Electronic Thesis and Dissertation Repository

Thesis Format

Monograph

Degree

Master of Science

Program

Biochemistry

Supervisor

Garnham, Christopher

Affiliation

Agriculture and Agri-Food Canada

2nd Supervisor

Junop, Murray

Co-Supervisor

Abstract

Fumonisins are a family of mycotoxins produced by Aspergillus and Fusarium spp. fungi whose contamination of food and feed presents a worldwide agroeconomic threat. Certain Aspergillus spp. post-biosynthetically convert their B series fumonisins (FB) to less toxic forms, replacing their terminal amine with a ketone (FPy) or hydroxyl (FLa). AnFAO is an enzyme responsible for FB deamination in Aspergilli, however FLa-generating enzymes remain unknown. Additionally, Fusarium’s potential fumonisin-detoxifying ability has not been characterized. Herein, I identify FLa-generating enzymes (FUM13) and characterize the fumonisin-deaminating activity of an F. verticillioides AnFAO homolog (FvFAO). A. niger and F. verticillioides FUM13 isoforms reduced FPy fumonisins to FLa forms overnight with low efficiency. I also report that FvFAO deaminates FB fumonisins ~5x less efficiently than AnFAO. These results provide insight into differences in fumonisin self-protection mechanisms between these genera, and may lead to the development of new tools for treatment of fumonisin-contaminated food and feed.

Summary for Lay Audience

Mycotoxins are toxic secondary metabolites produced by crop-dwelling fungi. They are not required for growth and development, however, they help fungi break down plant matter to more easily absorb nutrients. Fumonisins are a family of mycotoxins produced by various Fusarium and Aspergillus spp. Their colonization of various crops, and subsequent fumonisin production, presents a significant agroeconomic threat as they are associated with many diseases in plants, animals, and humans.

B series fumonisins (FB) are the most abundant and most toxic. They are composed of a linear poly-carbon backbone with many functional groups, one of the most important being a terminal amine (–NH2). This amine is primarily responsible for their toxicity, allowing FB fumonisins to inhibit ceramide synthase, an enzyme involved in sphingolipid biosynthesis.

Notably, certain Aspergillus spp. convert their FB fumonisins to less toxic FPy and FLa forms. Their only difference is the replacement of the amine by a carbonyl (=O) in the FPy form, and a hydroxyl (–OH) in the FLa form. Our lab hypothesized that there is a sequential, enzymatic detoxification process where FB fumonisins are converted to FPy forms, which are then converted to FLa forms. The enzyme responsible for FB to FPy conversion has already been identified and characterized. However, the FLa-generating enzyme(s) remained unknown.

In this thesis, I identify an enzyme from A. niger and F. verticillioides (FUM13) capable of FPy to FLa conversion. FUM13 converts the FPy carbonyl to the FLa hydroxyl in vitro, albeit very slowly. Importantly, this work led to the investigation of F. verticillioides’ potential to post-biosynthetically modify its own fumonisins.

Neither FPy nor FLa production by F. verticillioides has previously been reported in the literature. However in this thesis, in addition to identifying an F. verticillioides FLa-generating enzyme for the first time, I biochemically characterize the newly discovered FPy-generating activity an additional F. verticillioides enzyme (FvFAO). Overall, this work adds to our growing understanding of fumonisin self-protection methods in Fusaria and Aspergilli, and provides a starting point for the development of new tools to mitigate fumonisin contamination in food and feed.

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