Functional Catalysis by Heterochiral Peptides Designed to Mimic Carbonic Anhydrase for Carbon Dioxide Sequestration

Enzymes, as natural catalysts, play an essential role in catalyzing diverse biological reactions with high efficiency and selectivity. Mimicking enzyme active sites with short peptide sequences is a challenging but valuable scientific endeavor. In this study, we explore the design of heterochiral heptapeptides comprising alternating l- and d-amino acids to generate structurally stable and functional mimics of human carbonic anhydrase II (hCAII), a key enzyme for carbon dioxide hydration, with the aim of carbon capture and sequestration. Four Zn2+-coordinated heptapeptides, Ac-HyHyHfF-CONH2 (KK1), Ac-HPhYhFf-CONH2 (KK2), Ac-HhHfFyF-CONH2 (KK3), and Ac-HhYfHfF-CONH2 (KK4), were synthesized and structurally validated by 2D NMR, confirming the formation of organized conformations. Among these, KK1 exhibited the highest catalytic performance toward p-nitrophenyl acetate (p-NPA) hydrolysis, with a catalytic efficiency of 71.67 ± 1.64 × 10–2 M–1 s–1. KK2 and KK3 showed moderate activity, whereas KK4 was catalytically inactive. The observed differences in catalytic activity correlate with peptide secondary structure and self-assembly behavior, where peptides forming ordered α-helical-like nanostructures demonstrate enhanced activity due to favorable Zn2+ coordination and catalytic pocket formation. The peptide catalysts displayed increased activity at higher pH and temperatures, maintaining functionality up to 80 °C, in contrast to hCAII, which denatures above 50 °C. Furthermore, CO2 sequestration experiments confirmed catalytic activity, with KK1 showing the highest efficiency in bicarbonate formation and subsequent CaCO3 precipitation. Although the catalytic efficiency of these peptides remains lower than that of native hCAII, their significantly smaller size (∼1–1.1 kDa) and ability to function under non-physiological conditions highlight their potential as minimalist and robust catalytic systems. Additionally, cytotoxicity studies using HEK-293 cells demonstrated over 80% cell viability, confirming their biocompatibility. This study establishes heterochiral peptide design as a promising strategy for developing stable, tunable, and biocompatible catalysts for CO2 conversion and sequestration applications.