Notably, although His and Glu have low distribution percentages (see orange bars in Figure ), we consider both residues, along with Tyr, as potential proton acceptors. This is because His is already a well-accepted proton acceptor in other metal containing CA, while Glu498 is relevant due to our MD trajectory showing it associates with CO2 via a water chain, despite being positioned at a considerable distance of 13.6 Å from CO2 (cf. Figure ). A systematic study of CO2 hydration, focusing on these three residues (His, Glu, and Tyr), is discussed in the subsequent sections. interact with CO2, it is crucial to verify this using quantum chemical calculations. Accordingly, we performed potential energy surface (PES) scans employing hybrid QM/MM calculations for the hydration of CO2. For this transformation to occur using His584 and Glu498 as proton acceptor residues (cf. section S.3.a and S.3.b), we found that both require very high activation energy barriers of >32 kcal/mol and >35 kcal/mol (cf. Figure , S10), respectively, with producing an unstable product. A detailed discussion is provided in the SI (cf. S.3). As such, our QM/MM calculations corroborate the conclusions drawn from the MD simulations, thereby nullifying the possibility of His584 and Glu498 residues as potential proton acceptors in metal independent CA and leaving Tyr476 as the viable option.