Topology scheme of Ci-VSP.

(A) Topology scheme of Ci-VSP with the transmembrane voltage sensor (VSD) and the intracellular catalytic domain (CD), which comprises the phosphatase (PD) and the C2-domain. VSD and CD are connected via the linker sequence M240–K257. (B) Three-dimensional structure of the Ci-VSP CD based on the crystal structure by Liu et al. (PDB entry 3V0H) [14]. The P- (orange), the TI- (blue), and the WPD loop (yellow) form the active site of the CD. The linker (colored in red, with R253 shown as sticks) is oriented toward the TI loop, as proposed by Liu et al. [14]. (C) Amino acid alignment of Ci-VSP’s linker (M240–K257), active site (H362–R369), and the TI loop (R398–S414) with regions of the homolog PTEN (Hs, Homo sapiens). Amino acid identities are highlighted with a gray background. Putative interacting partners in the linker and the TI loop are denoted with blue and red asterisks, respectively. Single and multiple altered Ci-VSP mutants are aligned as well (in red letters: mutated positions). In the single neutralization mutants, the X stands for N (in case aspartate was the native residue) or for Q (in case of glutamate).