Fig. 4: Comparison of experimental in vitro assembly data to a simple equilibrium model.
For all panels, experimental data are plotted in green (assembled I53-40), purple (assembled I53-50), and orange (unassembled components). For all plots, error bars are standard deviations of three assembly replicates. a Quantitative analysis of equimolar assemblies are represented as Hill plots (Θ, assembly efficiency; L, free components). Hill coefficients are equal to the slopes of dashed regression lines, whose x-intercepts (dotted drop-lines with arrows) approximate the apparent KDs of assembly. Partitioning into high and low MW species is plotted in 3D space for b I53-40 and c I53-50 assemblies as a function of input [component]. This inclusive equilibrium model considers several nearly complete structures (black surface scatters, see “Methods”). Likewise, the smallest on-pathway species are summed with [T] and [P] (silver surface scatters). Gray lines emphasize output of the inclusive model at equimolar concentrations. d, e The same data as above, with the simplified equilibrium model in the 2D [T]0 = [P]0 plane. Only complete 120-subunit architectures (black) and free components (silver) are considered, and therefore plotted on axes where T = P. Solid lines represent the model output at the indicated best fit ΔGcon and dashed lines at best fit ±0.25 kcal/mol. Insets highlight the agreement between data and model at low [T]0 = [P]0. Cooperativity analyses for f, g I53-40 and h, i I53-50 report average peak area as a function of [variable component]. Raw signals are normalized to equimolar assemblies. The inclusive equilibrium models were modified to plot relative peak area of high MW (black lines) and low MW fractions (silver lines) as a function of [variable component]. The silver line-thickness is equal to twice the standard deviation associated with experimentally determined signal factors (see “Methods”). The values for best fit ΔGcon are indicated. For determination of ΔGcon for I53-50, only data where initial [T]0 ≥ [P]0 were considered. In h, i, the purple and orange colored areas emphasize the difference between inclusive equilibrium models and experimental data when [P]0 > [T]0. Source data are provided as a Source Data file.
