Cystine knot mutations affect the folding of the glycoprotein hormone alpha-subunit. Differential secretion and assembly of partially folded intermediates

The common glycoprotein hormone alpha-subunit (GPH-alpha) contains five intramolecular disulfide bonds, three of which form a cystine knot motif (10-60, 28-82, and 32-84). By converting each pair of cysteine residues of a given disulfide bond to alanine, we have studied the role of individual disulf...

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Bibliographic Details
Published in:The Journal of biological chemistry 2000-05, Vol.275 (20), p.15413-15421
Main Authors: Darling, R J, Ruddon, R W, Perini, F, Bedows, E
Format: Article
Language:English
Subjects:
Alanine
Amino Acid Substitution
Cell Line
Cysteine
Cystine
Disulfides
Dithiothreitol - pharmacology
glycoprotein hormone
Glycoprotein Hormones, alpha Subunit - chemistry
Glycoprotein Hormones, alpha Subunit - metabolism
Humans
Models, Molecular
Mutagenesis, Site-Directed
Point Mutation
Protein Folding
Protein Structure, Secondary
Recombinant Proteins - chemistry
Recombinant Proteins - metabolism
Transfection
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Summary:The common glycoprotein hormone alpha-subunit (GPH-alpha) contains five intramolecular disulfide bonds, three of which form a cystine knot motif (10-60, 28-82, and 32-84). By converting each pair of cysteine residues of a given disulfide bond to alanine, we have studied the role of individual disulfide bonds in GPH-alpha folding and have related folding ability to secretion and assembly with the human chorionic gonadotropin beta-subunit (hCG-beta). Mutation of non-cystine knot disulfide bond 7-31, bond 59-87, or both (leaving only the cystine knot) resulted in an efficiently secreted folding form that was indistinguishable from wild type. Conversely, the cystine knot mutants were inefficiently secreted (
ISSN:0021-9258
DOI:10.1074/jbc.275.20.15413