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Title: Novel chassis for engineered pathways and automating biological experiments in the biotechnology industry
Author: Asra, Aisha Mariam
ISNI:       0000 0004 7965 0021
Awarding Body: UCL (University College London)
Current Institution: University College London (University of London)
Date of Award: 2019
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For the pharmaceutical industry, there has been a movement towards greener chemistry to reduce environmental impact. This movement has sparked interest from the pharmaceutical and chemicals industry in synthetic biology, which involves increasing the use of biological hosts and enzymatic processes to produce pharmaceutical products. This increase in biocatalysis and synthetic biology has sparked more of a need for diverse and versatile enzymes in order to customise the end product as well as diverse and robust host organisms to accommodate the reactions. Synthetic biology also opens opportunities for the discovery of complex molecules which have been inaccessible by traditional synthetic chemistry routes. Using enzymes from the toluene metacleavage pathway found in Pseudomonas putida and an omega-transaminase, an engineered metabolic pathway was constructed in Escherichia coli and in various P. putida strains for the production of novel amines. In this study the conversion of benzoate to 2-hydroxymuconic semialdehyde (2-HMSA) has been investigated in a whole cell biotransformation reaction to compare the feasibility of using P. putida to E. coli as an alternative host organism for industrial processes. P. putida KT2440 showed activity comparable to E. coli, indicating that this could be a suitable organism for use in industry. Nineteen omega-transaminases were screened to select a transaminase which was able to convert 2-HMSA to a novel amine. Results showed that (R) - selective transaminase appeared to have activity with 2-HMSA, with an arthrobacter mutant transaminase (ArRMut11) showing the highest conversion rates. The conversion of modified starting materials, p-toluic acid, m-toluic acid and 2,3- dimethylbenzoate were investigated with the engineered pathway. Identifying the amine products of the transaminase conversion of 2-HMSA and the modified starting substrates was attempted using a combination of HPLC and mass spectrometry, however the exact structures were not confirmed.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available