Abstract
BackgroundGlucoamylase is an exo-type enzyme that converts starch completely into glucose from the non-reducing ends. To meet the industrial requirements for starch processing, a glucoamylase with excellent thermostability, raw-starch degradation ability and high glucose yield is much needed. In the present study we selected the excellent Carbohydrate-Activity Enzyme (CAZyme) producer, Bispora sp. MEY-1, as the microbial source for glucoamylase gene exploitation.Methodology/Principal FindingsA glucoamylase gene (gla15) was cloned from Bispora sp. MEY-1 and successfully expressed in Pichia pastoris with a high yield of 34.1 U/ml. Deduced GLA15 exhibits the highest identity of 64.2% to the glucoamylase from Talaromyces (Rasamsonia) emersonii. Purified recombinant GLA15 was thermophilic and showed the maximum activity at 70°C. The enzyme was stable over a broad pH range (2.2–11.0) and at high temperature up to 70°C. It hydrolyzed the breakages of both α-1,4- and α-1,6-glycosidic linkages in amylopectin, soluble starch, amylose, and maltooligosaccharides, and had capacity to degrade raw starch. TLC and H1-NMR analysis showed that GLA15 is a typical glucoamylase of GH family 15 that releases glucose units from the non-reducing ends of α-glucans. The combination of Bacillus licheniformis amylase and GLA15 hydrolyzed 96.14% of gelatinized maize starch after 6 h incubation, which was about 9% higher than that of the combination with a commercial glucoamylase from Aspergillus niger.Conclusion/SignificanceGLA15 has a broad pH stability range, high-temperature thermostability, high starch hydrolysis capacity and high expression yield. In comparison with the commercial glucoamylase from A. niger, GLA15 represents a better candidate for application in the food industry including production of glucose, glucose syrups, and high-fructose corn syrups.
Highlights
Starch is a polysaccharide carbohydrate of plant cells, and is composed solely of aglucose units that are linked by a-1,4- or a-1,6-glycosidic bonds
Amylopectin is an a-1,4linked D-glucose polymer with approximately 5% of a-1,6-linked branches, whereas amylose is a linear polymer essentially consisting of a-1,4-linked glucopyranose residues [1]
A gene fragment of 704 bp was cloned from the genome of Bispora sp
Summary
Starch is a polysaccharide carbohydrate of plant cells, and is composed solely of aglucose units that are linked by a-1,4- or a-1,6-glycosidic bonds. It has two homopolysaccharide types: amylopectin and amylose. Starch is mainly used to produce glucose, which can be further utilized to produce high-fructose syrups, bioethanol, organic acids and amino acids [2, 3]. Glucoamylase is an exo-type enzyme that converts starch completely into glucose from the non-reducing ends. To meet the industrial requirements for starch processing, a glucoamylase with excellent thermostability, raw-starch degradation ability and high glucose yield is much needed. Methodology/Principal Findings: A glucoamylase gene (gla15) was cloned from Bispora sp.
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