title: High-level expression of biologically active human prolactin from recombinant baculovirus in insect cells.
abstract: We examined the feasibility of high-level production of recombinant human prolactin, a multifunctional protein hormone, in insect cells using a baculovirus expression system. The human prolactin cDNA with and without the secretory signal sequence was cloned into pFastBac1 baculovirus vector under the control of polyhedrin promoter. Prolactin was produced upon infection of either Sf9 or High-Five cells with the recombinant baculovirus containing the human prolactin cDNA. The production of recombinant prolactin varied from 20 to 40 mg/L of monolayer culture, depending on the cell types. The prolactin polypeptide with its own secretory signal was secreted into the medium. N-terminal amino acid sequence analysis of the recombinant polypeptide purified from the culture medium indicated that the protein was processed similar to human pituitary prolactin. Carbohydrate analysis of the purified protein indicated that a fraction of the recombinant prolactin made in insect cells appeared to be glycosylated. Also, both secreted and nonsecreted forms of the recombinant prolactin in insect cells were biologically equivalent to the native human prolactin (pituitary derived) in the Nb2 lymphoma cell proliferation assay.
authors.affiliation: Ross Products Division, Abbott Laboratories, Columbus, Ohio 43215, USA.
authors.forename: T
authors.initials: T
authors.lastname: Das
authors.forename: P W
authors.initials: PW
authors.lastname: Johns
authors.forename: V
authors.initials: V
authors.lastname: Goffin
authors.forename: P
authors.initials: P
authors.lastname: Kelly
authors.forename: B
authors.initials: B
authors.lastname: Kelder
authors.forename: J
authors.initials: J
authors.lastname: Kopchick
authors.forename: K
authors.initials: K
authors.lastname: Buxton
authors.forename: P
authors.initials: P
authors.lastname: Mukerji
mesh_terms: D000818:Animals; D016367:Baculoviridae / Q000235:genetics*; D002455:Cell Division; D002460:Cell Line; D003001:Cloning, Molecular; D004591:Electrophoresis, Polyacrylamide Gel; D015870:Gene Expression; D006031:Glycosylation; D006801:Humans; D009844:Oligosaccharides / Q000032:analysis; D011388:Prolactin / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism*; D011994:Recombinant Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D020539:Sequence Analysis, Protein; D018411:Spodoptera / Q000166:cytology / Q000821:virology; D014407:Tumor Cells, Cultured
chemical_list: D009844:Oligosaccharides; D011994:Recombinant Proteins; D011388:Prolactin