Bacterial bio-factories for the production of valuable chemicals

Grigoriou, Stylianos (2020) Bacterial bio-factories for the production of valuable chemicals. PhD thesis, University of Nottingham.

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Abstract

This thesis focuses on the development of novel enzymatic approaches for chiral amine synthesis utilizing amine transaminases (ATAs) and ‘smart’ amine donors. Different strategies were combined for the expansion of the existing biocatalytic toolbox, paving the way towards green alternatives for the enantioselective production of valuable chemicals.

In the first chapter, the wider application of the ‘smart’ amine donors is presented. Initially, a panel of 400 ketones/aldehydes was screened, using o-xylylenediamine as the amine donor, with a commercially available ATA. The colored precipitate was used as a positive indicator, to reveal a pattern for the carbonyl tolerance of the ATA. Subsequently, the commercial value of the corresponding chiral amines was evaluated with the assistance of the industrial partner Key Organics. The viable substrates were thoroughly investigated with different ‘smart’ diamine donors, in an effort to reduce the production costs. 1,5-Diaminopentane (cadaverine) was the most effective donor and was used for the preparation of five ketones on preparative scale, from which three became commercially available through Key Organics.

In the second chapter, the development of a novel whole-cell system, mediating ATA reaction in a self-sufficient manner, is presented. A selection of (S)- and (R)-selective ATAs were screened against ‘smart’ amine donors utilizing a panel of carbonyl-containing compounds. Five (S)-selective ATAs were identified and transformed in Corynebacterium glutamicum cells, which have been engineered to produce cadaverine and 1,4- diaminobutane (putrescine). Due to endogenous side-reactions and complications with expression, three ATAs were successfully expressed and utilized for the whole-cell transamination of a model ketone substrate. New strains with enhanced ATA production were developed and the reaction conditions were optimized to afford an enantiomerically pure (>99%) (S)- pyrroline with 42% isolated yield.

Item Type: Thesis (University of Nottingham only) (PhD)
Supervisors: O'Reilly, Elaine
Hill, Phil
King, John
Keywords: Chemical production, Chiral amine synthesis, Amine transaminases, ATAs, Smart amine donors
Subjects: Q Science > QD Chemistry > QD241 Organic chemistry
Faculties/Schools: UK Campuses > Faculty of Science > School of Chemistry
Item ID: 63935
Depositing User: Grigoriou, Stylianos
Date Deposited: 12 Jan 2021 15:14
Last Modified: 12 Jan 2021 15:15
URI: http://eprints.nottingham.ac.uk/id/eprint/63935

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