Figure 3

Functional assays of purified human β-cardiac S1L construct co-purified with murine myosin light chains. (A) time courses of ATP turnover, using NADH-coupled assay, with constant [S1L-eGFP-FLAG] and [MgATP] but with [F-actin] (concentration relating to monomers) at either 5 (a), 10 (b), 15 (c), 20 (d), 30 (e), 40 (f), 60 (g), 80 (h), or 100 (i). (B) Actin-activated steady-state ATPase activity as a function of [F-actin]. The solid line through the data points is the best fit of Eq. (1) to the data with best-fit parameters presented in Table 1. Data from three independent protein preparations shown in different colors. (C) The sliding velocities of actin filaments in the in-vitro motility assay at 25 °C for two different protein preparations (Prep 1 and Prep 2). The data are superimposed on mean ± 95% CI from 25–30 filaments for each preparation. (D). The fraction of motile filaments after one (for Prep 1) or two (for Prep 2) affinity purifications (“deadheadings”). The data are superimposed on mean ± 95% CI from 5 fields of view per flow cell for each preparation. (E) Actin filament sliding velocities versus filament lengths (same data as in C for Prep 1). (F) Actin filament sliding velocities versus filament lengths (same data as in C for Prep 2). Straight horizontal lines in E and F indicate mean values suggesting minimal dependence of velocity on filament lengths at lengths > 3 µm.