Spin label studies of microsomal membranes from Acanthamoeba castellanii in different states of differentiation

Abstract

Two fatty acid spin labels—[I(1,14)], stearic acid bearing a paramagnetic nitroxide group on carbon 16, and [I(12,3)], stearic acid bearing a paramagnetic nitroxide group on carbon 5—have been used to compare the physical properties of lipid in rough and smooth microsomal membranes from trophozoites and cysts of Acanthamoeba castellanii. Arrhenius plots of rotational correlation times ( τ c ) calculated from the spectra for I(1,14) showed an abrupt discontinuity in slope for membranes from both trophozoites and cysts. This occurred at temperatures ranging from −3 to 1 °C for smooth microsomes and from 8 to 11 °C for rough microsomes for both cysts and amoebae. The value of τ c at 29 °C, the culturing temperature, in effect scores fluidity of the membrane matrix, and did not show any significant difference for either rough or smooth microsomes during the transition from exponential to stationary phase growth. However, smooth microsomes from cysts showed a 14% increase in fluidity relative to trophozoites, and the fluidity of rough microsomes from cysts tended to be lower. An order parameter ( S) calculated from spectra for I(12,3) did not change as a function of encystment for the smooth membranes and increased only slightly for rough microsomes. The activation energy ( E a) for Arrhenius plots of τ c above the inflection temperature increased as a result of encystment, indicating a greater degree of molecular interaction within the cyst membranes. Moreover, the τ c plots for both rough and smooth microsomal membranes from trophozoites tended to converge at 29 °C, the growth temperature, whereas plots for cyst membranes were virtually parallel, bracketing those for the trophozoite membranes. This suggests that the trophozoite is able to regulate its membrane fluidity and that cysts, which are resting cells, have lost this regulatory capacity.