TY - JOUR
T1 - Effect of poly and mono unsaturated fatty acids on stability and structure of recombinant S100A8/A9
AU - Asghari, Hamideh
AU - Goodarzvand Chegini, Koorosh
AU - Amini, Abbas
AU - Gheibi, Nematollah
PY - 2016
Y1 - 2016
N2 - Recombinant pET 15b vectors containing the coding sequences S100A8 and S100A9 are expressed in Escherichia coli BL21 (DE3) and purified using Ni-NTA affinity chromatography. The structural changes of S100A8/A9 complex are analyzed upon interaction with poly/mono unsaturated fatty acids (UFAs). The thermodynamic values, Gibbs free energy and the protein melting point, are obtained through thermal denaturation of protein both with and without UFAs by thermal scanning of protein emission using the fluorescence spectroscopy technique. The far-ultraviolet circular dichroism spectra show that all studied unsaturated fatty acids, including arachidonic, linoleic, alpha-linolenic and oleic acids, induce changes in the secondary structure of S100A8/A9 by reducing the alpha-helix and beta-sheet structure. The tertiary structure of S100A8/A9 has fluctuations in the fluorescence emission spectra after the incubation of protein with UFAs. The blue shift of emission maximum wavelength and the increase in fluorescence intensity of anilino naphthalene-8-sulfonic acid confirm the partial unfolding is caused by the conformational changes in the tertiary structure in the presence of UFAs. The structural changes in S100A8/A9 and its lower stability in the presence of UFAs may be necessary for S100A8/A9 to play a biological role in the inflammatory milieu.
AB - Recombinant pET 15b vectors containing the coding sequences S100A8 and S100A9 are expressed in Escherichia coli BL21 (DE3) and purified using Ni-NTA affinity chromatography. The structural changes of S100A8/A9 complex are analyzed upon interaction with poly/mono unsaturated fatty acids (UFAs). The thermodynamic values, Gibbs free energy and the protein melting point, are obtained through thermal denaturation of protein both with and without UFAs by thermal scanning of protein emission using the fluorescence spectroscopy technique. The far-ultraviolet circular dichroism spectra show that all studied unsaturated fatty acids, including arachidonic, linoleic, alpha-linolenic and oleic acids, induce changes in the secondary structure of S100A8/A9 by reducing the alpha-helix and beta-sheet structure. The tertiary structure of S100A8/A9 has fluctuations in the fluorescence emission spectra after the incubation of protein with UFAs. The blue shift of emission maximum wavelength and the increase in fluorescence intensity of anilino naphthalene-8-sulfonic acid confirm the partial unfolding is caused by the conformational changes in the tertiary structure in the presence of UFAs. The structural changes in S100A8/A9 and its lower stability in the presence of UFAs may be necessary for S100A8/A9 to play a biological role in the inflammatory milieu.
KW - Escherichia coli
KW - circular dichroism
KW - recombinant antibodies
KW - stability
KW - unsaturated fatty acids
UR - http://handle.uws.edu.au:8081/1959.7/uws:32966
U2 - 10.1016/j.ijbiomac.2015.11.065
DO - 10.1016/j.ijbiomac.2015.11.065
M3 - Article
SN - 0141-8130
VL - 84
SP - 35
EP - 42
JO - International Journal of Biological Macromolecules
JF - International Journal of Biological Macromolecules
ER -