title: Kinetics of folding and assembly of the human chorionic gonadotropin beta subunit in transfected Chinese hamster ovary cells.
abstract: We have employed Chinese hamster ovary (CHO) cell lines transfected with either the wild type human chorionic gonadotropin beta (hCG-beta) gene alone (CHO beta cells) or in conjunction with the gene expressing the alpha subunit (CHO alpha,beta cells) to study the folding pathway of the hCG-beta subunit. In both CHO beta and CHO alpha,beta cells, the earliest detectable hCG-beta precursor, p beta 1, which had two of six potential disulfide bonds (34-88 and 38-57) formed, was converted to p beta 2, a form that, following the formation of disulfide bonds between cysteines 9-90 and 23-72, migrated more slowly than p beta 1 by sodium dodecyl sulfate-polyacrylamide gel electrophoresis under nonreducing conditions. The t1/2 for the conversion of p beta 1 to p beta 2 in CHO alpha,beta and CHO beta cells was 5 min, demonstrating that the alpha subunit had no effect on the rate of this conversion. Furthermore, the tryptic-releasable peptides generated from nonreduced p beta 1 or p beta 2 were the same in both CHO alpha,beta and CHO beta cells. Thus, both the rate and order of disulfide bond formation during the conversion of the folding intermediate p beta 1 into p beta 2 were the same whether or not the alpha subunit was present. A comparison between cell types expressing different alpha/beta subunit ratios revealed that the higher the glycoprotein hormone alpha subunit to beta subunit ratio, the greater the rate and extent of hCG heterodimer assembly.
authors.affiliation: Eppley Institute for Research in Cancer and Allied Diseases, University of Nebraska Medical Center, Omaha 68198-6805.
authors.forename: E
authors.initials: E
authors.lastname: Bedows
authors.forename: J R
authors.initials: JR
authors.lastname: Huth
authors.forename: R W
authors.initials: RW
authors.lastname: Ruddon
mesh_terms: D000818:Animals; D016466:CHO Cells; D006063:Chorionic Gonadotropin / Q000737:chemistry* / Q000235:genetics; D002852:Chromatography, Ion Exchange; D006224:Cricetinae; D004220:Disulfides / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D007700:Kinetics; D010449:Peptide Mapping; D011233:Precipitin Tests; D011487:Protein Conformation; D014162:Transfection; D014357:Trypsin / Q000378:metabolism
chemical_list: D006063:Chorionic Gonadotropin; D004220:Disulfides; D014357:Trypsin