title: Bacterial expression of a natively folded extracellular domain fusion protein of the hFSH receptor in the cytoplasm of Escherichia coli.
abstract: We have expressed the extracellular domain of the hFSH receptor as a fusion protein with thioredoxin in the cytoplasm of an Escherichia coli strain that contains mutations in both the thioredoxin reductase and the glutathione reductase genes. The chimeric protein isolated following induction of expression was purified in a soluble form and binds hFSH with an affinity approximating that of native receptor. This truncated form of the receptor displays the same specificity as intact receptor and does not bind hCG. The protein is expressed at levels that exceed 5 mg/L in the bacterial cytoplasm. Expression of the properly folded extracellular domain of the hFSH receptor in the cytoplasm of E. coli allows the facile and economical purification of large quantities of material. This will facilitate the determination of the structure of the hormone-binding domain of this glycoprotein receptor as well as the production of epitope-specific antibodies.
authors.affiliation: Center for Reproductive Science and Department of Obstetrics and Gynecology, Columbia University, 630 West 168th Street, New York, New York 10032, USA.
authors.forename: Leslie
authors.initials: L
authors.lastname: Lobel
authors.forename: Susan
authors.initials: S
authors.lastname: Pollak
authors.forename: Brandie
authors.initials: B
authors.lastname: Lustbader
authors.forename: Jeffrey
authors.initials: J
authors.lastname: Klein
authors.forename: Joyce W
authors.initials: JW
authors.lastname: Lustbader
mesh_terms: D000595:Amino Acid Sequence; D001483:Base Sequence; D015153:Blotting, Western; D003593:Cytoplasm / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D000939:Epitopes; D004926:Escherichia coli / Q000378:metabolism*; D006801:Humans; D008957:Models, Genetic; D008969:Molecular Sequence Data; D011485:Protein Binding; D017510:Protein Folding; D017434:Protein Structure, Tertiary; D011962:Receptors, FSH / Q000737:chemistry*; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D013879:Thioredoxins / Q000378:metabolism
chemical_list: D000939:Epitopes; D011962:Receptors, FSH; D011993:Recombinant Fusion Proteins; D013879:Thioredoxins