Studies on the relevance of the glycan at Asn-52 of the alpha-subunit of human chorionic gonadotropin in the alphabeta dimer.
Glycosylation of Asn-52 of the alpha-subunit (alphaAsn-52) is required for bioactivity of the alphabeta-dimeric human chorionic gonadotropin (hCG), although at a molecular level the effect of the glycan at alphaAsn-52 is not yet understood. To study the role of this glycan for heterodimer stability, the beta-subunit was recombined in solution with either the alpha-subunit or the alpha-subunit enzymically deglycosylated at alphaAsn-52. Enzymic deglycosylation avoids modification of the glycans at alphaAsn-78 and disturbing the protein folding. The efficiency of recombination after 16 h is 80%, independent of whether alphaAsn-52 is glycosylated or not. The dissociation constant of the hCG complex, with or without the glycan at alphaAsn-52, is less than 1 x 10(-5) s(-1), indicating that the glycan at alphaAsn-52 does not contribute significantly to the stability of the dimer. CD and NMR spectra indicate a local conformational difference between both alphabeta-dimeric hCG variants, most probably involving amino acids of the hCG beta-subunit close to the glycan at alphaAsn-52. These data explain the native-like receptor-binding abilities of hCG lacking the glycan at alphaAsn-52. It is proposed that for bioactivity the glycan at alphaAsn-52 is necessary for inducing and stabilizing a conformational change in hCG upon binding to the receptor, resulting in activation of the signal-transduction pathway.
Erbel
Paul J A
PJ
Department of Bio-Organic Chemistry, Bijvoet Center, Utrecht University, P.O. Box 80.075, NL-3508 TB Utrecht, The Netherlands.
Haseley
Simon R
SR
Kamerling
Johannis P
JP
Vliegenthart
Johannes F G
JF
D001216:Asparagine / Q000737:chemistry*; D006063:Chorionic Gonadotropin / Q000737:chemistry*; D002942:Circular Dichroism; D019281:Dimerization; D006801:Humans; D007700:Kinetics; D008958:Models, Molecular; D010084:Oxidation-Reduction; D011134:Polysaccharides / Q000737:chemistry*; D011487:Protein Conformation; D020349:Surface Plasmon Resonance; D014961:X-Ray Diffraction
D006063:Chorionic Gonadotropin; D011134:Polysaccharides; D001216:Asparagine