ERGIC-53 IS A FUNCTIONAL MANNOSE-SELECTIVE AND CALCIUM-DEPENDENT HUMAN HOMOLOG OF LEGUMINOUS LECTINS

Based on sequence homologies with leguminous lectins, the intermediate compartment marker ERGIC-53 was proposed to be a member of a putative new class of animal lectins associated with the secretory pathway. In dependently, a promyelocytic protein, MR60, was purified by mannose-co lumn chromatography, and a cDNA was isolated that matched MR60 peptide sequences. This cDNA was identical to that of ERGIC-53 and homologies With the animal lectin family of the galectins were noticed. Not all peptide sequences of MR60, however, were found in ERGIC-53, raising th e possibility that another protein associated with ERGIC-53 may posses s the lectin activity. Here, we provide the first direct evidence for a lectin function of ERGIC-53. Overexpressed ERGIC-53 binds to a manno se column in a calcium-dependent manner and also co-stains with mannos ylated neoglycoprotein in a morphological binding assay. By using a se quential elution protocol we show that ERGIC-53 has selectivity for ma nnose and low affinity for glucose and GlcNAc, but no affinity for gal actose. To experimentally address the putative homology of ERGIC-53 to leguminous lectins, a highly conserved protein family with an invaria nt asparagine essential for carbohydrate binding, we substituted the c orresponding asparagine in ERGIC-53. This mutation, as well as a mutat ion affecting a second site in the putative carbohydrate recognition d omain, abolished mannose-column binding and co-staining with mannosyla ted neoglycoprotein. These findings establish ERGIC-53 as a lectin and provide functional evidence for its relationship to leguminous lectin s. Based on its monosaccharide specificity, domain organization, and r ecycling properties, we propose ERGIC-53 to function as a sorting rece ptor for glycoproteins in the early secretory pathway.