Biological catalysis regulated by cucurbit[7]uril molecular containers

Area of Science:

• Supramolecular Chemistry
• Enzyme Inhibition
• Chemical Biology

Background:

• Enzyme activity modulation is crucial for biological processes and therapeutic interventions.
• Cucurbiturils (CBs) are macrocyclic hosts capable of forming stable inclusion complexes with various guests.
• Developing controllable enzyme inhibitors remains a significant challenge in chemical biology.

Purpose of the Study:

• To synthesize and characterize novel 'two-faced' inhibitors possessing both enzyme-binding and cucurbituril-binding domains.
• To investigate the controllable inhibition of bovine carbonic anhydrase (BCA) and acetylcholinesterase (AChE) using these inhibitors and CB[7].
• To elucidate the mechanism of reversible and irreversible enzyme inhibition through supramolecular complex formation.

Main Methods:

• Synthesis of bifunctional molecules with enzyme inhibitor and cucurbituril recognition sites.
• Enzymatic activity assays (UV/Vis) to monitor catalytic rates of BCA and AChE.
• Fluorescence displacement assays, 1H NMR spectroscopy, and kinetic measurements (Ka, kon, koff) to characterize complex formation and dissociation.

Main Results:

• Successfully synthesized two-faced inhibitors 1-5 targeting BCA and AChE.
• Demonstrated reversible on-off switching of BCA activity via transient BCA-CB[7] ternary complex formation and dissociation.
• Achieved stable, irreversible inhibition of AChE through the formation of thermodynamically stable AChE-CB[7] ternary complexes.

Conclusions:

• The 'two-faced' inhibitor strategy enables tunable control over enzyme catalytic activity.
• Reversible inhibition was achieved with BCA, while irreversible inhibition was observed with AChE, depending on complex stability.
• This approach offers a distinct mechanism for enzyme activity control compared to natural allosteric regulation.