Syntheses of Probes for the Elucidation of Biosynthetic Pathways and Interactions of Natural Products

• Author / Creator

  Sanichar, Randy

• A new platform for the semi-synthetic preparation of a known nanomolar inhibitor of the Plasmodium falciparum lysyl-tRNA synthetase, cladosporin, from Cladosporium cladosporioides and its analogues was developed. Our work has shown that it is produced biosynthetically by a highly reducing (HR) and a non-reducing (NR) iterative type I polyketide synthase (PKS) pair, namely the Cla2 and Cla3. The heterologous expression of these enzymes in Saccharomyces cerevisiae was shown to produce cladosporin, thus confirming the identity of the putative gene cluster. In vitro incubation of the purified Cla3 enzyme with the synthesized natural pentaketide intermediate (or analogues) demonstrated the recognition and further elaboration of the intermediate to the completed product, cladosporin (or analogues). Incorporation of the natural pentaketide intermediate by Cla3 indicated a 5+3 assembly between the HR-PKS, Cla2 and the NR-PKS, Cla3 during cladosporin biosynthesis.
  Probes for the study of Coenzyme A (CoA) dependent natural product biosynthetic machinery were developed. Four CoA analogues were made in single steps through a transamidation reaction. The transformation was catalyzed by boric acid in water. This transformation offers access to useful compounds for the study of enzyme-catalyzed reactions, especially processes involving acyl/peptidyl carrier proteins (A/PCP) in polyketide synthases (PKS), fatty acid synthases (FAS), and nonribosomal peptide synthetases (NRPS). Currently work is ongoing to use these probes in the study of the Cla2/Cla3 interaction in cladosporin biosynthesis.
  A new Dess-Martin periodinane mediated oxidative rearrangement reaction was uncovered for the production of α-keto thioesters. The reaction proceeds through the oxidation of a β-hydroxy thioester to a β-keto thioester, followed by an α-hydroxylation and further oxidation to form a vicinal tricarbonyl thioester. This product then rearranges with loss of CO2, to form the α-keto product. The mechanism of the rearrangement was elucidated using 13C labelling and analysis of proposed intermediates as well as the products of the reaction. This process allows easy preparation of α-keto thioesters, which are vital intermediates in the preparation of pharmaceutically important heterocyclic scaffolds such as the quinoxalinones.
  The enantioselective preparation of spin-labelled α-amino acids for electron paramagnetic resonance (EPR) spectroscopy was studied. Spin labelled amino acids which are currently utilized are mostly racemic, labile or difficult to couple. Our work provides a simple approach towards the enantioselective preparation of two new spin labelled amino acids. This was achieved through alkylation of the chiral Ni(II) complex of the Schiff base of (S)-N-(2-benzoylphenyl)-1-benzylpyrrolidine-2-carboxamide and glycine with the spin-labelled electrophile of choice.

• Subjects / Keywords

  • Organic
  • Synthetic
  • Spin-label
  • Dess-Martin Periodinane Oxidative Rearrangement
  • Natural Products
  • Cladosporin
  • Anti-malarial

• Graduation date

  Fall 2018

• Type of Item

  Thesis

• Degree

  Doctor of Philosophy

• DOI

  https://doi.org/10.7939/R3NC5SV43

• License

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