Preparation of lactose-free pasteurized milk with a recombinant thermostable β-glucosidase from Pyrococcus furiosusReport as inadecuate




Preparation of lactose-free pasteurized milk with a recombinant thermostable β-glucosidase from Pyrococcus furiosus - Download this document for free, or read online. Document in PDF available to download.

BMC Biotechnology

, 13:73

Protein and enzyme technology

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 Ca. 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.

KeywordsRecombinant β-glucosidase Thermostable enzyme Pichia pastoris Milk lactose hydrolysis Pasteurization AbbreviationsP. furiosusPyrococcus furiosus

E. coliEscherichia coli

S. cerevisiaeSaccharomyces cerevisiae

PCRPolymerase chain reaction

SDS-PAGESodium dodecyl sulfate polyacrylamide gel electrophoresis

YPDYeast extract peptone dextrose

DODissolved oxygen

DEAEDiethylaminoethyl cellulose

pNPp-nitrophenol

pNPGp-nitrophenol-β-D-glucopyranoside

HPLCHigh-Performance Liquid Chromatography.

Electronic supplementary materialThe online version of this article doi:10.1186-1472-6750-13-73 contains supplementary material, which is available to authorized users.

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Author: Bin Li - Zemin Wang - Shiwu Li - William Donelan - Xingli Wang - Taixing Cui - Dongqi Tang

Source: https://link.springer.com/







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