title: Effects of ionic strength on the thermal unfolding process of granulocyte-colony stimulating factor.
abstract: This paper reports the effect of ionic strength on the process of thermal unfolding of recombinant methionyl human granulocyte-colony stimulating factor (rmethuG-CSF) at acid pH. We previously reported that the protein aggregates were formed at the highest temperature at pD 2.1 in the pD range of 5.5-2.1 and that the aggregation proceeded a little at pD 2.1 because of the strong repulsive interaction between the unordered structures that play the role of a precursor for the aggregation. In the present study temperature-dependent IR spectra and far-UV CD spectra were measured for rmethuG-CSF in aqueous solutions containing various concentrations of NaCl at acid pH. Second derivative and curve-fitting analysis were performed to examine the obtained IR spectra. The results revealed that the structure of rmethuG-CSF becomes less stable with increasing ionic strength at all pDs investigated (pD 2.1, 2.5, and 4.0). We have also demonstrated that, at pD 2.1, the temperature at which the protein aggregation starts becomes lower and that the amount of the aggregates becomes larger with the addition of NaCl. This is probably because the addition of NaCl masks the repulsive electrostatic interaction between the unordered structures.
authors.affiliation: Product Development Section, CMC R&D Laboratories, Pharmaceutical Division, Kirin Brewery Co., Ltd, Takasaki, Gunma 370-0013.
authors.forename: Katsuyoshi
authors.initials: K
authors.lastname: Yamazaki
authors.forename: Takafumi
authors.initials: T
authors.lastname: Iwura
authors.forename: Rika
authors.initials: R
authors.lastname: Ishikawa
authors.forename: Yukihiro
authors.initials: Y
authors.lastname: Ozaki
mesh_terms: D002942:Circular Dichroism; D000069585:Filgrastim; D016179:Granulocyte Colony-Stimulating Factor / Q000737:chemistry / Q000378:metabolism*; D006801:Humans; D009994:Osmolar Concentration; D011489:Protein Denaturation; D011994:Recombinant Proteins; D013056:Spectrophotometry, Ultraviolet; D017550:Spectroscopy, Fourier Transform Infrared; D055672:Static Electricity
chemical_list: D011994:Recombinant Proteins; D016179:Granulocyte Colony-Stimulating Factor; D000069585:Filgrastim