Production and characterization of thermostable acidophilic β-mannanase from Aureobasidium pullulans NRRL 58524 and its potential in mannooligosaccharide production from spent coffee ground galactomannan

The maximum yield of the crude mannanase from Aureobasidium pullulans NRRL 58524 was 8.42 +/- 0.18 U mL(-1) when cultured for 72 h in the optimized medium containing 3% (w v(-1)) defatted spent coffee grounds (SCG) and 0.67% (w v(-1)) ammonium sulphate. Two forms of mannanase were observed in the crude enzyme and the principal mannanase was enriched to apparent homogeneity via sequential filtration and anion exchange chromatography. The molecular mass of the enzyme was approximately 63 kDa as determined by SDS-PAGE. The enriched mannanase was active at high temperatures (45-75 degrees C) and a pH range from 3 to 6 with the maximum activity at 55 degrees C and pH 4.0. The enzyme was relatively thermostable with more than 75% of its initial activity remained after a 12 h incubation at 55 degrees C. The half-lives of the enriched mannanase were over 8 and 6 h at 60 and 65 degrees C, respectively. The enzyme was not adversely affected by chelator and most ions tested. This enzyme could hydrolyze both glucomannan and galactomannan and exhibited limited catalytic activity on beta-glucan. When the crude mannanase was used to hydrolyze galactomannan extracted from SCG, the maximum yield of reducing sugars mainly comprising of mannobiose (16.27 +/- 0.84 mg 100 mg(-1)), and mannotriose (2.85 +/- 0.20 mg 100 mg(-1)) was obtained at 58.22 +/- 2.04 mg 100 mg(-1) dry weight, under optimized condition (84.87 U g(-1) mannanase, 41 h 34 min incubation at 55 degrees C and pH 4.0). These results suggested the prospect of the enzyme in mannan hydrolysis and mannooligosaccharide production at a larger scale.

Automated summary

The crude mannanase from Aureobasidium pullulans NRRL 58524 achieved maximum yield in the optimized medium with 3% defatted spent coffee grounds and 0.67% ammonium sulphate. The enzyme showed thermostability and maximum activity at 55 degrees C and pH 4.0. It also had limited catalytic activity on beta-glucan and was not affected by most ions and chelators. The potential of this enzyme in mannan hydrolysis and mannooligosaccharide production at a larger scale was demonstrated.