Quantitative analysis of protein adsorption on a planar surface by Fourier transform infrared spectroscopy: lysozyme adsorbed on hydrophobic silicon-containing polymer.
The adsorption of hen egg white lysozyme onto a solid polytris(trimethylsiloxy)silylstyrene (pTSS) surface from a D(2)O solution at pD 7 containing 100 mM NaCl and 10 mM sodium deuterated phosphate was monitored at 25 degrees C by Fourier transform infrared spectroscopy using the attenuated total reflection (ATR) method. The infrared spectrum attributed to only the adsorbed lysozyme was derived from the observed spectrum, and the amount of adsorbed lysozyme was determined as a function of time and lysozyme concentration. The kinetics of adsorption could be decomposed into two components, one of which was a process with a time constant of larger than 4 h(-1) and the other was a process with one of about 0.1 h(-1). These spectra showed that the lysozyme adsorbed in the faster process had a higher beta-structure content than the dissolved lysozyme. It was also found that the slower adsorption induced some conformational change in the lysozyme adsorbed in the faster process and/or that adsorbed in the slower process. After adsorption for 24 h, the pTSS surface was rinsed out with lysozyme-free solution. The resultant spectra of the surface indicated that the lysozyme adsorbed in the faster process was bound irreversibly on the surface and was changed to a conformer with a higher beta-structure content during the slower process. The experimental procedures and the theoretical applications for such a quantitative analysis in the ATR spectroscopic method are presented in detail.
Yokoyama
Yasuhiro
Y
Faculty of Engineering, Gifu University, 1-1 Yanagido, Gifu 501-1193, Japan.
Ishiguro
Ryo
R
Maeda
Hirotaka
H
Mukaiyama
Mayumi
M
Kameyama
Keiichi
K
Hiramatsu
Koichi
K
D000327:Adsorption; D000818:Animals; D002645:Chickens; D057927:Hydrophobic and Hydrophilic Interactions; D009113:Muramidase / Q000737:chemistry* / Q000378:metabolism*; D011108:Polymers / Q000737:chemistry*; D011487:Protein Conformation; D012825:Silicon / Q000737:chemistry*; D017550:Spectroscopy, Fourier Transform Infrared / Q000379:methods; D013499:Surface Properties
D011108:Polymers; C421864:hen egg lysozyme; D009113:Muramidase; D012825:Silicon