Biochimica et Biophvsica Acta, 684 (1982) 53-58 53 Elsevier Biomedical Press BBA71002 TWO PURIFIED FRACTIONS OF ALAMETHICIN HAVE DIFFERENT CONDUCTANCE PROPERTIES IGOR VODYANOY ~, JAMES E. HALL a T.M. BALASUBRAMANIAN b and GARLAND R. MARSHALL b " Department of Physiology and Bioph.vsics, Universi(v of California. lrt~ine, lrt~ine, ('A 92717 and I, Department of Physiology and Biopllvsics, Washington Universi(v, St. Louis, MO 63110 (U.S.A.) (Received May 20th, 1981) Key words: Alamethicin fraction; Single channel conductance; Conductance The properties of two purified alamethicin fractions, Fraction 4 and Fraction 6, have been studied in phosphatidylethanolamine (PE) membranes and phosphatidylserine (PS) membranes. Membranes doped with Fraction 4 show well-defined single channel conductance (mean lifetime about 20 ms). The autocorrelation function of the current fluctuations has one relaxation time of the same order as the mean lifetime of the single channels, and the current response to a voltage pulse follows an exponential with only one time constant. The conductance of a membrane doped with Fraction 6 has a voltage-independent part and a current-voltage curve with a slope that is half the slope of the Fraction 4 current-voltage curve. In the presence of Fraction 6, PS membranes and PE membranes both have symmetrical current-voltage curves even with Fraction 6 added to only one side. We did not detect any well-defined single channel levels in the presence of Fraction 6, and autocorrelation analysis of the current due to Fraction 6 gave two characteristic correlation times: a fast time (about 5 ms) and a slow time (about 50 ms). High current level kinetics of Fraction 6 also show two time constants. A possible explanation for the differences between the two fractions is that Fraction 6 monomers have a lower dipole moment than those of Fraction 4. The difference in channel stability can be explained by a lowered tendency of the monomers to line up parallel to the field. The negative branch and voltage-independent conductance can be explained by lowered energy of insertion of monomers into the membrane, and lowered energy of interaction between the monomers and the electric field. Introduction Alamethicin (antibiotic U 22324), made by the fungus Trichoderma viride [1] was first used as a model of excitability in lipid bilayers by Mueller and Rudin [2]. Considerable additional work has been done since to understand the mechanism of alamethicin conductance (for review, see Ref. 3). Natural alamethicin shows rather sophisticated conductivity behavior dependent on kind of lipid, membrane thickness, temperature, current and membrane potential. But lack of well-characterized 0005-2736/82/0000-0000/$02,75 © 1982 Elsevier Biomedical Press purified derivatives had led to difficulties in inter- preting some experiments. Because alamethicin provides a simple example of protein-lipid interac- tion, the removal of this difficulty is important to the study of lipid protein interactions. We have initiated a study of the behavior in lipid bilayers of two recently available fractions of purified alamethicin. The structure of Fraction 4 is already known [4,5] and when that of Fraction 6 is availa- ble, it promises to shed light on the molecular basis of alamethicin's ability to induce a strongly voltage-dependent conductance.