Thesis
Establishing a chemoenzymatic platform for the synthesis of nucleoside analogues
- Creator
- Rights statement
- Awarding institution
- University of Strathclyde
- Date of award
- 2025
- Thesis identifier
- T17348
- Person Identifier (Local)
- 202169762
- Qualification Level
- Qualification Name
- Department, School or Faculty
- Abstract
- Nucleosides are essential molecular tools for use in chemical biology workflows and as therapeutics. The importance of nucleosides as antiviral agents has been demonstrated by the recent COVID-19 pandemic triggered by SARS-COV-2. A major challenge in the field of nucleoside research is the lack of preparative methods to access novel nucleoside scaffolds. Existing chemical approaches for the synthesis of nucleoside analogues are burdened by multi-step syntheses requiring environmentally harmful reagents, often producing complex mixtures of regio- and stereoisomers. Enzymatic approaches provide a more step-efficient and environmentally sustainable alternative for nucleoside synthesis. Traditionally, industrial enzymatic methods have used nucleoside phosphorylases (NPs) which produce nucleosides via a two-step approach. In contrast, nucleoside 2′ deoxyribosyltransferases (NDT) enzymes catalyse the synthesis of nucleoside analogues stereospecifically via the transglycosylation of nucleobases with a nucleoside donor. This work describes the development of a platform to prepare a broad range of non-canonical nucleosides using NDT-2 (LlNDT-2) derived from Lactobacillus leichmanii. The screening of over 40 nucleobase substrates established a comprehensive structure-activity profile as well as novel mild reaction conditions to form these nucleosides (including 100% H2O). X-ray structural analysis with ligands bound identified the pyrimidine and purine acceptor substrates by LlNDT2. Additionally, overexpression of wildtype NDT and corresponding mutants enhanced the understanding of substrate promiscuity in transglycosylations of 2’-deoxy and ribonucleosides is described, including using crude cell lysate for a more streamlined enzymatic process. Lastly, the enzymatic scale up to one gram of nucleoside analogues allows facile access to 15N labelled materials. This approach provides the basis to establish a convergent, step-efficient chemoenzymatic platform for the scalable preparation of high value 2'-deoxy and ribose nucleosides.
- Advisor / supervisor
- Burley, Glenn
- Resource Type
- DOI
- Embargo Note
Relations
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File | 2025-06-03 | Embargo |