Dataset Information

Hydrolysis of Oligosaccharides by a Thermostable ?-Galactosidase from Termitomyces eurrhizus.

ABSTRACT: The genus of Termitomyces purchased from the market has been identified as Termitomyces eurrhizus using the Internal Transcribed Spacer (ITS) method. An ?-galactosidase from T. eurrhizus (TEG), a monomeric protein with a molecular mass of 72 kDa, was purified 146 fold by employing ion exchange chromatography and gel filtration. The optimum pH and temperature was 5.0 and 60 °C, respectively. TEG was stable over pH 2-6, and also exhibited good thermostablility, retaining 100% of the original activity after incubation at 60 °C for 2 h. Inhibition of the enzyme activity by N-bromosuccinimide (NBS) constituted evidence for an essential role of tryptophan in the catalytic action of the isolated enzyme. Besides 4-nitro-phenyl ?-d-galactophyranoside (pNPGal), natural substrates could also be effectively hydrolyzed by TEG. Results of thin-layer chromatography (TLC) revealed complete enzymatic hydrolysis of raffinose and stachyose to galactose at 50 °C within 6 h. These properties of TEG advocate its utilization for elevating the nutritional value of soymilk.

SUBMITTER: Zhang W

PROVIDER: S-EPMC4691104 | biostudies-literature | 2015

REPOSITORIES: biostudies-literature

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Hydrolysis of Oligosaccharides by a Thermostable α-Galactosidase from Termitomyces eurrhizus.

Zhang Weiwei W Du Fang F Wang Li L Zhao Liyan L Wang Hexiang H Ng Tzi Bun TB

International journal of molecular sciences 20151208 12

The genus of Termitomyces purchased from the market has been identified as Termitomyces eurrhizus using the Internal Transcribed Spacer (ITS) method. An α-galactosidase from T. eurrhizus (TEG), a monomeric protein with a molecular mass of 72 kDa, was purified 146 fold by employing ion exchange chromatography and gel filtration. The optimum pH and temperature was 5.0 and 60 °C, respectively. TEG was stable over pH 2-6, and also exhibited good thermostablility, retaining 100% of the original activ ...[more]

PMID: 26670230

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Project description:BACKGROUND:?-Galactosidases are enzymes that act on galactosides present in many vegetables, mainly legumes and cereals, have growing importance with respect to our diet. For this reason, the use of their catalytic activity is of great interest in numerous biotechnological applications, especially those in the food industry directed to the degradation of oligosaccharides derived from raffinose. The aim of this work has been to optimize the recombinant production and further characterization of ?-galactosidase of Saccharomyces cerevisiae. RESULTS:The MEL1 gene coding for the ?-galactosidase of S. cerevisiae (ScAGal) was cloned and expressed in the S. cerevisiae strain BJ3505. Different constructions were designed to obtain the degree of purification necessary for enzymatic characterization and to improve the productive process of the enzyme. ScAGal has greater specificity for the synthetic substrate p-nitrophenyl-?-D-galactopyranoside than for natural substrates, followed by the natural glycosides, melibiose, raffinose and stachyose; it only acts on locust bean gum after prior treatment with ?-mannosidase. Furthermore, this enzyme strongly resists proteases, and shows remarkable activation in their presence. Hydrolysis of galactose bonds linked to terminal non-reducing mannose residues of synthetic galactomannan-oligosaccharides confirms that ScAGal belongs to the first group of ?-galactosidases, according to substrate specificity. Optimization of culture conditions by the statistical model of Response Surface helped to improve the productivity by up to tenfold when the concentration of the carbon source and the aeration of the culture medium was increased, and up to 20 times to extend the cultivation time to 216 h. CONCLUSIONS:ScAGal characteristics and improvement in productivity that have been achieved contribute in making ScAGal a good candidate for application in the elimination of raffinose family oligosaccharides found in many products of the food industry.

Dataset Information

Hydrolysis of Oligosaccharides by a Thermostable ?-Galactosidase from Termitomyces eurrhizus.

Publications

Hydrolysis of Oligosaccharides by a Thermostable α-Galactosidase from Termitomyces eurrhizus.

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