Inserting unstructured peptides between the S4–S5 linker and S5–S6 core domain does not much affect force activation. (A) A diagram and recording from 374-GGS are presented as in Fig. 3. This insertion shortens the open duration and thus lowers basal activities, which is evident when the activity at 10⁻³ M Ca²⁺ here and that of the wild type at the same [Ca²⁺] shown in Fig. 3 A are compared. (B) An over fivefold increase in nP_o observed in both the wild type and 374-12GGS upon 150-mmHg pressure. Because the saturation level could not be determined before lytic pressure even at high [Ca²⁺] because of its shortened open durations and less stable seal under high pressures compared with wild type, the quantification was conducted using the fold increase of nP_o stimulated by 150-mmHg pressure from comparable basal activity between 374-12GGS and wild type at different [Ca²⁺] (10⁻³ M for 374-12GGS and 10⁻⁵ M for wild type). No clear difference in mechanosensitivity can be discerned by this measure. P = 0.35. Means ± SD (n = 3 for both).