The dimorphic fungus Candida albicans, a harmless commensal in approximately 60% of healthy adults, is also an opportunistic pathogen in humans. Immuno-compromised individuals are mainly at risk, for example those under immunosuppressive therapies, HIV/AIDS patients, the elderly and early neonates. Diseases caused include invasive candidiasis, which has a 40-60% mortality rate. The majority of invasive candidiasis is caused by C. albicans forming biofilms on medical devices (e.g. catheters, heart valves) from which yeast cells disperse and enter the blood stream. The adherence properties of C. albicans are crucial for colonisation and biofilm formation. An important adhesive factor in the early stages of colonisation is Als1 (Agglutinin-like Sequence protein 1). Another crucial virulence factor is hyphal-expressed Als3, which contributes to adhesion and invasion of host cells. These proteins belong to the Als family of glycoproteins (Als1-7 and Als9), all of which share similar domain organisation: an N-terminal adhesion region comprised of two immunoglobulin-like domains, plus central and C-terminal regions, which are highly glycosylated. Previously, the N-terminus of Als9 had been shown to bind the flexible C-termini of host cell proteins as ligands. Due to their relevance in C. albicans adhesion, the N-terminal domains of Als1 and Als3 are ideal templates for ligand binding studies and the development of antifungal compounds. Shortened constructs Als1 and Als3 with improved solubility and structural properties (relative to the full length adhesins) were employed in this project. Here I describe the X-ray crystallographic structure of sNT-Als1 with a peptide ligand, the backbone resonance assignments by Nuclear Magnetic Resonance (NMR) of the ligand-free protein and ligand-binding experiments using Differential Scanning Fluorimetry (DSF). Comparison of the structure and ligand binding behavior of sNT-Als1 with other Als proteins will guide the design of therapeutic molecules that block the adhesive properties of these adhesins, and thus the formation of disease-causing biofilms by C. albicans.
| Identifer | oai:union.ndltd.org:bl.uk/oai:ethos.bl.uk:656798 |
| Date | January 2014 |
| Creators | Hale, Lisa |
| Contributors | Cota, Ernesto |
| Publisher | Imperial College London |
| Source Sets | Ethos UK |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
| Source | http://hdl.handle.net/10044/1/24906 |
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