Staff Publications

Staff Publications

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    'Staff publications' is the digital repository of Wageningen University & Research

    'Staff publications' contains references to publications authored by Wageningen University staff from 1976 onward.

    Publications authored by the staff of the Research Institutes are available from 1995 onwards.

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Record number 307864
Title High-level expression of biologically active recombinant bovine follicle stimulating hormone in a baculovirus system
Author(s) Wiel, D.F.M. van de; Rijn, P.A. van; Meloen, R.H.; Moormann, R.J.M.
Source Journal of Molecular Endocrinology 20 (1998)1. - ISSN 0952-5041 - p. 83 - 98.
DOI https://doi.org/10.1677/jme.0.0200083
Department(s) ID Lelystad, Institute for Animal Science and Health
Publication type Refereed Article in a scientific journal
Publication year 1998
Abstract Superovulation treatment of cows can benefit from the application of very pure recombinant bovine FSH (rbFSH), which is produced in nonmammalian cells. rbFSH is completely free of LH, and therefore can possibly reduce the variability in the results of superovulation. Furthermore, it does not contain brain-tissue-derived proteins and, when produced under serum-free conditions, it is free of other mammalian substances or potentially infectious material. We have produced rbFSH in insect cells, with the ultimate aim of inducing superovulation in cattle. Sf21 insect cells were coinfected with two recombinant baculoviruses, containing the cDNAs of bovine FSH α- and β-subunits respectively. High levels of production of bioactive rbFSH were obtained after cloning cDNA that contained a major part of the 3' untranslated region of the bFSHβ gene. Maximum production of rbFSH 1-5 μg/ml (as measured by immunoassay) was obtained 70-90 h after infection. The recombinant material was highly potent in two in vitro bioassays, giving biological activities of 13 IU/ml (Y1 cell rounding assay), 22 IU/ml (Y1 cell cAMP assay), and 23 IU/ml (bovine oocyte maturation inhibition assay), and had a lower but significant activity of 6 IU/ml in the rat Sertoli cell assay. rbFSH was purified by immunoaffinity chromatography, using a monoclonal antibody directed against the human FSH β-subunit. The purified heterodimer appeared to be homogeneous by SDS-PAGE, whereas the free β- subunit appeared as a doublet, possibly indicating differently glycosylated forms. Intact heterodimer and both subunits were further identified by western blot analysis, and showed apparent molecular masses of 20 kDa (α- subunit), 23 kDa (β-subunit) and 32.5 kDa (heterodimer). This insect-cell- produced rbFSH did not bind to wheat germ agglutinin, thus indicating that glycosidic side-chains may not contain terminal sialic acid. The relevance of a large 3' untranslated region in bFSHβ cDNA to the level of production of rbFSH, and the possible implications of the pattern of glycosylation for the biological activity of the recombinant hormone are discussed.
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