Bimutation breeding of Aspergillus niger strain for enhancing β-mannanase production by solid-state fermentation

A parent strain Aspergillus niger LW-1 was mutated by the compound mutagenesis of vacuum microwave (VMW) and ethyl methane sulfonate (EMS). A mutant strain, designated as A. niger E-30, with high- and stable-yield beta-mannanase was obtained through a series of screening. The beta-mannanase activity of the mutant strain E-30, cultivated on the basic fermentation medium at 32 degrees C for 96 h, reached 36,675 U/g dried koji, being 1.98-fold higher than that (18,501 U/g dried koji) of the parent strain LW-1. The purified E-30 beta-mannanase, a glycoprotein with a carbohydrate content of 19.6%, had an apparent molecular weight of about 42.0 kDa by SDS-PAGE. Its optimal pH and temperature were 3.5 and 65 degrees C, respectively. It was highly stable at a pH range of 3.5-7.0 and at a temperature of 60 degrees C and below. The kinetic parameters K-m and V-max, toward locust bean gum and at pH 4.8 and 50 degrees C, were 3.68 mg/mL and 1067.5 U/mg, respectively. The beta-mannanase activity was not significantly affected by an array of metal ions and EDTA, but strongly inhibited by Ag+ and Hg2+. In addition, the hydrolytic conditions of konjak glucomannan using the purified E-30 beta-mannanase were optimized as follows: konjak gum solution 240 g/L (dissolved in deionized water), hydrolytic temperature 50 degrees C, beta-mannanase dosage 120 U/g konjak gum, and hydrolytic time 8 h. (C) 2011 Elsevier Ltd. All rights reserved.