As a member of the ubiquitous ammonium transporter/methylamine permease/Rhesus (Amt/MEP/Rh) family of membrane protein channels, the 50 kDa Rhesus channel (Rh50) has been implicated in ammonia (NH3) and, more recently, also in carbon dioxide (CO2) transport. Here we present molecular dynamics simulations of spontaneous full permeation events of ammonia and carbon dioxide across Rh50 from Nitrosomonas europaea. The simulations show that Rh50 is functional in its crystallographic conformation, without the requirement for a major conformational change or the action of a protein partner. To assess the physiological relevance of NH3 and CO2 permeation across Rh50, we have computed potentials of mean force (PMFs) and permeabilities for NH3 and CO2 flux across Rh50 and compare them to permeation through a wide range of lipid membranes, either composed of pure lipids or composed of lipids plus an increasing cholesterol content. According to the PMFs, Rh50 is expected to enhance NH3 flux across dense membranes, such as membranes with a substantial cholesterol content. Although cholesterol reduces the intrinsic CO2 permeability of lipid membranes, the CO2 permeabilities of all membranes studied here are too high to allow significant Rh50-mediated CO2 flux. The increased barrier in the PMF for water permeation across Rh50 shows that Rh50 discriminates 40-fold between water and NH3. Thus, Rh50 channels complement aquaporins, allowing the cell to regulate water and NH3 flux independently. The PMFs for methylamine and NH3 are virtually identical, suggesting that methylamine provides an excellent model for NH3 in functional experiments.