The effect of pH on stereospecific opiate binding to mouse brain membranes

High-affinity, in vitro stereospecific binding of 3H-di-hydromorphine or 3H-naloxone to brain membranes shows a marked dependence on pH; maximal binding, observed at pH 7.5-8.0, is abruptly and reversibly reduced as the pH is lowered, with the binding half-maximal at about pH 6.8. A similar pH dependence of stereospecific binding is observed with the quaternary agonist N-methyl morphine, but non-specific 3H-di-hydromorphine of 3H-naloxone binding (that not displaced by a 100-fold excess of unlabelled levorphanol) shows only a slight decrease in this pH range. Binding of agonist and antagonist at pH 6.8 show the same differences with respect to trypsin sensitivity and to the effects of Na+ and Mn++ ions that are seen at pH 8.0. It is concluded that the ability of low pH to reduce stereospecific binding is due to protonation of a membrane-bound site, and that this site is probably the anionic site of the opiate receptor. Of the four anionic groups commonly found in biological membranes, only phosphate exhibits a pK close to that of this effect. Taken with other evidence, the results suggest that the opiate receptor may be a phosphoprotein.