Model of the %IHM/SRX impact on β-cardiac myosin Velocity Index. (a) Velocity Index versus surface occupancy data for 2HEP WT were fit to the modified Hill equation (solid line) (Supplemental text) and used to represent an β-cardiac myosin population with 16% SRX that allowed a family of curves to be generated for hypothetical IHM/SRX percentages ranging from 0 to 100% (b). The 95% confidence limits (gray shaded area) for the 2HEP WT fit were used to estimate the error in the model predicted %IHM/SRX for a given construct. All fits were performed by calculating the minimum COST function from the least squared error analysis (example for 2HEP WT, inset). For 2HEP WT, the model best fit returns a best fit of 14% IHM/SRX with an estimate error of ±13% IHM/SRX. Velocity Index versus surface occupancy data for 15HEP WT with model predicted 69% IHM/SRX curve and 95% confidence limits (gray shaded area) error estimates of ±6% IHM/SRX. Predicted %IHM/SRX and the errors in the estimate for all constructs were as follows: WT: 2HEP (14 ± 13%), 15HEP (69 ± 6%), 25HEP (76 ± 4%); E525K: 2HEP (16 ± 12%), 15HEP (62 ± 6%), and 25HEP (71 ± 4%). (b) Family of curves for hypothetical %IHM/SRX ranging from 0 to 100% based on the original WT 2HEP modified Hill dose–response curve in a, which assumed a 16%IHM/SRX (see Supplemental text for details).