Molecular determinants for the distinct pH sensitivity of Kir1.1 and Kir4.1 channels

Kir1.1 (ROMK1) is inhibited by hypercapnia and intracellular acidosis with midpoint pH for channel inhibition (pK(a)) of similar to 6.7. Another close relative, Kir4.1 (BIR10), is also pH sensitive with much lower pH sensitivity (pK(a) similar to 6.0), although it shares a high sequence homology with Kir1.1. To find the molecular determinants for the distinct pH sensitivity, we studied the structure-functional relationship using site-directed mutagenesis. An NH2-terminal residue (Lys-53) was found to be responsible for the low pH sensitivity in Kir4.1. Mutation of this lysine to valine (K53V), a residue seen at the same position in Kir1.1, markedly increased channel sensitivity to CO2/pH. Reverse mutation on Kir1.1 (V66K) decreased the CO2/pH sensitivities. Interestingly, mutation of these residues to glutamate greatly enhanced the pH sensitivity in both channels. Other contributors to the distinct pH sensitivity were histidine residues in the COOH terminus, whose numbers are fewer in Kir4.1 than Kir1.1. Mutation of two of these histidine residues in Kir1.1 (H342Q/H354N) reduced CO2/pH sensitivities, whereas the creation of two histidines (S328H/G340H) in Kir4.1 increased the CO2/pH sensitivities. Combined mutations of the lysine and histidine residues in Kir4.1 (K53V/S328H/G340H) gave rise to a channel that had CO2/pH sensitivities almost identical to those of the wild-type Kir1.1. Thus the residues demonstrated in our current studies are likely the molecular basis for the distinct pH sensitivity between Kir1.1 and Kir4.1.