Host-induced changes in the cell surface N-linked glycoproteins, from Aspergillus fumigatus. Search for specific targets with potential for clinical therapy and/or diagnosis.

This dissertation is presented to obtain a Master degree in Structural and Functional Biochemistry

The fungus Aspergillus fumigatus is responsible for causing invasive aspergillosis in human lungs, a fatal disease in immunocompromised patients. The development of such diseases is typically associated with a deficient immune response in the host as well as with phenotypic changes at cellular level of the fungus itself. The sequencing of the fungal genome has allowed the study of the proteome and its constituents, making it possible to explain the reason to such changes. The collection of carbohydrate moieties present in N- and O-linked glycoproteins and glycolipids, which protrude outwards from the cell membrane, has been defined as the exoglycome. Furthermore, studies have demonstrated that changes suffered by the exoglycome of A. fumigatus are the main cause of the fungus infectious potential. Therefore, identification and characterization of the different carbohydrate structures that comprise the fungal exoglycome has become of great importance in order to increase the knowledge of the fungus pathogenicity. In this study, several experimental techniques developed in proteomics and glycomics areas were used in an attempt to identify the main components of the cell membrane proteome of A. fumigatus as well as the N-linked oligosaccharides structure that comprise the fungal exoglycome. Two methods for glycoprotein detection were used that are based in the non-covalent binding of lectins to specific oligosaccharides and the oxidation of carbohydrate groups followed by conjugation with a chromogenic or tagged substrate. In an attempt to identify the glycoproteins that comprise the proteome was performed, mass spectrometry was used, however the results were inconclusive. Certain factors, such as lack of homology between the sequenced peptides from membrane proteins with protein sequences already described in the databases were evaluated and questioned, but with no conclusive answers.