Degree Type

Thesis

Date of Award

2012

Degree Name

Master of Science

Department

Biochemistry, Biophysics and Molecular Biology

First Advisor

Reuben J Peters

Abstract

Terpenoids form a large superfamily of natural products with over 55,000 known. Of these, the labdane-related diterpenoids (LRDs) form the largest class of these natural products including phytohormones such as gibberellins which are key compounds involved in the growth and development of plants. LRDs proceed via the protonation-initiated bicyclization of (E,E,E)-geranylgeranyl diphosphate to various stereoisomers of labdadienyl/copalyl diphosphate from a class II diterpene cyclase. Little is understood about the mechanism of class II diterpene cyclases. This catalyzed reaction proceeds through a labda-13-en-8-yl+ intermediate which is quenched by deprotonation through a catalytic basic residue. Here we explore an active site His in various class II diterpene cyclases that could be potential candidates for this deprotonation. We performed site-directed mutagenesis on these residues to smaller side-chain residues and found that these enzymes were incorporating water into the catalyzed reaction to form various stereoisomers copal-8-ol diphosphate. These results provide new insight into class II diterpene cyclases, specifically the role of a His as the potential catalytic base and the ability of these cyclases to incorporate water into the catalyzed reaction.

Copyright Owner

Jared Criswell

Language

en

Date Available

2013-10-31

File Format

application/pdf

File Size

64 pages

Included in

Biochemistry Commons

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