Nature's approach toward ring formation and structural diversity in ergot alkaloid biosynthesis

Ergot alkaloids are fungal-derived secondary metabolites well known for a diverse array of pharmacological effects both beneficial and detrimental to human health. Historically, the ergot alkaloids have been known to cause ergotism in populations that consume grain contaminated by ergot alkaloid producing fungus. However, naturally isolated and semi-synthetic derivatives of certain ergot alkaloids have also been used to treat migraines, Parkinsonism, and tumor growth. Different fungal species such as Aspergillusfumigatus, Claviceps purpurea, and Neotyphodium lolii produce ergot alkaloids that have distinct structural features yet share a common tetracyclic ergoline ring scaffold. Mechanistic details regarding the formation of the common ergoline ring are not well understood, though the genes encoding the enzymes that carry out these cyclizations are believed to be well conserved across divergent fungal species. Here we describe in vivo gene disruption experiments in Aspergillusfumigatus that allowed us to identify candidate enzymes that were directly involved with the intramolecular cyclization of the ergoline ring. Additionally, we discuss the cloning and heterologous expression of these genes to further characterize their catalytic function. Old Yellow Enzyme homologues from the ergot gene clusters of Aspergillus fumigatus and Neotyphodium loii were shown to catalyze the formation of the D ring of the ergoline skeleton. These enzymes catalyzed either reductase or isomerase type reactions to yield distinct pathway intermediates. Mutational analysis was used to engineer an Old Yellow Enzyme that displayed both reductase and isomerase activities, thereby elucidating the mechanistic basis behind this switch in enzymatic activity. These findings present a mechanistic rationale behind nature's biosynthetic strategy toward ring cyclization and the introduction of structural diversity into the ergot alkaloid class of natural products.

Department dc:contributor.department

Massachusetts Institute of Technology. Dept. of Chemistry.

Grantor dc:publisher

Massachusetts Institute of Technology

Year dc:date.issued

2011

Author dc:creator

• Cheng, Johnathan Zandrew

Advisor dc:contributor.advisor

• Sarah E. O'Connor.

• Chemistry.

Statement dc:rights

• M.I.T. theses are protected by copyright. They may be viewed from this source for any purpose, but reproduction or distribution in any format is prohibited without written permission. See provided URL for inquiries about permission.

Licence dc:rights.uri

• http://dspace.mit.edu/handle/1721.1/7582

Language dc:language.iso

eng

Handle dc:identifier.uri

http://hdl.handle.net/1721.1/65472

OAI identifier oai:identifier

oai:dspace.mit.edu:1721.1/65472