Date of Award

2017

Document Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Department

Biotechnology Science and Engineering

Committee Chair

Robert L. McFeeters

Committee Member

Hana McFeeters

Committee Member

Luis R. Cruz-Vera

Committee Member

Mary J. Bossard

Committee Member

Emanuel Waddell

Subject(s)

Fungal diseases of plants, Cryptococcus, Mannose, Lectins, Biotechnology

Abstract

Fungal pathogens represent a growing concern in human health, and the advancement of new therapeutic options is essential. The encapsulated basidiomycotic yeast Cryptococcus causes severe meningoencephalitis and cryptococcosis results in over 600,000 deaths a year. The prevalence of cryptococcal infection is increasing, driving the need for new and improved treatments. Scytovirin is a high mannose binding lectin from the cyanobacterium Scytonema varium. Originally pursued as an antiviral, herein is reported novel antifungal activity with detailed characterization of activity against Cryptococcus. Scytovirin specifically binds Man4, the D3 arm of Man9, a carbohydrate moiety found on cryptococcal glycoproteins. Localization of Scytovirin in Cryptococcus has found that the protein accumulates near the cell wall, where it likely binds an essential mannoprotein. Additionally, Scytovirin exhibits synergy with common antifungals, inferring the utilization of a novel inhibitory mechanism. The extreme carbohydrate specificity of Scytovirin, made possible by its unique fold, results in a benign safety profile. To further understand the structure/function relationship of Scytovirin, a similar protein, Myxovirin, has been discovered and is being characterized. Myxovirin affords numerous advantages including a single carbohydrate binding domain that simplifies NMR studies, an N-terminus that is amenable to alterations, and a smaller size which is beneficial in protein therapeutic development. In addition to Scytovirin inhibition of Cryptococcus, the broader antifungal capabilities of high mannose binding lectins have been investigated, including studies of the lectin Griffithsin and the fungi Botrytis cinerea, Aspergillus niger, Saccharomyces cerevisiae, and Pichia pastoris. Griffithsin, specific for terminal mannoses on Man9, was found to inhibit Cryptococcus. This result infers the binding of Man9 to be a major factor in the inhibition of this pathogen. Furthermore, it was discovered that Scytovirin is able to inhibit the growth of pathogenic molds B. cinerea and A. niger. In both cases, localization of Scytovirin determined that binding was occurring near the cell wall. It is clear from this research that the use of high mannose binding lectins to inhibit fungi represents an untapped potential in the field of medicinal mycology. As the threat of pathogenic fungi continues to increase, further efforts into antifungal development need to be a priority.

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