Rapid equilibrium between monomeric, dimeric and tetrameric forms of the 46‐kDa mannose 6‐phosphate receptor at 37°C

Abstract

At 4 degrees C, detergent-solubilized 46-kDa mannose 6-phosphate receptor (MPR 46) exists as a mixture of dimeric and tetrameric forms [A. Waheed, A. Hille, U. Junghans & K. von Figura (1990) Biochemistry 29, 2449-2455]. At 37 degrees C, MPR 46 exists as a mixture of monomeric, dimeric and tetrameric forms, which can be separated by sucrose density centrifugation or chromatography on a mannose-6-phosphate affinity matrix. Monomeric MPR 46 did not bind to the affinity matrix, while dimeric and tetrameric receptors were eluted with increasing concentrations of mannose 6-phosphate. Depending on the incubation temperature, dimeric MPR 46 preferentially associates to tetramers (less than or equal to 16 degrees C) or dissociates to monomers (greater than or equal to 25 degrees C). The presence of mannose 6-phosphate shifts the equilibrium to higher quaternary structure, with a maximal effect at 37 degrees C. Incubation of dimeric or tetrameric receptor at pH 4.0 induces a rapid dissociation of about a third of the receptor with t1/2 of less than 2 min, followed by a phase of slow dissociation. Readjusting the pH to 7.5 induces a rapid formation of dimers and tetramers, which is promoted by the presence of mannose 6-phosphate or an increase of the receptor concentration. These observations may indicate that the recycling of the receptor between the Golgi apparatus, where it binds ligands at near-neutral pH, and the acidic prelysosomes, where it releases the ligands, is associated with a change of its quaternary structure, which in turn may affect the recycling kinetics. In keeping with this assumption, we observed in baby hamster kidney cells over-expressing MPR 46 and in membrane prepared from these cells monomeric, dimeric and tetrameric forms of the receptor.