Insulin receptor carbohydrate units contain poly-N-acetyllactosamine chains.
The insulin receptor was immunoprecipitated from cultured human lymphocytes (IM-9) and rat hepatocytes (Fao) after biosynthetic labeling with [3H]glucosamine or [3H]mannose, and the nature of the carbohydrate units was investigated. Digestion of the receptor from IM-9 lymphocytes with E. freundii endo-beta-galactosidase increased the migration of the insulin receptor alpha- and beta-subunits on sodium dodecyl sulfate-polyacrylamide gels and sharpened the electrophoretic bands; the alpha-subunit was converted from an apparent mol wt (Mr) of 123,000 to a Mr of 118,000, and the beta-subunit from a Mr of 92,000 to 89,000. The susceptibility of the insulin receptor to this enzyme indicates that its carbohydrate units contain poly-N-acetyllactosamine sequences. Affinity chromatography of receptor glycopeptides on Concanavalin-A-Sepharose revealed that the poly-N-acetyllactosamine units were attached to multiantennary glycopeptides that accounted for over 75% of the [3H]glucosamine incorporated into the IM-9 lymphocyte insulin receptor; the remaining radioactivity was present in polymannose units (primarily Man8GlcNAc2) and biantennary complex saccharides. Several differences in the carbohydrate chains of the insulin receptor from the Fao and IM-9 cells indicated that glycosylation was cell specific despite the occurrence of poly-N-acetyllactosamine chains in both cell types. The IM-9 lymphocyte receptor glycopeptides were larger (Mr, 3,200-9,500) and more susceptible to endo-beta-galactosidase than those from the Fao receptor (Mr, 3,000-5,000). Moreover, the released saccharides from the Fao receptor were found by exoglycosidase digestions and chromatographic comparison to standards to contain terminal sialic acid in both alpha 2----3 and alpha 2----6 linkage to galactose, whereas the IM-9 carbohydrate units contained only alpha 2----3-linked sialic acid.
Edge
A S
AS
Department of Medicine, Harvard Medical School, Boston, Massachusetts.
Kahn
C R
CR
Spiro
R G
RG
D000818:Animals; D002236:Carbohydrate Conformation; D002240:Carbohydrate Sequence; D002241:Carbohydrates / Q000032:analysis*; D002460:Cell Line; D002846:Chromatography, Affinity; D005944:Glucosamine / Q000378:metabolism; D006026:Glycoside Hydrolases; D006031:Glycosylation; D006801:Humans; D008099:Liver / Q000032:analysis; D008214:Lymphocytes / Q000032:analysis; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009844:Oligosaccharides / Q000032:analysis / Q000378:metabolism; D011134:Polysaccharides / Q000032:analysis*; D051381:Rats; D011972:Receptor, Insulin / Q000032:analysis* / Q000378:metabolism; D001616:beta-Galactosidase / Q000378:metabolism
D002241:Carbohydrates; D009844:Oligosaccharides; D011134:Polysaccharides; C037199:poly-N-acetyllactosamine; D011972:Receptor, Insulin; D006026:Glycoside Hydrolases; C026093:keratan-sulfate endo-1,4-beta-galactosidase; D001616:beta-Galactosidase; D005944:Glucosamine