Statistical optimization of halophilic chitosanase and protease production by Bacillus cereus HMRSC30 isolated from Terasi simultaneous with chitin extraction from shrimp shell waste

Abstract

Chitosanase is an industrially important enzyme involved in the production of bioactive chitooligosaccharides. In the present study, Bacillus cereus HMRSC30 isolated from the Indonesian shrimp paste was found to have high ability to assimilate shrimp shell powder (SSP), and selected as the microbial tool to produce low-cost chitosanase under statistical optimization. The highest chitosanase production (4.85 U/mL) was achieved under the optimal conditions: 20 g/L SSP, 3 g/L (NH4)2SO4, 0.3 g/L Mg2SO4, 1.5 g/L KH2PO4, 1.5 g/L K2HPO4, pH 5.5 with a 15% (v/v) inoculum size at 35 °C for 3 days incubation. Besides chitosanase, a high amount of protease (33.47–72.19 U/mL) was also synthesized during 7 days fermentation. The optimal fermentation conditions also simultaneously deproteinized (76.36–97.86%) and demineralized (24.07–67.22%) the SSP. These observations indicated the potential of the strain HMRSC30 in a biological process for the enzyme production simultaneous with chitin extraction. The purified chitosanase with an apparent MW of 41 kDa exhibited the highest specificity to 75–85% deacetylated chitosan in soluble form (100%), followed by colloidal form (73.6%) and soluble chitin (73.4%) in an acidic pH of 5.5 at 55 °C. Additionally, it was discovered that the chitosanase obtained was a halophilic enzyme which was fully activated by 2.5% (w/v) NaCl and displayed activity of 81.73% even in the presence of 7.5% (w/v) NaCl. The enzyme yielded a putative dimer, and trimer of glucosamine as the predominant hydrolysis products. This study revealed that the properties of chitosanase from the strain HMRSC30 would be useful in industrial applications.