High-Affinity Copolymers Inhibit Digestive Enzymes by Surface Recognition

Abstract

This account presents a general method for the construction of polymeric surface binders for digestion enzymes. Two prominent parts, namely, the modification of the copolymer composition and the screening assay for the most powerful inhibitors are both amenable to parallelization. The concept hinges on the appropriate selection of amino-acid-selective comonomers, their free radical copolymerization, and subsequent screening of the resulting copolymer library for efficient enzyme inhibition. A microscale synthetic procedure for the copolymerization process was developed, which produces water-soluble affinity polymers that can be stored for years at room temperature. Initial parallel screening was conducted in standard enzyme assays to identify polymeric inhibitors, which were subsequently subjected to determination of IC values for their target enzyme. For all digestion enzymes, except elastase, a number of polymer inhibitors were found, some of which were selective toward one or two protein targets. Since the key monomers of the best inhibitors bind to amino acid residues in the direct vicinity of the active site, we conclude that efficient coverage of the immediate environment by the copolymers is critical. Strong interference with enzymatic activity is brought about by blocking the substrate access and product exit to and from the active site.

Bioactive Synthetic Polymers

“…In these works, the authors demonstrated that screening large numbers of copolymers synthesized by free‐radical polymerization was a viable strategy for identifying strong binding interactions between synthetic polymers and immunoglobulins, [ 40 ] or digestive enzymes. [ 41 ]…”

“…In these works, the authors demonstrated that screening large numbers of copolymers synthesized by free-radical polymerization was a viable strategy for identifying strong binding interactions between synthetic polymers and immunoglobulins, [40] or digestive enzymes. [41] In another complementary strategy for synthetic polymerprotein binding, Xu and co-workers analyzed the dispersity of amino acid residues on the surfaces of folded proteins to provide design criteria for synthetic heteropolymers. [42] Interestingly, computational analysis of model proteins such as horseradish peroxidase suggested many small patches (1-2 nm) of neutral hydrophilic, hydrophobic, positively, or negatively charged groups on the folded protein surfaces, with interpatch distances of 1-2 nm.…”

Design of Functional Nanoparticles for Intractable Disease Therapy

“…Additionally, typically, PPIs are larger than 1000 Å 2 and involve more than 20 amino acid contacts. 3,4) Synthetic polymers, such as dendrimers, 5) linear polymers, [6][7][8] and polymer nanoparticles (NPs), 9,10) have the potential to work as protein affinity reagents by mimicking protein-protein interactions 11) (Fig. 1b).…”

Associative and Dissociative Processes in Non‐Covalent Polymer‐Mediated Intracellular Protein Delivery

“…Non-covalent polymer-protein binding studies, sometimes involvingd elivery experiments, is ab road field, and additional reports relatedt ot his topic that highlight various binding interactions and characterization methods can be found in Refs. [58][59][60][61][62][63][64][65][66][67][68][69][70][71].B inding studies showcasing availablet echniques could potentially contain criticali nformation necessary for future protein-delivery optimization studies and should not be overlooked.…”