ABSTRACT
Ligand-binding activity of rat testis homogenate increases during storage for 24 h at 4°C, and rapidly deteriorates after 48 h at the same temperature. Inhibition of cellular proteases with phenylmethanesulphonylfluoride (PMSF) preserves the activity for 7 days, and, lyophilized preparations of the tissue homogenate retain their ligand-binding activity for prolonged periods of time during storage at 4°C. The rate of increase in ligand-binding capacity is temperature-dependent. Enhancement of activity is also noted in the purified membrane fraction and appears to be due to unmasking of receptor molecules in the membrane. Even in the presence of PMSF, there is a rapid and irreversible loss of ligand-binding activity when either fresh or reconstituted freeze-dried receptor preparations are pre-incubated at 37°C, although in the former case this is preceeded by a brief period of activation. The destabilization of the reconstituted lyophilized preparation at 37°C is induced by the dehydration step in the freeze-drying process and not by freezing alone. It is therefore unlikely that the rapid inactivation at 37°C is enzymatically induced; rather it is due to destabilization of membrane architecture and/or receptor accessibility. The inherent inaccuracies of measurements of kinetic parameters and assessments of the number of receptor sites in such unstable systems by Scatchard plots of binding data are pointed out.
Evidence is presented which indicates that in the presence of luteinizing hormone the spontaneous inactivation of receptors is inhibited and that there is an apparent "conservation" of receptor sites. A mechanism whereby this is accomplished is proposed, on the basis of the kinetics of inactivation.
Conformational changes in the chicken receptor for endocytosis of glycoproteins. Modulation of ligand-binding activity by Ca2+ and pH.