Studies and application of the enzymes of fluorometabolite biosynthesis in Streptomyces cattleya
Abstract
This thesis focuses on studies investigating the structure of intermediates involved in
fluorometabolite biosynthesis, and the potential applications of the fluorinase enzyme in
positron emission tomography (PET).
Chapter 1 introduces the rare natural occurrence of fluorinated compounds. The bacterium
Streptomyces cattleya is known to biosynthesise two fluorinated secondary metabolites: the
toxin fluoroacetate (FAc) and the antibiotic 4-fluorothreonine (4-FT). The enzymes and
intermediates identified on this fluorometabolite biosynthetic pathway in S. cattleya, prior to
this research, are discussed in detail.
Chapter 2 presents studies towards the unambiguous structural identification of (3R,4S)-5-
deoxy-5-fluoro-D-ribulose-1-phosphate (5-FRulP) as the third fluorinated intermediate on the
biosynthetic pathway to fluoroacetate and 4-fluorothreonine in S. cattleya.
Chapter 3 describes the synthetic routes to key molecules, necessary as reference
compounds and substrates, to underpin the subsequent studies in this thesis. In particular,
synthetic routes to 5'-deoxy-5'-fluoroadenosine (5'-FDA), 5'-deoxy-5'-fluoroinosine (5'-FDI),
5-deoxy-5-fluoro-D-ribose (5-FDR) and 5-deoxy-5-fluoro-D-xylose (5-FDX) are described.
Chapter 4 describes the use of the fluorinase enzyme from S. cattleya as a tool for the
synthesis of new [¹⁸F]-labelled sugars with potential application in positron emission
tomography (PET). A new route to 5-deoxy-5-[¹⁸F]fluoro-D-ribose ([¹⁸F]FDR) is developed
in a two-step enzymatic synthesis. A total of three potential radiotracers ([¹⁸F]FDA,
[¹⁸F]FDR and [¹⁸F]FDI) are synthesised using fluorinase-coupled enzyme reactions.
In addition, in vitro studies are reported with these [¹⁸F]-labelled sugars to investigate their
uptake and potential as PET radiotracers in cancer cells. A preliminary rat imaging study with
[¹⁸F]FDA is reported.
Chapter 5 details the experimental procedures for the compounds synthesised in this
research and the biological procedures for chemo-enzymatic syntheses and protein
purification.
Type
Thesis, PhD Doctor of Philosophy
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