Investigating the Functional Roles of the Sialidase NanH and Glycopeptidase Amuc_1438 in the Enzymatic Degradation of Mucin




Medley, Brendon

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The mucosal layer within the gastrointestinal tracts of animals and humans serves a critical function in safeguarding the epithelial layer against pathogens, including Clostridium perfringens. This layer consists of mucin-based glycoproteins, forming a dual-layered structure: an inner layer firmly attached to the epithelial cells, acting as a protective barrier, and an outer layer that fosters a favourable environment for commensal organisms. Both commensal and pathogenic bacterial species possess an array of enzymes designed for mucin degradation, fulfilling two main purposes: utilizing the abundant carbohydrate network as an energy and carbon source, and breaking down the mucus layer during pathogenic invasion. However, our understanding of these enzymatic tools employed by the gut microbiota remains incomplete, leaving gaps in our knowledge concerning pathogenic invasion and host-microbe interactions. This thesis presents a comprehensive analysis of two enzymes involved in mucin degradation: the intracellular sialidase NanH from Clostridium perfringens and the previously uncharacterized Amuc_1438 from Akkermansia muciniphila. The thesis investigates the structure of NanH, comparing it to its extracellular counterpart NanI, and provides structural insight into the specificity for sialic acids linked to glycans with α(2,3) over α(2,6) linkages. Additionally, through structural and functional investigations, Amuc_1438 is revealed to possess glycopeptidase activity, targeting specifically glycopeptides containing a Tn-antigen on a serine or threonine residue. Intriguingly, this work outlines that Amuc_1438 likely belongs to an uncharacterized family of glycopeptidases. The primary aim of this thesis is to demonstrate how these two distinct enzymes fit into the enzymatic pathway of mucin degradation observed in both commensal and pathogenic gut bacteria. By shedding light on the structural characteristics and functional roles of NanH and Amuc_1438, this research contributes to a deeper understanding of the intricate enzymatic processes involved in mucin degradation, thus enhancing our knowledge of pathogenic mucosal-invasion strategies and host-microbe interactions.



Mucin, enzyme, Protein, Hydrolysis, Health, Gut