The molecular mechanism of ion channel gating remains unclear. Using approaches such as proline scanning mutagenesis and homology modeling, we localize the gate of the K+ channels controlled by the βγ subunits of G proteins at the pore-lining bundle crossing of the second transmembrane (TM2) helices. We show that the flexibility afforded by a highly conserved glycine residue in the middle of TM2 is crucial for channel gating. In contrast, flexibility introduced immediately below the gate disrupts gating. We propose that the force produced by channel-Gβγ interactions is transduced through the rigid region below the helix bundle crossing to bend TM2 at the glycine that serves as a hinge and open the gate.