Conserved extracellular cysteine residues in the inwardly rectifying potassium channel Kir2.3 are required for function but not expression in the membrane

The mouse potassium channel Kir2,3 possesses conserved extracellular cystei ne residues at positions 113 and 145, We have investigated the role of thes e cysteines in structure/function and membrane trafficking. Cysteine to ser ine mutations resulted in the absence of potassium currents in oocytes and coexpression of these mutants with wild-type channel showed a dominant nega tive inhibition of wild-type currents. FLAG-tagged channels expressed in oo cytes were detected in the cell membrane by anti-FLAG antibody for wild-typ e and mutant channels, In vitro translation using the reticulocyte lysate s ystem showed that mutation of these residues did not affect processing nor insertion into membranes. Cysteine residues at 113 and 145 are therefore re quired for function of the Kir2,3 channel but not for processing into the c ell membrane; disulfide bonds between subunits are unlikely. (C) 1999 Feder ation of European Biochemical Societies.