products by afflicted individuals. Lactase (β-D-galactosidase) is an enzyme secreted by intestinal villi that hydrolyses the disaccharide lactose into glucose and galactose [1,2] and is essential for the digestion of bovine milk [3] which contains an average of 4.8% lactose. This enzyme has been cloned and functionally characterized in Saccharomyces cerevisiae, expression in batch culture produced low yields (~10 mg/L) which are not suitable for industrial applications [13] Still, this β-glucosidase serves as a model system to use for lactose hydrolysis since it displays extreme stability and high catalytic activity in the presence of lactose
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