The sugar binding activity of MR60, a mannose-specific shuttling lectin, requires a dimeric state

MR60 is an intracellular membrane protein which has been shown to act as a mannoside specific lectin and to be identical to ERGIC-53, a protein charac teristic of the endoplasmic reticulum-Golgi apparatus-intermediate compartm ent, acting as a shuttle. According to its primary sequence, this MR60/ERGI C-53 protein contains a luminal domain including the carbohydrate recogniti on domain, a stem, a transmembrane segment and a cytosolic domain. The endo genous MR60/ERGIC-53 protein is spontaneously oligomeric, (dimers and hexam ers), In this paper, we study the relationship between the oligomerization state and the sugar binding capacity by using recombinant proteins. The exp ression of the recombinant proteins was evidenced by immunocytochemistry an d by immunoprecipitation followed by SDS-PAGE analysis. The full size recom binant protein binds mannosides and is oligomeric, up to the hexameric form . Two truncated proteins lacking the transmembrane and the cytosolic domain s were prepared and characterized. A long one, containing the cysteine 466 close to the C-terminal end of the recombinant protein but lacking the cyst eine 475, close to the C-terminal end of the native protein, does bind mann osides and forms dimers but no higher oligomeric forms. A shorter one, lack ing both the cysteines 466 and 475, does not bind mannosides and does not f orm dimers or higher polymers. The two cysteines in the carbohydrate recogn ition domain (C190 and C230) are not involved in the stabilization of oligo mers, In conclusion, this study shows that the luminal moiety of MR60/ERGIC -53 contains a device allowing both its oligomeric pattern and its sugar bi nding capability.