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A Novel Highly Thermostable Multifunctional Beta-Glycosidase from Crenarchaeon Acidilobus saccharovorans.

ABSTRACT: We expressed a putative ?-galactosidase Asac_1390 from hyperthermophilic crenarchaeon Acidilobus saccharovorans in Escherichia coli and purified the recombinant enzyme. Asac_1390 is composed of 490 amino acid residues and showed high sequence similarity to family 1 glycoside hydrolases from various thermophilic Crenarchaeota. The maximum activity was observed at pH 6.0 and 93°C. The half-life of the enzyme at 90°C was about 7 hours. Asac_1390 displayed high tolerance to glucose and exhibits hydrolytic activity towards cellobiose and various aryl glucosides. The hydrolytic activity with p-nitrophenyl (pNP) substrates followed the order pNP-?-D-galactopyranoside (328?U?mg(-1)), pNP-?-D-glucopyranoside (246?U?mg(-1)), pNP-?-D-xylopyranoside (72?U?mg(-1)), and pNP-?-D-mannopyranoside (28?U?mg(-1)). Thus the enzyme was actually a multifunctional ?-glycosidase. Therefore, the utilization of Asac_1390 may contribute to facilitating the efficient degradation of lignocellulosic biomass and help enhance bioconversion processes.

SUBMITTER: Gumerov VM

PROVIDER: S-EPMC4619763 | biostudies-literature | 2015

REPOSITORIES: biostudies-literature

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A Novel Highly Thermostable Multifunctional Beta-Glycosidase from Crenarchaeon Acidilobus saccharovorans.

Gumerov Vadim M VM Rakitin Andrey L AL Mardanov Andrey V AV Ravin Nikolai V NV

Archaea (Vancouver, B.C.) 20151011

We expressed a putative β-galactosidase Asac_1390 from hyperthermophilic crenarchaeon Acidilobus saccharovorans in Escherichia coli and purified the recombinant enzyme. Asac_1390 is composed of 490 amino acid residues and showed high sequence similarity to family 1 glycoside hydrolases from various thermophilic Crenarchaeota. The maximum activity was observed at pH 6.0 and 93°C. The half-life of the enzyme at 90°C was about 7 hours. Asac_1390 displayed high tolerance to glucose and exhibits hydr ...[more]

PMID: 26539062

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Project description:Streptococcus intermedius is a known human pathogen and belongs to the anginosus group (S. anginosus, S. intermedius, and S. constellatus) of streptococci (AGS). We found a large open reading frame (6,708 bp) in the lac operon, and bioinformatic analysis suggested that this gene encodes a novel glycosidase that can exhibit ?-d-galactosidase and N-acetyl-?-d-hexosaminidase activities. We, therefore, named this protein "multisubstrate glycosidase A" (MsgA). To test whether MsgA has these glycosidase activities, the msgA gene was disrupted in S. intermedius. The msgA-deficient mutant no longer showed cell- and supernatant-associated ?-d-galactosidase, ?-d-fucosidase, N-acetyl-?-d-glucosaminidase, and N-acetyl-?-d-galactosaminidase activities, and all phenotypes were complemented in trans with a recombinant plasmid carrying msgA. Purified MsgA had all four of these glycosidase activities and exhibited the lowest Km with 4-methylumbelliferyl-linked N-acetyl-?-d-glucosaminide and the highest kcat with 4-methylumbelliferyl-linked ?-d-galactopyranoside. In addition, the purified LacZ domain of MsgA had ?-d-galactosidase and ?-d-fucosidase activities, and the GH20 domain exhibited both N-acetyl-?-d-glucosaminidase and N-acetyl-?-d-galactosaminidase activities. The ?-d-galactosidase and ?-d-fucosidase activities of MsgA are thermolabile, and the optimal temperature of the reaction was 40°C, whereas almost all enzymatic activities disappeared at 49°C. The optimal temperatures for the N-acetyl-?-d-glucosaminidase and N-acetyl-?-d-galactosaminidase activities were 58 and 55°C, respectively. The requirement of sialidase treatment to remove sialic acid residues of the glycan branch end for glycan degradation by MsgA on human ?1-antitrypsin indicates that MsgA has exoglycosidase activities. MsgA and sialidase might have an important function in the production and utilization of monosaccharides from oligosaccharides, such as glycans for survival in a normal habitat and for pathogenicity of S. intermedius.

Dataset Information

A Novel Highly Thermostable Multifunctional Beta-Glycosidase from Crenarchaeon Acidilobus saccharovorans.

Publications

A Novel Highly Thermostable Multifunctional Beta-Glycosidase from Crenarchaeon Acidilobus saccharovorans.

Similar Datasets

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