Two dimensional near-infrared-infrared correlation spectroscopy studies of protein hydration and denaturation

点击翻译按钮获取中文摘要

Attenuated total reflection (ATR)/infrared (IR) spectra were measured for human serum albumin (HSA) in aqueous solutions over a pH range of5.0-3.2.Generalized two-dimensional (2D) correlation analysis was applied to the amide I region of the spectra to investigate pH-dependent changes in secondary structures and in hydrogen bondings of side chains of HSA. The synchronous and asynchronous 2D spectra were generated from the pH-dependent spectral variations for the three states of HSA, the N isomeric form (pH 5.0-4.4), the N-F transition (pH 4.6-3.8), and the F isomeric form (pH 3.8-3.0). The most interesting finding in the 2D spectra is identification offour bands at 1740, 1715, 1705, and 1696 cm1 due to a OO stretching mode of free and hydrogen bonded (weak, medium, and strong) COOH groups ofHSA. "FT-NIR spectra of HSA at different concentration in aqueous solutions have been measured from 45-80 ーC. Concentration dependent 2D correlation analysis at different temperature were applied to … More these spectra in two range 5500-7500 cm1 and 4200-4900 cm"1, respectively. Absorption bands arising from the combination vibrations of O-H stretching in water dominate the NIR spectra of the aqueous solution, especially at the 5500-7500 cm'1 region. The hydration and inter-, intra-molecular interaction changes, mainly due to the variation of the hydrogen bonding between different molecular units have been provided by the 2D-correlation patterns in thisrange. "Molten globule-like states of ovalbumin (OVA) in acid aqueous solutions have been investigated by generalized 2D FT-NIR correlation spectroscopy. This new method allows us to explore in changes in hydration and secondary structure simultaneously.FT-NIR spectra were measured for OVA aqueous solutions with concentrations of 1, 2, 3, 4 and 5 wt% over a pH range of2.4-5.4.Concentration-perturbed 2D correlation spectra were calculated for'the spectra in the 4850-4200 cm1 and 7500-5350 cm"1 regions at different pH. The 2D-NIR synchronous spectrum in the 4850-4200 cm"1 region shows a significant change upon going from pH 5.4 to pH 3.6.An autopeak at 4265 cm"1 due to a combination of symmetric CH2 stretching mode and CH2 bending mode of side chains seen at pH 5.0 disappears completely in the synchronous spectrum at pH 3.6.This suggests that some amino acid residues of OVA are subjected to micro environmental changes with decreasing pH." Less