The asparagine-linked oligosaccharides of the human chorionic gonadotropin beta subunit facilitate correct disulfide bond pairing

J Biol Chem. 1995 May 19;270(20):11851-9. doi: 10.1074/jbc.270.20.11851.

Abstract

The role of asparagine (N)-linked oligosaccharide chains in intracellular folding of the human chorionic gonadotropin (hCG)-beta subunit was determined by examining the kinetics of folding in Chinese hamster ovary (CHO) cells transfected with wild-type or mutant hCG-beta genes lacking one or both of the asparagine glycosylation sites. The half-time for folding of p beta 1 into p beta 2, the rate-determining step in beta folding, was 7 min for wild-type beta but 33 min for beta lacking both N-linked glycans. The p beta 1-->p beta 2 half-time was 7.5 min in CHO cells expressing the beta subunit missing the Asn13-linked glycan and 10 min for the beta subunit missing the Asn30-linked glycan. The inefficient folding of hCG-beta lacking both N-linked glycans correlated with the slow formation of the last three disulfide bonds (i.e. disulfides 23-72, 93-100, and 26-110) to form in the hCG-beta-folding pathway. Unglycosylated hCG-beta was slowly secreted from CHO cells, and beta subunit-folding intermediates retained in cells for more than 5 h were degraded into a hCG-beta core fragment-like protein. However, coexpression of the hCG-alpha gene enhanced folding and formation of disulfide bonds 23-72, 93-100, and 26-110 of hCG-beta lacking N-linked glycans. In addition, the molecular chaperones BiP, ERp72, and ERp94, but not calnexin, were found in a complex with unglycosylated, unfolded hCG-beta and may be involved in the folding of this beta form. These data indicate that N-linked oligosaccharides assist hCG-beta subunit folding by facilitating disulfide bond formation.

Publication types

  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Asparagine / chemistry*
  • CHO Cells
  • Carbohydrate Sequence
  • Chorionic Gonadotropin / chemistry
  • Chorionic Gonadotropin / genetics
  • Chorionic Gonadotropin / metabolism*
  • Chorionic Gonadotropin, beta Subunit, Human
  • Cricetinae
  • Cystine / metabolism*
  • Glycoprotein Hormones, alpha Subunit / genetics
  • Glycoprotein Hormones, alpha Subunit / metabolism
  • Glycosylation
  • Macromolecular Substances
  • Molecular Chaperones / metabolism
  • Molecular Sequence Data
  • Oligosaccharides / metabolism*
  • Peptide Fragments / chemistry
  • Peptide Fragments / genetics
  • Peptide Fragments / metabolism*
  • Protein Folding*
  • Protein Processing, Post-Translational*
  • Recombinant Fusion Proteins / metabolism

Substances

  • Chorionic Gonadotropin
  • Chorionic Gonadotropin, beta Subunit, Human
  • Glycoprotein Hormones, alpha Subunit
  • Macromolecular Substances
  • Molecular Chaperones
  • Oligosaccharides
  • Peptide Fragments
  • Recombinant Fusion Proteins
  • Cystine
  • Asparagine