Production optimization, purification and characterization of a heat-tolerant acidic pectinase from Bacillus sp. ZJ1407

Abstract

Medium compositions for a heat-tolerant acidic pectinase production from Bacillus sp. ZJ1407 were optimized via response surface methodology (RSM) and its enzymatic properties were investigated. A 2-level factorial design was used to estimate the main effect of factors, and to screen the significant factors. A central composite design was used to find out the optimal concentrations of screened key factors. Lactose, tryptone and (NH4)2SO4 were found to have a significant influence on the pectinase activity (p <0.05). The optimal medium compositions were as follows: lactose 44.8g/l, tyrptone 30.9g/l, (NH4)2SO4 1.35g/l, MnSO4·H2O 0.2g/l, MgSO4 0.4g/l and NaCl 3.5g/l. Pectinase was purified to homogeneity by ammonium sulphate precipitation, DEAE-cellulose ion-exchange chromatography and Sephadex G-100 size-exclusion chromatography. The molecular weight of the purified pectinase determined by SDS-PAGE was about 23kDa, and its final specific activity was 110.47U/mg. Its optimal temperature and pH were 37°C and 5.0, respectively. Pectinase was very stable within a pH range of 3.0–5.0, and showed a high thermo-stability at 80 and 90°C. Ba2+ could significantly promote the activity of pectinase, and Mn2+ heavily inhibited its activity. This study provides new insight into the future development and use of pectinase from Bacillus sp. ZJ1407.