Site-specific and complete enzymic 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 biological activity of this hormone. To obtain detailed insight into the function of N-glycosylation, the availability of site-specifically and fully deglycosylated alpha-subunits obtained under non-denaturing conditions is a prerequisite. NMR spectroscopy in combination with FAB-mapping demonstrates that only Asn52 of the alpha-subunit is accessible to digestion by peptide-N4-(N-acetyl-beta-glucosaminyl)asparagine amidase F under native conditions. Treatment of native alpha hCG with endo-beta-N-acetylglucosaminidase B results in full deglycosylation yielding alpha hCG with one GlcNAc residue at both Asn52 and Asn78.
Van Zuylen
C W
CW
Bijvoet Center, Department of Bio-Organic Chemistry, Utrecht University, The Netherlands.
De Beer
T
T
Rademaker
G J
GJ
Haverkamp
J
J
Thomas-Oates
J E
JE
Hård
K
K
Kamerling
J P
JP
Vliegenthart
J F
JF
D000581:Amidohydrolases / Q000378:metabolism; D000595:Amino Acid Sequence; D002240:Carbohydrate Sequence; D015292:Glycoprotein Hormones, alpha Subunit / Q000378:metabolism*; D006031:Glycosylation; D006801:Humans; D009682:Magnetic Resonance Spectroscopy; D008969:Molecular Sequence Data; D043524:Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase; D016339:Spectrometry, Mass, Fast Atom Bombardment
D015292:Glycoprotein Hormones, alpha Subunit; D000581:Amidohydrolases; D043524:Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase