Optically detected structural change in the N-terminal region of the voltage-sensor domain.

The voltage-sensor domain (VSD) is a functional module that undergoes structural transitions in response to membrane potential changes and regulates its effectors, thereby playing a crucial role in amplifying and decoding membrane electrical signals. Ion-conductive pore and phosphoinositide phosphatase are the downstream effectors of voltage-gated channels and the voltage-sensing phosphatase, respectively. It is known that upon transition, the VSD generally acts on the region C-terminal to S4. However, whether the VSD also induces any structural changes in the N-terminal region of S1 has not been addressed directly. Here, we report the existence of such an N-terminal effect. We used two distinct optical reporters-one based on the Förster resonance energy transfer between a pair of fluorescent proteins, and the other based on fluorophore-labeled HaloTag-and studied the behavior of these reporters placed at the N-terminal end of the monomeric VSD derived from voltage-sensing phosphatase. We found that both of these reporters were affected by the VSD transition, generating voltage-dependent fluorescence readouts. We also observed that whereas the voltage dependencies of the N- and C-terminal effects appear to be tightly coupled, the local structural rearrangements reflect the way in which the VSD is loaded, demonstrating the flexible nature of the VSD.