Cholesterol triggers clustering of PI(4,5)P ₂ molecules. (A) Size-aggregation frequency analysis (right panel) for systems without (gray color) and with 30 mol% cholesterol (brown color). The analysis was averaged over the two membrane leaflets considering the last 1,000 ns of simulations. Data are shown as the average of both bilayer leaflets, and the error is given as standard deviation. Two monomers were considered as a dimer if any of their atoms had a distance of <1.2 nm. The right panel represents a snapshot of a lateral partial density analysis with a dimer present as an example. (B) Time evolution of the cluster fractional occupation number distribution of PI(4,5)P₂ head groups for systems without (subpanels a and b), and with 30 mol% cholesterol (subpanels c and d). The calculation was made for each membrane leaflet separately. Each panel represents a 1,200 ns MD simulation of a system with eight PI(4,5)P₂ molecules (head groups only). The fractional occupation number was calculated every nanosecond and it is represented with a black and white scale bar.