Document Type

Dissertation

Degree

Doctor of Philosophy (PhD)

Department

Chemistry

First Advisor's Name

Kathleen S. Rein

First Advisor's Committee Title

Committee chair

Second Advisor's Name

David Becker

Second Advisor's Committee Title

Committee member

Third Advisor's Name

Watson Lees

Third Advisor's Committee Title

Committee member

Fourth Advisor's Name

Lidia Kos

Fourth Advisor's Committee Title

Committee member

Fifth Advisor's Name

Xiaotang Wang

Fifth Advisor's Committee Title

Committee member

Keywords

Karenia brevis, dinoflagellate, brevetoxin, epoxide hydrolase

Date of Defense

6-30-2015

Abstract

Polyether compounds are a subgroup of natural products with regular occurrence of multiple C-O-C motifs. The biosynthetic origin of polycylic polethers has been studied and the majority of them are derived from polyketide or terpene pathways. Normally, the polycyclic polyethers can be divided into two groups based on their structural features: the first group features multiple rings that are interconnected by carbon-carbon single bond, which are produced by a biosynthetic cascade of exo epoxide-opening reactions; the other group has multiple fused cyclic ethers and are formed by an cascade of endo epoxide-opening reactions.

Karenia brevis (K. brevis) is known as principle harmful bloom (HAB) organism of the Gulf of Mexico which can cause red tides. Brevetoxins (PbTx) are a suit of cyclic polyether ladder compounds produced by K. brevis. Brevetoxins are neurotoxins that can bind to voltage-gated sodium channels in nerve and muscle cells, resulting in disruption of normal neurological processes causing the human illness which is clinically described as neurotoxic shellfish poisoning (NSP).

Inspired by Cane-Celmer-Wesley’s proposal regarding monensin biosynthesis, Nakanishi and Shimizu proposed a biosynthetic pathway for brevetoxin which suggests that PKS-mediated synthesis of the polyene is followed by epoxidation to afford a polyepoxide which then undergoes an epoxide-opening cascade, catalyzed by an epoxide hydrolase (EH).

To find evidence to support the hypothesis that an epoxide hydrolase from polyether ladder producing dinoflagellates will catalyze the construction of the polyether ladder framework from polyepoxide substrates, and to study the role of epoxide hydrolase in the biosynthesis of polyether ladder compounds, it is necessary to identify and produce one or more epoxide hydrolase from dinoflagellates. The methods to detect epoxide hydrolase activity in K. brevis and different techniques to obtain epoxide hydrolases from K. brevis are discussed. A microsomal EH identified from a K. brevis EST library was cloned and expressed. The characterization of this EH, including substrate selectivity and enantioselectivity as well as its potential to catalyze the critical ento-tet cyclization epoxy alcohol, is discussed.

Identifier

FIDC000133

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