An amylopullulanase (ApuNP1) from Geobacillus thermoleovorans NP1: biochemical characterization and its potential...

An amylopullulanase was produced by Geobacillus thermoleovorans NP1. The optimum enzyme production occurred at 45°C and pH 7.0 (12 hr). NP1 amylopullulanase (ApuNP1) exhibited the maximal activity at 50°C and pH 6.0 and was stable between 30-50°C, and pH 3.0-12.0 for 24 hr. The enzyme showed two bands with molecular weights of 112 and 107 kDa in sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). The amylopullulanase retained 100% of its activity in the presence of 10 mM of Ca²⁺, Ba²⁺, Zn²⁺, Mg²⁺, Cu²⁺, EDTA, and PMSF. While the enzyme showed resistance to 5% of TritonX-100, Tween 20, and Tween 80, the activity was inhibited by 5% β-mercaptoethanol and H₂O₂. While the hydrolysis products of pullulan were maltose, maltotriose, and maltodextrin, the starch was hydrolyzed to maltose, maltotriose, and maltodextrin units. This shows that NP1 pullulanase is a type II pullulanase (amylopullulanase). After the liquefaction assay, 12% glucose content was measured with a refractometer in the presence of 20% starch. According to the wash performance tests, the mixture of ApuNP1 and 1% detergent removed almost all of the stains. This novel thermo-acidic amylopullulanase has a potency to be used in detergent, starch, food, baking, textile, and cosmetic industries.