Mechanism of NH₄⁺ Recruitment and NH₃ Transport in Rh Proteins

Summary In human cells, membrane proteins of the rhesus (Rh) family excrete ammonium and play a role in pH regulation. Based on high-resolution structures, Rh proteins are generally understood to act as NH₃ channels. Given that cell membranes are permeable to gases like NH₃, the role of such proteins remains a paradox. Using molecular and quantum mechanical calculations, we show that a crystallographically identified site in the RhCG pore actually recruits NH₄⁺, which is found in higher concentration and binds with higher affinity than NH₃, increasing the efficiency of the transport mechanism. A proton is transferred from NH₄⁺ to a signature histidine (the only moiety thermodynamically likely to accept a proton) followed by the diffusion of NH₃ down the pore. The excess proton is circulated back to the extracellular vestibule through a hydrogen bond network, which involves a highly conserved and functionally important aspartic acid, resulting in the net transport of NH₃.