ESR determinations of membrane permeability in a yeast sterol mutant

Yeast sterol mutants were subjected to ESR analysis in an attempt to elucidate how altered sterol composition correlates with membrane permeability. The technique requires spin labeling the intact yeast cells with a small, water-soluble nitroxide probe (2,2,5,5 tetramethyl-3-pyrrolin-1-oxyl-3-carboxylic acid, PCA), suspending cells in a NiCl₂ solution, and measuring the extent of Ni²⁺ entry through the membrane by its magnetic dipolar line broadening effect on the PCA signal. The wild type, A184D, was found to be impermeable to Ni²⁺ during all growth phases while the sterol mutant $\text{erg}\text{6}\text{2}$ was readily permeable to Ni²⁺. Other sources of line broadening such as increased rotational correlation time and cell nonviability are shown to be negligible. Internal Ni²⁺ concentrations for $\text{erg}\text{6}\text{2}$ and kinetics of Ni²⁺ entry were determined.