Potassium channels are formed from four subunits, each of which contributes to the K+-selective pore. In mammals, the inwardly-rectifying (Kir) potassium channel subunits Kir4.1 and Kir5.1 co-assemble to form novel pH-sensitive heteromeric channels which show unique single channel bursting behaviour and subconductance states. In this study, we have identified orthologs of Kir4.1 and Kir5.1 from Xenopus tropicalis (XT) where the XTKir4.1 shares >90% identity with rat Kir4.1, and XTKir5.1 an overall 63% identity with ratKir5.1, which increases to >80% if the more divergent distal termini are excluded. Electrophysiological analysis of co-expressed XTKir4.1/XTKir5.1 channels revealed them to have an almost identical pH-sensitivity to the mammalian channels, but that their rate of time-dependent activation at hyperpolarising potentials was markedly different. However, the principal difference is in their kinetic behaviour. We found that the subconductance states in XTKir4.1-XTKir5.1 heteromeric channels have a much longer duration. In particular, they have a long S1 sublevel opening which is almost 15 times of the average dwell time duration of the S1 sublevels observed in the rat Kir4.1-Kir5.1 channels (7.5ms vs 0.5ms). We then created a ratKir4.1-XTKir5.1 dimer in order to assess whether this difference is conferred by the XTKir5.1 subunit. We found that ratKir4.1-XTKir5.1 heteromeric channels also exhibit these markedly long S1 sublevels, thus confirming that the XTKir5.1 subunit is responsible for this difference. These novel channels will therefore provide a greater insight into the mechanism of K+ channel gating at the single molecule level and help in the identification of the domains and/or residue(s) responsible for controlling the transitions between these states. This will have a major impact on our understanding of the structural basis of the sublevel transitions which occur during channel opening in this and other K+ channels.