SITE-SPECIFIC AND COMPLETE ENZYMATIC DEGLYCOSYLATION OF THE NATIVE HUMAN CHORIONIC-GONADOTROPIN ALPHA-SUBUNIT

Numerous studies have shown that glycosylation of the alpha-subunit of human chorionic gonadotropin (alpha hCG) is essential for the biologi cal activity of this hormone. To obtain detailed insight into the func tion of N-glycosylation, the availability of site-specifically and ful ly deglycosylated alpha-subunits obtained under non-denaturing conditi ons is a prerequisite. NMR spectroscopy in combination with FAB-mappin g demonstrates that only Asn52 of the alpha-subunit is accessible to d igestion by peptide-N-4-(N-acetyl-beta-glucosaminyl)asparagine amidase F under native conditions. Treatment of native alpha hCG with endo-be ta-N-acetylglucosaminidase B results in full deglycosylation yielding alpha hCG with one GlcNAc residue at both Asn52 and Asn78.