Phosphatase activity of the voltage-sensing phosphatase, VSP, shows graded dependence on the extent of activation of the voltage sensor

Key points The voltage-sensing phosphatase (VSP) consists of the voltage sensor and the phosphatase domain. The voltage sensor movement is coupled to the phosphatase activity. To uncover the coupling mechanisms between the two domains, we made a voltage sensor mutant of VSP. Our analyses showed that this voltage sensor moves in two steps. Measurements revealed that the phosphatase activity of this mutant is associated with both the first and second step of the voltage sensor movements. Results suggest that the phosphatase activity of VSP shows graded dependence on the extent of activation of the voltage sensor. AbstractThe voltage-sensing phosphatase (VSP) consists of a voltage sensor and a cytoplasmic phosphatase region, and the movement of the voltage sensor is coupled to the phosphatase activity. However, its coupling mechanisms still remain unclear. One possible scenario is that the phosphatase is activated only when the voltage sensor is in a fully activated state. Alternatively, the enzymatic activity of single VSP proteins could be graded in distinct activated states of the voltage sensor, and partial activation of the voltage sensor could lead to partial activation of the phosphatase. To distinguish between these two possibilities, we studied a voltage sensor mutant of zebrafish VSP, where the voltage sensor moves in two steps as evidenced by analyses of charge movements of the voltage sensor and voltage clamp fluorometry. Measurements of the phosphatase activity toward phosphatidylinositol 4,5-bisphosphate revealed that both steps of voltage sensor activation are coupled to the tuning of phosphatase activities, consistent with the idea that the phosphatase activity is graded by the magnitude of the movement of the voltage sensor.