Permeation mechanisms of hydrogen peroxide and water through Plasma Membrane Intrinsic Protein aquaporins

Hydrogen peroxide (H₂O₂) transport by aquaporins (AQP) is a critical feature for cellular redox signaling. However, the H₂O₂ permeation mechanism through these channels remains poorly understood. Through functional assays, two Plasma membrane Intrinsic Protein (PIP) AQP from Medicago truncatula, MtPIP2;2 and MtPIP2;3 have been identified as pH-gated channels capable of facilitating the permeation of both water (H₂O) and H₂O₂. Employing a combination of unbiased and enhanced sampling molecular dynamics simulations, we investigated the key barriers and translocation mechanisms governing H₂O₂ permeation through these AQP in both open and closed conformational states. Our findings reveal that both H₂O and H₂O₂ encounter their primary permeation barrier within the selectivity filter (SF) region of MtPIP2;3. In addition to the SF barrier, a second energetic barrier at the NPA (asparagine-proline-alanine) region that is more restrictive for the passage of H₂O₂ than for H₂O, was found. This behavior can be attributed to a dissimilar geometric arrangement and hydrogen bonding profile between both molecules in this area. Collectively, these findings suggest mechanistic heterogeneity in H₂O and H₂O₂ permeation through PIPs.