Deciphering the evolutionary origin of the enantioselectivity of short-chain dehydrogenases from plants toward 1-borneol
Abstract Enzyme engineering has produced numerous methods to optimize enzymes for biotechnological processes; however, less is known about how natural evolution creates new functionalities. We investigate the evolutionary emergence of enantioselectivity in plant borneol dehydrogenases (BDHs), which feature hydrophobic active-sites and are enantioselective towards dibornane-type monoterpenols. Ancestral sequence reconstruction provided a trajectory from the oldest unselective BDH ancestor N30 (E=12) toward a more recent selective ancestor N32, involving 19 mutations: 18 mutations are peripheral, one (I111L) occurs in the active-site. The mutation L111I in the hydrophobic pocket increased the selectivity of N30, while the back-mutation I111L decreased the selectivity of N32. Additional peripheral mutations (V136L/G169A/V183I) were required for high selectivity. Crystal structures suggested that protein dynamics, rather than structural changes shape these catalytic properties; this was confirmed by ML/MM simulations of ligand binding. Funnel-metadynamics simulations revealed a correlation between the active-site’s solvent-accessible surface area (SASA) and selectivity. This potential evolutionary pathway shapes enantioselectivity, and guides future enzyme engineering campaigns.
Authors
- Ivana Drienovská (ORCID: https://orcid.org/0000-0003-1715-4236)
- Andrea M. Chánique (ORCID: https://orcid.org/0000-0003-0642-1351)
- Carl P. O. Helmer (ORCID: https://orcid.org/0000-0001-7195-9938)
- Robert Kourist (ORCID: https://orcid.org/0000-0002-2853-3525)
- Daniel Kracher (ORCID: https://orcid.org/0000-0002-3856-3170)
- Bernhard Loll (ORCID: https://orcid.org/0000-0001-7928-4488)
- Sebastian Brickel (ORCID: https://orcid.org/0000-0003-1972-0393)
- Shina Caroline Lynn Kamerlin (ORCID: https://orcid.org/0000-0002-3190-1173)
- Bruno Di Geronimo (ORCID: https://orcid.org/0000-0003-1822-7142)
- Jasmin Zuson (ORCID: https://orcid.org/0009-0001-9795-6652)
- Katarína Kavčiaková
- Rosa Teijeiro-Juiz (ORCID: https://orcid.org/0009-0000-3403-6755)
Institutions
- Uppsala University (SE)
- Institute of Molecular Biotechnology (AT)
- Georgia Institute of Technology (US)
- Georgia State University (US)
- Lund University (SE)
- Graz University of Technology (AT)
- BioTechMed-Graz (AT)
- Freie Universität Berlin (DE)
- Vrije Universiteit Amsterdam (NL)
Publication Details
- Journal
- Nature Communications
- Published
- 2026-08-14
- DOI
- https://doi.org/10.1038/s41467-026-76435-1
- Primary Topic
- Enzyme Catalysis and Immobilization
- Type
- article
- Field-Weighted Citation Impact
- 0.00
Funders
- Georgia Institute of Technology
- Georgia Research Alliance
- European Commission
- Deutsche Forschungsgemeinschaft
- Bundesministerium für Bildung und Forschung
- Austrian Science Fund
- Humboldt-Universität zu Berlin
- Freie Universität Berlin
- Hanns-Seidel-Stiftung
- BioTechMed-Graz
- Helmholtz-Zentrum Berlin für Materialien und Energie
- HORIZON EUROPE Framework Programme
- Deutsches Elektronen-Synchrotron
- Leibniz-Gemeinschaft