Samaddar, M and Catterall, JF and Dighe, RR (1997) Expression of biologically active beta subunit of bovine follicle-stimulating hormone in the methylotrophic yeast Pichia pastoris. In: Protein Expression and Purification, 10 (3). pp. 345-355.
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Follicle-stimulating hormone (FSH), a pituitary gonadotropin, is a heterodimer composed of an alpha subunit, which is common to all the glycoprotein hormones, noncovalently associated with the hormone-specific beta subunit, The objective of the present study is to develop a recombinant DNA expression system for the beta subunit of FSH that can be applied to study structure-function relationships while producing large quantities of the hormone subunit for immunocontraceptive, clinical, and veterinary purposes. We report here the expression of biologically active bovine FSH beta (bFSH beta) in the methylotrophic yeast Pichia pastoris. The Pichia-expressed FSH beta (pFSH beta) was secreted into the culture medium and was found to be immunologically very similar to pituitary-derived ovine FSH beta. Replacement of the cognate signal peptide with the yeast alpha mating factor signal peptide increased the level of expression from 230 ng/ml (cognate signal peptide) to 4 mu g/ml (alpha mating factor signal peptide) of the culture supernatant, pFSH beta(His.tag) (pFSH beta with six histidine residues at the C terminus) was purified to apparent homogeneity using one-step nickel affinity chromatography. The molecular weight of purified pFSH beta(His.tag) was approximately 22,000, which was slightly higher than that of the pituitary-derived ovine FSH beta, pFSH beta(His.tag) could assemble with the a subunit to yield a heterodimer capable of binding to the FSH receptors and also elicit biological response,These data show that pFSH beta(His.tag) is properly folded and biologically active.
|Item Type:||Journal Article|
|Additional Information:||Copyright of this article belongs to Elsevier Science.|
|Department/Centre:||Division of Biological Sciences > Molecular Reproduction, Development & Genetics (formed by the merger of DBGL and CRBME)|
|Date Deposited:||19 Jul 2009 06:26|
|Last Modified:||19 Sep 2010 05:26|
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