Supramolecularly gated enzymes: On-demand activity switching via host-guest competition for precision tumor therapy.

Dynamic control over enzyme function has been of great interest in the field of enzyme engineering and disease treatment, however, achieving on-demand regulation of native enzymes in living systems remains challenging due to the complexity of cellular environments and the lack of selective interactions. Here, we introduce a novel enzyme activity regulation system based on supramolecular host-guest recognition that enables reversible and on-demand regulation of native enzyme activity in living cells without covalent modification. This system utilizes the high-affinity interaction between cucurbit[7]uril (CB[7]) and amantadine (Ad) to precisely control enzyme function. CB[7] binds to key residues on the enzyme surface, blocking the active site and switching off enzyme activity. Upon intracellular administration of Ad, CB[7] is competitively removed, restoring enzyme activity and allowing for dynamic regulation of cellular functions. This strategy was successfully validated with model enzymes, including catalase (CAT) and β-galactosidase (β-Gal), demonstrating its broad applicability. Furthermore, we extended this approach to enzyme/prodrug therapy, achieving localized prodrug activation and targeted cell death while minimizing off-target effects and systemic toxicity. Thus, this non-covalent, reversible regulation of enzyme activity represents a significant advancement over traditional methods, offering a versatile and biocompatible tool for studying protein function and developing targeted therapeutic strategies.