Movement of NH₃ through the human urea transporter B: a new gas channel

Aquaporins and Rh proteins can function as gas (CO₂ and NH₃) channels. The present study explores the urea, H₂O, CO₂, and NH₃ permeability of the human urea transporter B (UT-B) (SLC14A1), expressed in Xenopus oocytes. We monitored urea uptake using [¹⁴C]urea and measured osmotic water permeability (Pf) using video microscopy. To obtain a semiquantitative measure of gas permeability, we used microelectrodes to record the maximum transient change in surface pH (ΔpHS) caused by exposing oocytes to 5% CO₂/33 mM HCO₃⁻ (pHS increase) or 0.5 mM NH₃/NH₄⁺ (pHS decrease). UT-B expression increased oocyte permeability to urea by >20-fold, and Pf by 8-fold vs. H₂O-injected control oocytes. UT-B expression had no effect on the CO₂-induced ΔpHS but doubled the NH₃-induced ΔpHS. Phloretin reduced UT-B-dependent urea uptake (Jurea*) by 45%, Pf* by 50%, and (- ΔpHS*)NH₃ by 70%. p-Chloromercuribenzene sulfonate reduced Jurea* by 25%, Pf* by 30%, and (ΔpHS*)NH₃ by 100%. Molecular dynamics (MD) simulations of membrane-embedded models of UT-B identified the monomeric UT-B pores as the main conduction pathway for both H₂O and NH₃ and characterized the energetics associated with permeation of these species through the channel. Mutating each of two conserved threonines lining the monomeric urea pores reduced H₂O and NH₃ permeability. Our data confirm that UT-B has significant H₂O permeability and for the first time demonstrate significant NH₃ permeability. Thus the UTs become the third family of gas channels. Inhibitor and mutagenesis studies and results of MD simulations suggest that NH₃ and H₂O pass through the three monomeric urea channels in UT-B.