Co-assembly of KCNQ1 a-subunits with KCNE1 b-subunits generates the I_Ks current implicated in the cardiac repolarization reserve. Both the S6 domain and the C-terminus of KCNQ1 are required for the modulation of KCNQ1 by KCNE1. To investigate the involvement of the cytoplasmatic end of S6 in the gating process of KCNQ1 and in the interaction with KCNE1, we performed scanning mutagenesis of residues 346-362 in KCNQ1. Substituting residues F351, A352, V355, Q356 and Q359 by alanine or tryptophan resulted in channels with altered gating properties compared to WT. These residues cluster on the same side on a a-helical wheel representation. The residues located on the opposite side showed no effect. However, substitutions of K354 and Q357, which did not modify KCNQ1 gating, impacted on the I_Ks current upon co-expression with KCNE1. These results favor an a-helical configuration of the S6 tail region, with one side presumably interacting with S4 and/or S5 thereby affecting the KCNQ1 gating, while the opposite side interacts with KCNE1.