The effects of ionic strength on the protein conformation and the fluidity of porcine intestinal brush border membranes. Fluorometric studies using N-[7-dimethylamino-4-methylcoumarinyl]maleimide and pyrene.
The effects of ionic strength on the conformation around the SH groups of the proteins and the lipid fluidity of porcine intestinal brush border membranes were studied using two fluorescent dyes, N-[7-dimethylamino-4-methylcoumarinyl]maleimide (DACM) and pyrene. The extent of DACM labeling to the SH groups of the membrane proteins was accelerated depending on the KCl concentrations in medium. A quenching study of DACM-labeled membranes with acrylamide showed that the proximity of the quencher to the fluorescence-labeled SH groups in the membrane proteins is increased with increasing ionic strength of medium. An implication of the conformational changes around SH groups in the membrane proteins with increase of ionic strength was also obtained from the stimulation of guanidine effect on the fluorescence parameters of DACM-labeled membranes by addition of KCl. On the other hand, the results of the quenching study with KI, excimer fluorescence, and polarization measurements of pyrene-labeled membranes suggested an increase of membrane fluidity on addition of KCl to medium. The temperature dependence of polarization of the complex strongly suggested that the rotational freedom of pyrene molecules embedded into the lipid layers of the membranes is increased by addition of KCl. In fact, the harmonic means of the rotational relaxation times of pyrene molecules in the membranes with and without 100 mM KCl were estimated to be about 2900 and 9000 ns at 25 degrees C, respectively. Based on these results, the salt-induced alterations of the conformation in the vicinity of the bound dyes of the membrane proteins and of the membrane fluidity are discussed.
D000818:Animals; D005470:Fluorometry; D019791:Guanidine; D006146:Guanidines / Q000494:pharmacology; D007202:Indicators and Reagents; D007421:Intestine, Small / Q000648:ultrastructure*; D008301:Maleimides; D008433:Mathematics; D008560:Membrane Fluidity; D008871:Microvilli / Q000648:ultrastructure; D009994:Osmolar Concentration; D011189:Potassium Chloride / Q000494:pharmacology; D011487:Protein Conformation; D011721:Pyrenes; D013552:Swine; D013696:Temperature
D006146:Guanidines; D007202:Indicators and Reagents; D008301:Maleimides; D011721:Pyrenes; C019347:N-(7-dimethylamino-4-methylcoumarinyl)maleimide; D011189:Potassium Chloride; C030984:pyrene; D019791:Guanidine