Intrinsic proton transfer activation of L-DOPA encoded carbonic anhydrase for efficient CO2 sequestration.

Carbonic anhydrase (CA), a zinc-bound carbon sequestrating metalloenzyme, is well-known for its catalytic function of interconverting carbon dioxide into bicarbonate: CO2 + H2O ↔ HCO3- + H+. The rate of hydration depends on intramolecular proton transfer within the active site and its surrounding cleft residues. The proton-transfer energy barrier requires certain hydrogen bond formation in the active site and cleft for water chain motion. Here, we genetically introduced metal binding catechol L-3,4-dihydroxyphenylalanine (L-DOPA), which formed hydrogen bonds with glycine and asparagine in the cleft region, while its additional hydroxyl groups facilitated the formation of new hydrogen bonds with an adjacent water molecule. This congener CA (rDCA) enhanced the intermediate stabilization and proton shuttling, leading to a more than twofold increase in substrate turnover and thermal stability and a more than threefold increase in catalytic efficiency compared to native CA (rCA). The formation of additional hydrogen bonds confirmed the accelerated carbon sequestration, highlighting rDCA as a promising catalyst for carbon capture and storage technologies.