Figure ) highlights its transient movement within the enzyme environment. All these observation indicate that, after its formation, HCO3 -cannot remain in any zone long enough to facilitate reverse catalysis. Consequently, our results support experimental findings suggesting that the dehydration of of HCO3 -is not achievable in metal-free CA. Nevertheless, the brief entry of HCO3 -into the hydrophilic zone aligns with the crystal structure, where HCO3 -was found co-crystallized in that zone. Thus, our simulations clarify how HCO3 -migrates from the hydrophobic to the hydrophilic zone after its formation and provide an explanation for its inability to perform the reversal of CO2 hydration.