Infrared Absorption Intensity Analysis as a New Tool for Investigation of Salt Effect on Proteins
- Received Date: 2009-08-05
- Infrared intensity, Salt effect, Fourier transform infrared spectroscopy, Secondary structure, Thermodynamic constant
Abstract: The native protein structures in buffer solution are maintained by the electrostatic force as well as the hydrophobic force, salt ions play an important role in maintaining the protein native structures, and their effect on the protein stability has attracted tremendous interests. Infrared spectroscopy has been generally used in molecular structure analysis due to its fingerprint resolution for different species including macromolecules as proteins. However spectral intensities have received much less attention than the vibrational frequencies. Here we report that the spectral intensities of protein amide I band, the finger prints for the protein secondary structures, are very sensitive to the local electric field known as Onsager reaction field caused by salt ions. IR absorbance thermal titrations have been conducted for a series of samples including simple water soluble amino acids, water soluble monomeric protein cytochrome c and dimeric protein DsbC and its single-site mutant G49R. We found that at lower temperature range (10-20 oC), there exists a thermal activated salting-in process, where the IR intensity increases with a rise in the temperature, corresponding to the ions binding of the hydrophobic surface of protein. This process is absent for the amino acids. When further raising the temperature, the IR intensity decreases, this is interpreted as the thermal activated breaking of the ion-protein surface binding. Applying Van't Hoff plot to the thermal titration curves, the thermodynamic parameters such as ?H and ?S for salting-in and ion unbinding processes can be derived for various protein secondary structural components, revealing quantitatively the extent of hydrophobic interaction as well as the strength of the ion-protein binding.
|Citation:||Heng Li, Yan-yan Xu, Yu-xiang Weng. Infrared Absorption Intensity Analysis as a New Tool for Investigation of Salt Effect on Proteins[J]. Chinese Journal of Chemical Physics , 2009, 22(6): 556-562. doi: 10.1088/1674-0068/22/06/556-562|