Partial purification and characterization of a recombinant β-mannanase from Aspergillus fumigatus expressed in Aspergillus sojae grown on carob extract

Abstract

Mannanases are of great importance as they are able to hydrolyze polysaccharides for industrial applications and can be easily produced by fermentation. Purification of a β-mannanase produced from carob extract in a bioreactor is of great importance in order to obtain information about its structural and functional features and to foresee its implementations. Therefore, the aims of this study were to purify a β-mannanase from Aspergillus fumigatus expressed in Aspergillus sojae grown on carob extract by implementation of various flocculant agents in the fermentation medium, followed by the determination of its physico-chemical, kinetic, and thermodynamic characteristics. Flocculants increase the visible size of the dispersed particles by agglomeration, simplifying the separation of solids and liquids, and thereby reducing process costs. Optimum flocculation conditions for chitosan- and PAC-PAM-implemented processes were determined as 40 mg/L of chitosan, 30 min of implementation time, no agitation and 3 g/L PAC-PAM, 30 min implementation time, and no agitation. Under these conditions, more than a 2-fold increase in the enzyme activity was achieved in both flocculation processes. Further investigations determined molecular weight, optimum pH, and temperature values of the produced β-mannanase to be 56–60 kDa, pH 6, and 60 °C, respectively. Additionally, Km and Vm determination of the enzyme for various substrates concluded that the β-mannanase showed the highest substrate specificity against locust bean gum among the substrates evaluated. Furthermore, experimental results for substrate specificity were further modeled and successfully represented by the Weibull model. In conclusion, this study provides valuable and important information about β-mannanase production and purification for industrial-scale applications.