Fourier transform infrartd (FTIR) spectroscopy is an established tool for the structural character- ization of proteins. However, many potential pitfalls exist for the unwary investigator. In this review we critically assess the application of FIlR spectroscopy to the determination of protein structure by (1) outlining the principles underlying protein secondary structure determination by FZZR spectroscopy. (2) highhghting the situations in which FZZR spectroscopy should be considered the technique of choice, (3) discussing the manner in which experiments should be conducted to derive as much physiologically relevant information as possible, and (4) outlining current methods for the determination of secondary structure from infrared spectm of proteins,

The Use and Misuse of FTIR Spectroscopy in the Determination of Protein Structure

Amide modes and protein conformation.

The basic knowledge accumulated over the last twenty years on the different vibrations of polypeptides were reviewed in Chapter 8. Because of the complexity of naturally occurring proteins, most of these data have been obtained from the study of model compounds, from simple amino acid derivatives to large synthetic polypeptides, which can be crystallized in a single secondary structure. This chapter covers biologically synthesized proteins. Data on this subject are much more recent because the advent of the new generation of Fourier transform spectrophotometers only now provides high quality spectra. Simultaneously, manipulations of the spectra have been made possible by the concomitant digitalization of the spectra and the availability of low cost computers in laboratories. It was only in 1986 that the race for determination of secondary structure from manipulated IR spectra started with a paper by Byler and Susi (1986), although it is only fair to say that the results of several attempts to obtain secondary structures had been published before. The number of papers using infrared spectros-copy (IR) to obtain secondary structures has been growing exponentially ever since. One purpose of the present review is to point out, through the description and the comparison of the different methods, that interpretation of the results still needs caution. Indeed, while some spectral features of the main secondary structures are well established, others are not. Moreover, no agreement exists on a “correct” mathematical treatment of the spectra. Both the intrinsic uncertainties in the assignments and the methodological diversity open the door to flawed conclusions if the user is not properly aware of these problems. Such warnings have been issued previously (Haris and Chapman, 1992; Surewicz et al., 1993; Haris and Chapman, 1992).

R. Mendelsohn

Papers

Thermal denaturation of globular proteins. Fourier transform-infrared studies of the amide III spectral region