Engineered protein-small molecule conjugates empower selective enzyme inhibition

Area of Science:

• Biochemistry
• Molecular Biology
• Drug Discovery

Background:

• Potent and specific ligands are crucial for precision medicine and fundamental biological research.
• Current ligand classes (proteins, peptides, small molecules) have limitations in achieving desired biological activity against complex targets.
• Greater molecular diversity is needed to develop ligands for challenging targets.

Purpose of the Study:

• To demonstrate a platform for discovering protein-small molecule (PriSM) hybrids.
• To combine the unique activities and shapes of small molecules with the engineerability of proteins.
• To create novel ligands with enhanced properties for biological applications.

Main Methods:

• A fibronectin protein library was displayed on yeast.
• A small molecule, acetazolamide, was conjugated to the protein via a maleimide-poly(ethylene glycol) linker.
• Magnetic and flow cytometry were used to sort and enrich specific binders to carbonic anhydrase isoforms.

Main Results:

• Isolated PriSMs demonstrated potent and specific inhibition of carbonic anhydrase isoforms.
• PriSMs showed superior efficacy compared to acetazolamide or the protein alone.
• An 80-fold increase in specificity and a 9-fold increase in potency were observed.
• The platform demonstrated flexibility through engineering various linker lengths, conjugation sites, and sequences.

Conclusions:

• The protein-small molecule (PriSM) hybrid platform significantly advances molecular diversity for engineerable ligands.
• PriSMs offer a promising approach to developing highly potent and specific inhibitors for therapeutic and research applications.
• The study highlights the potential of PriSMs to overcome limitations of traditional ligand classes.