Kinetics of the valinomycin-induced potassium ion leak from liposomes with potassium thiocyanate enclosed

Abstract

SUMMARYThe kinetics of the valinomycin-induced K + leak from egg lecithin liposomes with KCNS enclosed were studied by measuring the initial increase of the leak following the addition of the ionophore. Variation of liposome and valinomycin concentrations enabled a discrimination between the affinity of the ionophore for the lipid bilayer and the turnover rate of the transport process. The results indicate that valinomycin is distributed immediately between the outer liposomal bilayer and the aqueous phase. A partition constant of 0.0222 cm at 30 C was obtained for egg lecithin liposomes. By varying the temperature it was shown that the partition constant has a negative temperature coefficient. The Arrhenius activation energy of the transport process itself is 15.7"-~ I. I kcals/mole. Concerning the mechanism of the valinomycinmediated transport it was found that the complex formation can be described kinetically by the formation of a ternary complex in which K + and a thiocyanate ion are bound by the ionophore.By varying the fatty acid composition of the membrane constituents it was found that the turnover rate of the transport increased with increasing unsaturation of the phospholipids present in the membrane. This result suggests that the translocation of the complex is the rate-limiting step of the transport cycle. At 30 c'C a turnover constant of 328±44 potassium ions/molecule valinomycin per second was obtained for egg lecithin lipsomes.

1-Anilino-8-naphthalenesulfonate: A fluorescent probe of ion and ionophore transport kinetics and trans-membrane asymmetry

“…Our measured value of AH:~ is in good agreement with the values measured by Johnson and Bangham (1969) and Blok et al (1974) for 42K+ flux and KCNS efflux (respectively) through phospholipid membranes.…”

“…It has been shown by de Gier, Mandersloot and van Deenen (1968) that the nonelectrolyte permeability of liposomes increases with increasing degree of unsaturation and decreasing chain length. The lipid bilayer experiments were carried out at least 30~ above their phase transition temperature, whereas the k3b values of Table 6 are for a temperature 10 ~ above T m. The transport reaction has a large activation enthalpy (AH~: = 16.3 + 0.5 kcal) and extrapolation of the measured rates to 53~ (30~ above Tin) gives a k3b value of 310sec -1 in good agreement with the value of Blok et al (1974). The remaining difference between this value and the values for mitochondria (Haynes et al, 1974) and the lipid bilayer (Benz et al, 1973) may be due to the presence of integral proteins in the former system and the presence of organic solvents in the latter system.…”

“…Blok, de Gier and Van Deenen (1974) have given evidence for a similar mechanism for valinomycin-facilitated K + (CNS-) efflux from liposomes. It is further assumed that the interracial complexation and decomplexation steps are rapid, such that k 2 > k4 > k3b" The following expression can be derived for the dependence of the transport rate on…”

“…Blok et al (1974) calculated a turnover number of 328 _+ 44 and K z = 0.62 -3.0 M-1 and K 3 = 20 M-1 for valinomycin-induced KCNS permeability in phospholipid multilayers. and Benz et al (1973) have reported first order rate constants of ca.…”

Transmembrane electrophoresis of 8-anilino-1-naphthalenesulfonate through egg lecithin liposome membranes

“…The possibility that ANS permeation may be induced or increased by a valinomycin potassium complex is suggested by the observation that this complex may be associated with anions (Davis & Tosteson, 1971;Blok, De Gier & Van Deenen, 1974) and, more specifically, with ANS in water and chloroform (Feinstein & Felsenfeld, 1971) and possibly in phospholipid membranes (Haynes, 1972). Further, Feinstein and Felsenfeld (1971) have shown that the presence of valinomycin and potassium chloride increases the partition of ANS from water into chloroform.…”