Studies on the carbohydrate moiety of vitellogenin from the tobacco hornworm,Manduca sexta

Vitellogenin can be isolated in large quantities from the hemolymph of the tobacco hornworm, Manduca sexta by a combination of KBr density gradient ultracentrifugation, gel permeation and cation exchange chromatography. Glycopeptides generated by exhaustive pronase digestion of vitellogenin were separated by gel permeation chromatography on a Bio-Gel P-6 column. The major glycopeptide was shown to bind to concanavalin A-Sepharose but not to lentil lectin-Sepharose and was sensitive to treatment with endo-β-N-acetylglucosaminidase H. Analysis of the glycopeptide by high field proton NMR spectroscopy revealed that the primary structure is of the high mannose class containing nine mannose and two N-acetylglucosamine units. These results suggested that N-glycosylation of insect proteins, as in mammals, yeasts and plants, involves the en bloc transfer of an oligosaccharide containing the unit (Glu)₃(Man)₉(GlcNAc)₂ from a lipid intermediate to an asparagine residue followed by removal of glucose residues. Processing to more complex structures does not seem to occur in M. sexta vitellogenin. In vitro uptake with isolated M. sexta follicles showed that deglycosylation had no significant effect on uptake of ¹²⁵I-labeled vitellogenin.