Add time:08/16/2019         Source:sciencedirect.com

Digitonin is commonly used to permeabilize cell membranes and solubilize membrane components. It interacts specifically with cholesterol in the membrane which leads to the formation of pores. Thus far, the mechanism by which digitonin interacts with the membrane has only been described qualitatively. We investigated this interaction in model membranes that contain little or no cholesterol with a combination of isothermal titration calorimetry, dynamic light scattering, and zeta potential measurements. Digitonin partitions fully asymmetrically into large unilamellar vesicles of phosphocholine (PC) lipid at 20 °C (remaining in the outer leaflet only), with a partition coefficient of 0.22 ± 0.04 mM−1 and ΔH of partitioning of 23.3 ± 1.6 kJ mol−1. Beyond a digitonin/lipid ratio of ∼0.1 in the outer leaflet, digitonin micelles coexist with vesicles without solubilizing them—even at high digitonin concentrations. This “staying out” of digitonin was also observed with phosphoserine (PS), PC/PS, and PC/PS/cholesterol vesicles. The mechanism by which digitonin perturbs and solubilizes the membrane is very different when the membrane contains little or no cholesterol as opposed to 20–30 mol% cholesterol. The role of digitonin should thus be carefully considered in the design of preparative protocols and experiments in studies of cellular processes and membrane proteins.

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