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Transglycosylating glycoside hydrolase family 1 β-glucosidase from Penicillium funiculosum NCL1: Heterologous expression in Escherichia coli and characterization

Paper ID Volume ID Publish Year Pages File Format Full-Text
2868 140 2015 8 PDF Available
Title
Transglycosylating glycoside hydrolase family 1 β-glucosidase from Penicillium funiculosum NCL1: Heterologous expression in Escherichia coli and characterization
Abstract

•A β-glucosidase encoding gene was amplified from the potential celluloytic fungi Penicllium funiculosum NCL1.•The gene was successfully over-expressed in Escherichia coli.•The recombinant enzyme showed broad substrate specificity and excellent catalytic activity.•The enzyme possess transglycosylation ability by which glucose is converted to gentiobiose, a rare oligosaccharide with the prebiotic value.

A β-glucosidase cDNA belonging to glycoside hydrolase family 1 was cloned from RNA isolated from lactose grown cellulolytic fungus, Penicillium funiculosum NCL1 by reverse-transcriptase polymerase chain reaction. The gene (bgl6) contained an open reading frame of 1470 bp that encoded 490 amino acids. bgl6 gene was overexpressed in Escherichia coli BL21-CodonPlus and the recombinant enzyme (rBgl6) was purified. The rBgl6 was a monomer with the molecular weight of 52 kDa. The enzyme was optimally active at pH 6.0, 50 °C and stable in the wide range of pH and temperature. rBgl6 showed higher activity with pNPG, pNPGal cellobiose and lactose, while it showed substantial activity on pNPX. When the substrate was pNPG, the enzyme showed glucose tolerance at a concentration of 500 mM. rBgl6 exhibited enhanced activity with metal cations Ca2+, Mn2+, Mg2+ and Zn2+, while it showed stability with DTT and EDTA. The enzyme showed Km values of 0.125 and 0.25 mM, whereas Vmax of 256 and 166 μmol/mg with pNPG and cellobiose respectively. rBgl6 possessed transglycosylation activity and it converted glucose into gentiobiose, a rare oligosaccharide with greater prebiotic value. The yield of gentiobiose was 0.57 mg/ml for 5 mg/ml of glucose. These results indicated that rBgl6 could be a valuable candidate for the enzymatic synthesis of gentiobiose.

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Transglycosylating glycoside hydrolase family 1 β-glucosidase from Penicillium funiculosum NCL1: Heterologous expression in Escherichia coli and characterization
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Publisher
Database: Elsevier - ScienceDirect
Journal: Biochemical Engineering Journal - Volume 102, 15 October 2015, Pages 6–13
Authors
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Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering
Get Full-Text Now
Don't Miss Today's Special Offer
Price was $35.95
You save - $31
Price after discount Only $4.95
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Full-text PDF Download
Online Support
Any Questions? feel free to contact us