Epsilon Open Archive

Abstract

Cellulose and chitin, the main ways of storing biological energy in nature, also play a vital role in the structures of many organisms. Cellulose is the main structural component in plants whereas chitin is found in invertebrates and fungi. Gaining a better understanding of the degradation of these polymers can have direct or indirect economic impact. This thesis summarizes the structural perspectives of the cellulose and chitin degradation machinery. The white-rot fungus Phanerochaete chrysosporium has six cellobiohydrolases, which are expressed differentially with varying stimuli and time intervals. X-ray crystal structures of one of the six isozymes (Pc_Cel7D) suggested that it uses a retention mechanism and acts from the reducing end of cellulose chain. Homology modeling of the other enzymes supported the same sort of mechanism for all except one (Pc_Cel7B) and considerably different dynamic properties for two isozymes (Pc_Cel7A and Pc_Cel7B). Piromyces sp. strain E2 Cel9A and Cel6A as well as Piromyces equi Cel6A are modular structures, which function as parts of the fungal cellulosome of the respective organisms. Homology modeling supported the conclusion that Cel9A is an endoglucanase having a wide active site cleft and a conserved calcium-binding site with an inverting catalytic mechanism, whereas the Cel6As are processive cellobiohydrolases that act via an inverting mechanism that releases cellobiose from the non-reducing end of the cellulose chain. Brassica juncea endo acting chitinase is a pathogenesis-related protein that acts in defense of the plant. A homology model of the catalytic module was useful in designing mutants that helped us to understand the substrate binding and catalytic processes. X-ray crystal structures of the catalytic module and a mutant extended the knowledge of how the enzyme acts during the catalysis, with conformational changes opening and closing the enzyme. The homology model of yam, Dioscorea opposita, class IV endochitinase suggests that this enzyme catalyzes chitin cleavage via an inverting mechanism. Deletions in class IV chitinases compared to class I/II cluster at the ends of the substrate-binding cleft, shortening it by one glycosyl unit at each end. The shorter cleft might be expected to recognize and grasp a small section of exposed chitin on a fungal hyphal wall, more effectively attacking it.