Characterization of a deswapped triple mutant bovine odorant binding protein.

The stability and functionality of GCC-bOBP, a monomeric triple mutant of bovine odorant binding protein, was investigated, in the presence of denaturant and in acidic pH conditions, by both protein and 1-aminoanthracene ligand fluorescence measurements, and compared to that of both bovine and porcine wild type homologues. Complete reversibility of unfolding was observed, though refolding was characterized by hysteresis. Molecular dynamics simulations, performed to detect possible structural changes of the monomeric scaffold related to the presence of the ligand, pointed out the stability of the β-barrel lipocalin scaffold.

Dissociation and unfolding of bovine odorant binding protein at acidic pH.

Dissociation of bovine odorant binding protein (bOBP) dimers to monomers at pH 2.5 has been confirmed through size exclusion chromatography experiments. Moreover, structural and binding properties of the acidic monomer and neutral dimer have been compared using a combination of experimental (circular dichroism and fluorescence) and computational (molecular dynamics) techniques.

Unfolding and refolding of porcine odorant binding protein in guanidinium hydrochloride: equilibrium studies at neutral pH.

Unfolding and refolding studies on porcine odorant binding protein (pOBP) have been performed at pH 7 in the presence of guanidinium hydrochloride (GdnHCl). Unfolding, monitored by following changes of protein fluorescence and circular dichroism (CD), was found to be a reversible process, in terms of recovered structure and function. The equilibrium transition data were fitted by a simple two-state sigmoidal function of denaturant concentration and the thermodynamic folding parameters, derived from the two techniques, were very similar (average values: C(1/2) approximately 2.

Reversible unfolding of bovine odorant binding protein induced by guanidinium hydrochloride at neutral pH.

An analysis of the unfolding and refolding curves at equilibrium of dimeric bovine odorant binding protein (bOBP) has been performed. Unfolding induced by guanidinium chloride (GdnHCl) is completely reversible as far as structure and ligand binding capacity are concerned. The transition curves, as obtained by fluorescence and ellipticity measurements, are very similar and have the same protein concentration-independent midpoint (C1/2 approximately 2.