The influenza A virus M2 proton channel, is the target of anti-flu medications, amantadine and rimantadine. The functions of M2 is closely related to its conformation changes. While extensive progresses have been made regarding the structure-function relationship for the transmembrane helix, there have been much fewer studies for the other membrane associated segment, the C-terminal amphiphilic helix. We carried out FRET experiments by introducing into the amphiphilic helix an unnatural amino acid, PheCN, as the donor for the W residue located in the transmembrane helix. This allowed us to study the global conformation change of these membrane associated structures under channel activation or drug inhibition. The distance between amphipathic helix and transmembrane helix increased upon channel activation in the acidic environment. The degree of this distance increase was not affected by drug inhibition. Therefore we speculate that the conformation change of amphipathic helix is not related to the proton channel activity, but is more likely associated with the protein’s role in virus budding.