Exploiting cofactor-dependent oxidoreductases by enzyme discovery, engineering and reconstruction

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Abstract

Being among the most common cofactors, flavins and nicotinamides are employed by many oxidoreductases to enable their function in metabolic pathways of organisms. The crucial roles of these cofactor-dependent enzymes can also be deduced from the fact that they account for about one third of all enzyme activities1. A number of these enzymes are nowadays used for the production of high value compounds in modern society. When considering enzymes as biocatalysts for use in synthetic chemistry, they are typically appreciated for their high activity, high selectivity while only mild reaction condition can be used. Therefore, they are especially suitable for the manufacture of fine chemicals, pharmaceuticals and food additives, where high purity, chemo- and enantioselectivity and low hazards are called for. So, biocatalytic processes are often considered as green and efficient ways to replace legacy synthetic chemistry reactions especially for asymmetric reactions.
Clearly, enzymes have huge potential as biocatalysts for industrial applications. To satisfy the requirement for suitable biocatalysts, methods are continuously being developed and improved to generate robust and efficient enzymes. Here in this thesis, we applied different methods to study and explore the potential of cofactor-dependent oxidoreductases:
1. New deazaflavin-dependent flavin-containing thioredoxin reductases (DFTRs) were discovered to broaden the repertoire of this type of biocatalysts.
2. Protein engineering was used to modify F420:NADP+ oxidoreductases (FNOs) to establish a regeneration system of nicotinamide cofactors biomimetics.
3. Ancestral sequence reconstruction (ASR) was used to elucidate the evolutionary history of flavin-containing monooxygenases (FMOs).
4. ASR was also used to reconstruct how Baeyer-Villiger monooxygenases (BVMOs) attained their unique ability to perform B-V oxidations.
Original languageEnglish
QualificationDoctor of Philosophy
Awarding Institution
  • University of Groningen
Supervisors/Advisors
  • Fraaije, Marco, Supervisor
  • Slotboom, Dirk, Supervisor
Award date5-Dec-2023
Place of Publication[Groningen]
Publisher
DOIs
Publication statusPublished - 2023

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