Expression of biologically active β subunit of bovine follicle-stimulating hormone in the methylotrophic yeast Pichia pastoris

Abstract

Follicle-stimulating hormone (FSH), a pituitary gonadotropin, is a heterodimer composed of an a subunit, which is common to all the glycoprotein hormones, noncovalently associated with the hormone-specific β subunit. The objective of the present study is to develop a recombinant DNA expression system for the β 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 FSHβ (bFSHβ) in the methylotrophic yeast Pichia pastoris.ThePichia-expressed FSHβ (pFSHβ) was secreted into the culture medium and was found to be immunologically very similar to pituitary-derived ovine FSHβ. Replacement of the cognate signal peptide with the yeast α mating factor signal peptide increased the level of expression from 230 ng/ml (cognate signal peptide) to 4 μg/ml (α mating factor signal peptide) of the culture supernatant. pFSHβ^His.tag(pFSHβ with six histidine residues at the C terminus) was purified to apparent homogeneity using one-step nickel affinity chromatography. The molecular weight of purified pFSHβ^His.tag was approximately 22,000, which was slightly higher than that of the pituitary-derived ovine FSHβ. pFSHβ^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 pFSHβ^His.tag is properly folded and biologically active.