Use this URL to cite or link to this record in EThOS: http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.727351
Title: Biocatalytic imine reduction : isolation, application, structure and mechanism of imine reductases
Author: Wells, Elizabeth
Awarding Body: University of York
Current Institution: University of York
Date of Award: 2017
Availability of Full Text:
Access from EThOS:
Full text unavailable from EThOS. Thesis embargoed until 03 Nov 2020
Access from Institution:
Abstract:
Chiral amines are industrially useful chemicals found in the pharmaceutical, chemical and agrochemical industries. While many excellent methods have been developed for their synthesis using abiotic catalysis, these are often reliant on precious metals the global supply of which is becoming increasingly scarce. Additionally, those catalysts often require harsh, non-environmentally friendly reaction conditions such as high temperatures and pressures. Biocatalytic imine reduction, which involves the asymmetric reduction of imines to form chiral amines using imine reductases (IREDs), offers an efficient and sustainable synthesis of chiral amines which overcomes many of the limitations encountered in abiotic catalysis. Two NADPH-dependent IREDs were expressed and purified; SkR-IRED from Streptomyces kanamyceticus and SS-IRED from Streptomyces sp. GF3546, which catalyse the reduction of the model imine 2-methylpyrroline to (R)¬- and (S)- amine products respectively. The SkR¬-IRED monomer consists of an N-terminal Rossman fold motif and a C-terminal helical domain. SkR-IRED exists as a homodimer of two monomers which are linked by an unusual reciprocal domain sharing arrangement. The structure of SkR-IRED was used as a basis to study mechanism using mutagenesis experiments whichsuggested that residue Asp187 may be important for catalysis. The structure of SS-IRED was solved to a resolution of 3.2 Å in its apo form and revealed that the enzyme shares the structural features of SkR-IRED. Mutagenesis experiments suggested that residue Tyr169 is necessary for catalysis. Last, a novel IRED from the moss Physcomitrella patens (Pp-IRED) was expressed and purified. Pp-IRED was active towards the amine N-methyl-1-quinolin-6-ylmethanamine in the oxidative direction, the first IRED active towards this substrate. The structure of Pp-IRED was solved in both its apo form and in complex with NADPH (2.5 Å resolution). The structure was distinct from other IREDs as it did not display the reciprocal domain sharing arrangement seen in SkR-IRED and SS-IRED.
Supervisor: Grogan, Gideon Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID: uk.bl.ethos.727351  DOI: Not available
Share: