Kidney Blood Press Res 2001 Jan 01;243:142-8. doi: 10.1159/000054221.

Influences of the N- and C-termini of the distal nephron inward rectifier, ROMK.

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The inward rectifying potassium channels of the ROMK family are present in the distal nephron of the kidney. These channels have two membrane spanning portions, between which lies a hydrophobic domain thought to confer the majority of the conductive properties of the channel. The N- and C-termini are both intracellular. In this paper we have examined the contribution of the N- and C-termini to the pore by examining the interaction of Cs+ with the channels. ROMK1 has an additional 19 amino acids on its N-terminus in comparison to ROMK2. The C-terminus of ROMK2 was extended by addition of a streptavidin tag (sfROMK2). Currents were measured following expression in Xenopus oocytes using two-electrode voltage clamp. ROMK1, ROMK2 and sfROMK2 exhibited concentration- and voltage-dependent block of inward currents by extracellular Cs+. The Hill coefficients were not significantly different from one. The mean Kd values at 0 mV were 100.6 +/- 10.6, 63.1 +/- 3.9 and 40.6 +/- 9.4, respectively (p < 0.05). The electric distances (delta) were 0.94 +/- 0.06, 1.0 +/- 0.05 and 1.37 +/- 0.06 respectively. The delta of sfROMK2 was greater than either ROMK1 or ROMK2 (p < 0.001). ROMK1, ROMK2 and sfROMK2 are sensitive to extracellular Cs+. Block was both concentration- and voltage-dependent. sfROMK2 is most Cs+-sensitive. ROMK1 contains an additional N-terminal 19 amino acids. Thus the pore properties of these two isoforms are subtly different, and influenced by the N-terminus. The lower Kd in sfROMK2 suggests that the streptavidin tag, and perhaps the C-terminus, also affect the pore.

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Species referenced: Xenopus
Genes referenced: kcnj1