Year: 2026
Type: preprint
Abstract: Enzymes are unparalleled catalysts that combine high activity and selectivity with operation under mild conditions. Despite these advantages, their broader use as industrial biocatalysts is limited by insufficient stability under process-relevant conditions and by the difficulty of achieving site-specific chemical functionalization. Simultaneously enhancing protein stability and enabling site-specific functionalization through a single chemical strategy remains a central challenge, particularly, when addressing multimeric enzymes. Here, we show that a four-armed crosslinker with tetrahedral symmetry enables simultaneous stabilization and programmable functionalization of the tetrameric enzyme borneol dehydrogenase. A twostep crosslinking–modification reaction yields a well-defined and stable protein quaternary architecture with an incorporated reactive handle, enabling the installation of diverse functionalities, including bioorthogonal tags and functional groups for immobilization. The chemically engineered bicyclic enzyme exhibits remarkably enhanced thermal stability and resistance to organic cosolvents, which results in exceptional longevity supporting ten weeks of continuous flow operation. Unlike previous methods that decouple stabilization from modification, this strategy unites structural reinforcement and functionalization, thereby expanding the scope of chemical protein engineering.
Source: ChemRxiv
Institutions: Vrije Universiteit Amsterdam, University of Göttingen
Language: English
Subfield: Molecular Biology
Domain: Life Sciences
Open Access status: green
OpenAlex ID: W7204902528