UC Berkeley

The explosion of protein diversity through domain rearrangements and inter-domain coupling supported the evolution of multicellular organisms. To perform the advanced signaling necessary for multicellularity, sometimes unrelated protein domains combined to form novel domain architectures that over time evolved tight mechanisms of allosteric coupling. One such protein, the voltage sensing phosphatase (VSP), developed a sophisticated mechanism of inter-domain coupling, which enabled cells to integrate changes in membrane potential into chemical changes in a class of secondary signaling lipids called phosphatidylinositol-phosphates (PIPs).

Phosphoinositol phosphate signaling lipids (PIPs) are important second messengers that regulate ion channels, transporters, cell motility and endo/exocytosis. PIP concentrations are controlled by enzymes, including VSP, which has broad specificity for a diverse class of PIPs. VSP is a novel lipid phosphatase, which contains a voltage sensing domain (VSD) homologous to voltage-gated ion channels, and a lipid phosphatase domain (PD). Until now it was not known what properties of the cytosolic PD were allosterically regulated by the membrane-associated VSD. Using a pair of new PIP sensors to monitor enzyme activity and voltage clamp fluorometry to monitor conformational changes in the VSD, it becomes clear the Ciona intestinalis VSP (Ci-VSP) has two distinct voltage regulated enzyme active states: a faster low-voltage state with substrate preference for PIP3 and a slower high-voltage state with preference for PIP2. This novel 2-step allosteric switch for enzyme specificity enables membrane potential to function as an allosteric effector that dynamically regulates PIP concentration.

In this work, it is show that two unrelated domains, a VSD from voltage dependent ion channels and a lipid PD homologous to protein tyrosine phosphatases evolved a tight mechanism of allosteric regulation that transduces fluctuations in membrane potential into changes in the enzyme selectivity of a novel lipid phosphatase. This regulation of active site specificity in the PD by an allosteric effector domain represents a significant advancement in our understanding of allosteric regulation, which has previously been restricted to control of activity on only one type of substrate.