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Published on: June 20, 2019
Mechanistic Analysis of an Engineered Enzyme that Catalyzes the Formose Reaction
Sean Poust1,2, James Piety1,2, Arren Bar-Even3
1Department of Chemical and Biomolecular Engineering, University of California, 201 Gilman Hall, Berkeley, CA 94720-1462 (USA).
Abstract:
An enzyme that catalyzes the formose reaction, termed "formolase", was recently engineered through a combination of computational protein design and directed evolution. We have investigated the kinetic role of the computationally designed residues and further characterized the enzyme's product profile. Kinetic studies illustrated that the computationally designed mutations were synergistic in their contributions towards enhancing activity. Mass spectrometry revealed that the engineered enzyme produces two products of the formose reaction-dihydroxyacetone and glycolaldehyde-with the product profile dependent on the formaldehyde concentration. We further explored the effects of this product profile on the thermodynamics and yield of the overall carbon assimilation from the formolase pathway to help guide future efforts to engineer this pathway.
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