Infrared spectroscopy, a tool for probing bacterial peptidoglycan. Potentialities of infrared spectroscopy for cell wall analytical studies and rejection of models based on crystalline chitin

Abstract

Infrared spectroscopic measurements are used to obtain insights into the three-dimensional architecture of peptidoglycan (murein), the rigid component of almost all bacterial cell walls. The infrared spectra of various types of peptidoglycans (including all chemotypes and examples of the so called A and B groups) were compared to each other and to those obtained from crystalline chitin. All peptidoglycans investigated exhibited very similar infrared spectra. In particular the conformationally sensitive amide A, I and II absorption bands were found to be constantly centered around 3300 cm-1, 1657 cm-1 and 1534 cm-1 respectively; furthermore, the spectral region between 1200 cm-1 and 800 cm-1, characterized by several strong absorption bands connected to complex sugar ring modes, proved to be remarkably uniform. Additionally the infrared spectra remained significantly constant between -175 degrees C and + 75 degrees C and turned out to be rather independent of sample preparation (solvent replacement, freeze-drying and film producing). An analysis of band half-widths revealed no high crystalline state of order of peptidoglycan. On the basis of band positions and half-widths of amide bands, regular conformations like alpha helices of beta pleated sheets could be excluded. Several distinctive, fingerprint-like spectral features of the various murein samples permitted a facile identification of individual peptidoglycans. Moreover, infrared spectroscopy seems to be very promising as an analytical tool, e.g. for tracing variations of cell wall structure, detecting conformational changes and estimating crosslinking indices in a quick and simple way. The comparative analysis of amide band positions and band half-widths yielded substantial differences between infrared spectra of chitin and murein, thus rejecting previous models based on the assumption of a nearly crystalline chitin-like structure of the glycan chains of murein.