Factors affecting the dissociation and aggregation of human interferon gamma.
The biologically active form of interferon gamma (IFN-gamma) is a dimer consisting of two identical non-covalently bound polypeptide chains. We have studied spectroscopically the dimer-monomer dissociation equilibrium of human recombinant IFN-gamma and have found that the monomers possess approximately 50% lower Trp quantum yield than the dimers [Boteva et al. Biochemistry 1996;35:14825]. In the present study we characterise the conformational properties of the two states--monomeric and dimeric, and analyse the effects of the salt composition of human blood plasma, physiological cations K+, Na+, Ca2+ and Mg2+ and mechanical stress on the dimer-monomer equilibrium. A medium with electrolyte composition of human blood plasma increases both the association and dissociation rate constants without shifting significantly the dimer-monomer equilibrium. The physiological cations shift the equilibrium towards dissociation of dimers into monomers by lowering the activation energy and the free energy of the process thus decreasing the stability of IFN-gamma. Mechanical stress caused by stirring of the protein solution reduces irreversibly the Trp fluorescence by 75-80% and decreases significantly the alpha-helical content and favours the aggregation.
Zlateva
T
T
CNR Centre of Metalloproteins, Department of Biology, University of Padua, Italy.
Boteva
R
R
Salvato
B
B
Tsanev
R
R
D000178:Acrylamides; D002414:Cations, Monovalent; D019281:Dimerization; D006801:Humans; D007371:Interferon-gamma / Q000737:chemistry*; D007700:Kinetics; D008670:Metals; D017433:Protein Structure, Secondary; D011994:Recombinant Proteins; D013050:Spectrometry, Fluorescence; D014364:Tryptophan
D000178:Acrylamides; D002414:Cations, Monovalent; D008670:Metals; D011994:Recombinant Proteins; D007371:Interferon-gamma; D014364:Tryptophan