Reversible interconversions of sterol-binding and sterol-nonbinding forms of filipin as determined by fluorimetric and light scattering properties.
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Combined fluorimetric and Tyndall light scattering measurements have confirmed the existence of different forms of filipin; filipin herein refers to the filipin complex. In these studies, the ratio of two of the absorbance peaks of the filipin molecule (A338/A305) is a consistent indicator of the presence of active and inactive forms. The partial quantum efficiency (PQ) was used to measure the interaction between filipin and sterols. Light scattering (R90) was used to monitor changes in the aggregate size of the polyene. These analyses have shown that filipin can occur in active formsthose that interact with cholesteroland inactive formsthose that do not interact with cholesterolin aqueous solutions (Schroeder, F., Holland, J. F., and Bieber, L. L. (1972), Biochemistry 11, 3105). Inactive filipin is converted to the active form by incubation at 50 for 2 hr. The conversion of the nonbinding to the binding form in aqueous systems follows a first-order rate law and appears to involve conformational changes that are time and temperature dependent. The conversion of active to inactive forms of filipin is reversible and independent of the presence or absence of large micelles of filipin. Methods using nonaqueous solvents to isolate and purify filipin consistently produce this nonbinding form. Short-chain alcohols disrupt aqueous filipin micelles in the order of effectiveness: propanol > ethanol > methanol. Light scattering of filipin systems is decreased approximately by an order of magnitude with addition of excess cholesterol. Many of the apparent discrepancies in the literature concerning the interaction of filipin with sterols and sterol-containing membranes may result from the use of the sterol-nonbinding form of filipin and the use of concentrations so high that the observations are independent of sterol-binding properties. 1973, American Chemical Society. All rights reserved.
