Abstract
Maltooligosaccharide-forming amylases (MFAses) are promising enzymes for a variety of industrial applications. In this study, a maltooligosaccharide-forming amylase (BkAmy) isolated from Bacillus koreensis HL12 was first heterologous expressed and characterized. According to structural-sequence alignment, BkAmy contained seven conserved regions which are the signature of a novel GH13 subfamily. The gene was expressed in Pichia pastoris KM71 as an extracellular protein with a volumetric activity of 3.38 U/mL culture medium after 72 h induction by 3% (w/v) of methanol. The recombinant BkAmy migrated as a single protein band with an expected size approximately of 55 kDa. BkAmy exhibited the highest catalytic activity on soluble starch with a specific activity of 42.2 U/mg at 40 °C, pH 7.0. The enzyme exhibited 65% relative activity at 30 °C, indicating its advantage on application at moderate reaction temperature desirable for energy saving and reduction of side unwanted reactions. The enzyme exhibited a specific cleavage pattern by releasing maltose (G2), maltotriose (G3) and maltotetraose (G4) from cassava starch with the highest yield of 363 mg/g substrate equivalent to 36% conversion using 40 U/g substrate at 60 min. The work demonstrates the potential of this enzyme on maltooligosaccharide production from starch to create high value-added products in starch processing industries.
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Acknowledgements
The authors acknowledge the support provided by the Department of Biotechnology, Faculty of Science and Technology, Thammasat University. The authors are grateful to Natinee Junmat for soil sample collection. Daran Prongjit was supported by the Thailand Graduate Institute of Science and Technology (TGIST) (Grant number: SCA-CO-2563-12209-TH), National Science and Technology Development Agency.
Funding
This study was financially supported by Thammasat University Research Fund (Contract no. TUFT 042/2563). This study was also financially supported by the National Science and Technology Development Agency (Grant no. P18-52705).
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HL, BB and VC conceived and designed the work. HL, BB, NP and DP performed the experiments, and analyzed the data. HL and BB drafted the manuscript. BB and VC critically revised the manuscript for intellectual content. All authors have read and agree to the submission of the manuscript.
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Lekakarn, H., Bunterngsook, B., Pajongpakdeekul, N. et al. A novel low temperature active maltooligosaccharides-forming amylase from Bacillus koreensis HL12 as biocatalyst for maltooligosaccharide production. 3 Biotech 12, 134 (2022). https://doi.org/10.1007/s13205-022-03188-1
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DOI: https://doi.org/10.1007/s13205-022-03188-1