Epitope mapping of monoclonal antibodies specific for a macrophage lectin: a calcium-dependent epitope is in the carbohydrate recognition domain.
Mouse macrophage galactose/N-acetylgalactosamine-specific calcium-type lectin (mMGL) has a calcium-dependent conformational epitope which is a ligand-induced binding site. A monoclonal antibody (mAb) specific for this epitope (LOM-11) stabilize lectin activity. We performed mapping for this conformational epitope using trypsin fragments that contain a carbohydrate recognition domain (CRD) and chimeric recombinant proteins between mMGL and a human counterpart of this molecule. Binding site for the mAb LOM-11 was mapped within the C-terminal 59 amino acids of CRD. Binding sites for all four mAbs that block carbohydrate ligand binding were also mapped in the C-terminal half of CRD. These results indicated that the calcium-dependent site potentially involved in protein-protein interaction, regulatory or for coordinated binding, is mapped within CRD in addition to the independent carbohydrate binding site, and that both of the distinct sites may have spatial proximity.
Kimura
T
T
Laboratory of Cancer Biology and Molecular Immunology, Graduate School of Pharmaceutical Sciences, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, 113-0033, Tokyo, Japan.
Hosoi
T
T
Yamamoto
K
K
Suzuki
N
N
Imai
Y
Y
Irimura
T
T
D000911:Antibodies, Monoclonal / Q000276:immunology*; D001665:Binding Sites; D002118:Calcium / Q000494:pharmacology*; D002352:Carrier Proteins / Q000187:drug effects / Q000235:genetics / Q000276:immunology*; D018604:Epitope Mapping; D037102:Lectins / Q000187:drug effects / Q000235:genetics / Q000276:immunology*; D037181:Lectins, C-Type; D008264:Macrophages / Q000276:immunology*; D008565:Membrane Proteins; D010446:Peptide Fragments / Q000276:immunology; D011993:Recombinant Fusion Proteins / Q000276:immunology; D014357:Trypsin
D000911:Antibodies, Monoclonal; D002352:Carrier Proteins; D037102:Lectins; D037181:Lectins, C-Type; D008565:Membrane Proteins; D010446:Peptide Fragments; D011993:Recombinant Fusion Proteins; D014357:Trypsin; D002118:Calcium