Surface FTIR Techniques to Analyze the Conformation of Proteins/ Peptides in H2O Environment

Abstract

Proteins/peptides, which are involved in various biochemical processes in biological systems, contain infrared (IR) active vibrations. Among all the IR absorption bands of proteins/peptides, the amide I band arises mainly from the stretching vibration of the carbonyls (C=O) in backbone amide bonds and is sensitive to the conformations (such as α– helix, β–sheet, unstructured conformation, and so on) in a protein/peptide. Therefore, the amide I band has been used to monitor the biophysical/biochemical behavior of proteins/peptides in biological samples (e.g., living cells or tissues). However, obtaining reproducible IR spectra of proteins/peptides in H2O solution was challenging by direct transmission measurement using a liquid cell with milli-meter level path length, due to the intensive IR absorption of H2O around 1620 cm-1 which overlaps the amide I band. Thus, lots of the IR spectra of proteins/peptides were accomplished in D2O, which has IR absorption around 1200 cm-1. Since D2O may not be a favorable solvent for biological samples, the position of the amide I band of various conformations was needed as a reference for biological samples. Consequently, various surface FTIR techniques (such as Infrared Reflection-Absorption Spectroscopy or IRRAS, and Attenuated Total Reflection or ATR) have been developed to obtain the IR spectra of proteins/peptides in H2O environment and have been reviewed here.