Investigating the Antifungal Potential of Genetically Modified Hybrid Chitinase Enzymes Derived from Bacillus subtilis and Serratia marcescens

Abstract

Chitinases are glycosyl hydrolase enzymes that break down chitin, an integral component of fungal cell walls. Bacteria such as Bacillus subtilis and Serratia marcescens produce chitinases with antifungal properties. In this study, we aimed to generate hybrid chitinase enzymes with enhanced antifungal activity by combining functional domains from native chitinases produced by B. subtilis and S. marcescens. Chitinase genes were cloned from both bacteria and fused together using overlap extension PCR. The hybrid constructs were expressed in E. coli and the recombinant enzymes purified. Gel electrophoresis and computational analysis confirmed the molecular weights and isoelectric points of the hybrid chitinases were intermediate between the parental enzymes. Antifungal assays demonstrated that the hybrid chitinases inhibited growth of the fungus Fusarium oxysporumsignificantly more than the native enzymes. Our results indicate domain shuffling of bacterial chitinases is a promising approach to engineer novel antifungal proteins. This study provides insight into structure-function relationships of chitinases and strategies for generating biotherapeutics with enhanced bioactive properties.