Lectin analysis of human immunoglobulin G N-glycan sialylation.
The lectins Sambucus nigra agglutinin (SNA) and Ricinus communis agglutinin (RCA), specific for alpha2,6 linked sialylation, and terminal galactose respectively were used to study the occurrence, linkage and distribution of human immunoglobulin G (IgG) sialylation. SNA was shown to bind N-glycan alpha2,6-linked sialic acid only. Sialidase analysis confirmed that this is the dominant, if not exclusive linkage. Total IgG sialylation was estimated at 1.0 microg SA/mg IgG (or about 0.5 mole per mole) using a biochemical sialic acid assay. SNA displayed strong binding to the IgG Fab fragment in both its native and denatured state. In contrast, SNA failed to bind the IgG Fc fragment in its native form, but displayed strong binding after the Fc was denatured. This allowed the construction of quantitative assays capable of measuring both IgG Fab and Fc alpha2,6-sialylation without the need for enzymatic peptide digestion.
Dalziel
M
M
ICRF Breast Cancer Biology Group, Thomas Guy House, Guy's Hospital, London, UK. M.Dalziel@icrf.icnet.uk
McFarlane
I
I
Axford
J S
JS
D000581:Amidohydrolases; D001665:Binding Sites; D002236:Carbohydrate Conformation; D006801:Humans; D007140:Immunoglobulin Fab Fragments / Q000737:chemistry / Q000378:metabolism; D007141:Immunoglobulin Fc Fragments / Q000737:chemistry / Q000378:metabolism; D007074:Immunoglobulin G / Q000737:chemistry* / Q000378:metabolism*; D066298:In Vitro Techniques; D037102:Lectins / Q000378:metabolism*; D009439:Neuraminidase; D043524:Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase; D037121:Plant Lectins; D011134:Polysaccharides / Q000737:chemistry* / Q000378:metabolism*; D054788:Ribosome Inactivating Proteins; D012794:Sialic Acids / Q000737:chemistry
D007140:Immunoglobulin Fab Fragments; D007141:Immunoglobulin Fc Fragments; D007074:Immunoglobulin G; D037102:Lectins; D037121:Plant Lectins; D011134:Polysaccharides; C017707:Ricinus communis agglutinin-1; C050600:Sambucus nigra lectins; D012794:Sialic Acids; D009439:Neuraminidase; D054788:Ribosome Inactivating Proteins; D000581:Amidohydrolases; D043524:Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase