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Directed evolution of prenylated FMN-dependent Fdc supports efficient in vivo isobutene production

DOI: 10.1038/s41467-021-25598-0 DOI Help

Authors: Annica Saaret (University of Manchester) , Benoît Villiers (Global Bioenergies) , François Stricher (Global Bioenergies) , Macha Anissimova (Global Bioenergies) , Mélodie Cadillon (Global Bioenergies) , Reynard Spiess (University of Manchester) , Sam Hay (University of Manchester) , David Leys (University of Manchester)
Co-authored by industrial partner: Yes

Type: Journal Paper
Journal: Nature Communications , VOL 12

State: Published (Approved)
Published: September 2021
Diamond Proposal Number(s): 12788 , 17773

Open Access Open Access

Abstract: Isobutene is a high value gaseous alkene used as fuel additive and a chemical building block. As an alternative to fossil fuel derived isobutene, we here develop a modified mevalonate pathway for the production of isobutene from glucose in vivo. The final step in the pathway consists of the decarboxylation of 3-methylcrotonic acid, catalysed by an evolved ferulic acid decarboxylase (Fdc) enzyme. Fdc belongs to the prFMN-dependent UbiD enzyme family that catalyses reversible decarboxylation of (hetero)aromatic acids or acrylic acids with extended conjugation. Following a screen of an Fdc library for inherent 3-methylcrotonic acid decarboxylase activity, directed evolution yields variants with up to an 80-fold increase in activity. Crystal structures of the evolved variants reveal that changes in the substrate binding pocket are responsible for increased selectivity. Solution and computational studies suggest that isobutene cycloelimination is rate limiting and strictly dependent on presence of the 3-methyl group.

Journal Keywords: Biocatalysis; Biofuels; Metabolic engineering

Diamond Keywords: Biofuel; Enzymes

Subject Areas: Chemistry, Biology and Bio-materials, Energy


Instruments: I03-Macromolecular Crystallography , I04-1-Macromolecular Crystallography (fixed wavelength) , I04-Macromolecular Crystallography , I24-Microfocus Macromolecular Crystallography

Added On: 18/09/2021 11:29

Documents:
s41467-021-25598-0-2.pdf

Discipline Tags:

Catalysis Organic Chemistry Earth Sciences & Environment Climate Change Energy Bioenergy Sustainable Energy Systems Engineering & Technology Biotechnology Life Sciences & Biotech Structural biology Chemistry Biochemistry

Technical Tags:

Diffraction Macromolecular Crystallography (MX)