Temperature-Induced Changes in Lipid Fluidity Alter the Conformation of Proteins in Senescing Plant Membranes

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Abstract
Complementary biophysical techniques have provided evidence for an effect of temperature-induced changes in lipid fluidity on the conformation of proteins in senescing plant membranes. Smooth microsomal membranes were isolated from bean cotyledons (Phaseolus vulgaris L. (cv Kinghorn)) at various stages of senescence. Lipid fluidity was measured by fluorescence polarization after labelling the membranes with diphenyl hexatriene (DPH). Alterations in protein conformation were determined by labelling the membranes with the paramagnetic sulfhydryl reagent, 3-maleimido proxyl (3-MP), and following changes in the ratio of weakly immobilized to strongly immobilized (w/s) spin label. Plots of w/s as a function of temperature featured characteristic break points for each age of membrane. Corresponding break points at virtually identical temperatures were also observed in plots of DPH polarization versus temperature for membranes as well as for liposomes prepared from lipid extracts of membranes. In at least one instance these break points in DPH polarization did not correspond to liquid-crystalline to gel phase transitions in the lipid. The fluidity of microsomal membranes decreased with advancing senescence of the cotyledons, and there were also changes in the parameter w/s for membrane proteins with advancing age. The results indicate that subtle changes in the molecular ordering of lipid bilayers alter the relative proportions of weakly and strongly immobilized sulfhydryl groups in the membrane proteins, which can be interpreted as reflecting changes in protein conformation.