Optimization of Poly(<i>N</i>‑isopropylacrylamide) as an Artificial Amidase

Abstract

Poly­(<i>N</i>-isopropylacrylamide) microgel (NMG) has been developed by adding various functional groups to control surface charges, hydrophobicity, p<i>K</i>_a and protein adsorption capacity. Here, we developed and optimized NMG anchored with three types of functional groups as a polymeric catalyst to hydrolyze amide bonds under optimized mild conditions. Various optimization strategies were evaluated for efficient hydrolysis activity on a <i>p</i>-nitroaniline-based substrate by using a colorimetric assay. Based on the results, we propose a mechanism to hydrolyze amide bonds and determine the theoretical average distance, using NMG bearing functional group of 1-vinylimidazole as the study model. The hydrolysis of amide bonds was inhibited by a transition-state protease inhibitor, which also confirmed the proposed reaction model for NMG. These results provide an insight into the strategies developed to functionalize hydrogels through an enzyme-mimic approach for future robust bio- and chemical conversions as well as therapeutic utilities