Identification and Analysis of the Role of Chitinase Substrate in Francisella Novicida

Abstract

Francisella tularensis is a gram negative facultative intracellular pathogen which is a class A biothreat according to the CDC. This bacteria codes for two chitinases (ChiA and ChiB) and one chitin binding protein (CbpA). Chitin is the most abundant oligosaccharide in marine environments and in the exoskeleton of many insects. In order to digest this material, microorganisms produce chitinases capable of cleaving this polymer. The chitinases in Francisella novicida have previously been found by our lab to negatively regulate the bacterial biofilm, most likely cleaving the extracellular polysaccharide substance (EPS) with β-1,4 glycosidic linkages. These enzymes may enable the organism to use the resulting cleaved polymers as carbon and nitrogen sources for growth. Although Francisella encodes for several polysaccharide synthases, it has not been found to produce chitin. Therefore, we hypothesized that the chitinase-enzyme substrate that is self-produced in the F. novicida EPS and biofilm must be some other molecule also containing β-1,4 glycosidic linkages. The biochemical characterization of ChiB activity on various substrates will be evaluated and the role of the substrate in the biofilm will be examined. Based on the preliminary data from the van Hoek lab and other labs, we think that chitinases play important roles both in Francisella ecological persistence and macrophage mediated growth during Francisella infection. Identifying and characterizing the substrate for these chitinases will generate information on how Francisella uses them to persist in the environment and replicate in human macrophages.