Optimization of protease and chitinase production by Bacillus cereus SV1 on shrimp shell waste using statistical experimental design. Biochemical and molecular characterization of the chitinase

Abstract

The current increase in the amount of shell waste produced by the shrimp industries has led to the need to find new methods for its disposal. In this study, statistical methodologies were used for chitinase and protease co-production optimization by Bacillus cereus SV1 in media containing shrimp shell powder. Medium composition and culture conditions were optimized using two statistical methods: Plackett–Burman design was applied to find the key ingredients and conditions for the best yield of enzymes production, and central composite design was used to optimize the levels of the five significant variables: shrimp shell powder (SSP), NH4Cl, CaCl2, K2HPO4 and speed of agitation. The medium optimization resulted in protease and chitinase activities of 8,445.8 U ml−1 and 82.8 mU ml−1, respectively. The crude chitinase exhibited maximal activity at 55°C and pH 7.0 using colloidal chitin as substrate. The metal ions Fe2+ and Mg2+ increased chitinase activity, while Hg2+ strongly inhibited this activity. The nucleotide sequencing of SV1 chitinase revealed an open reading frame containing 2,025 bp and encoding 642 amino acids.