Quantitative orientation of alpha-helical polypeptides by attenuated totalreflection infrared spectroscopy

Spectral simulations of polarized attenuated total reflection (ATR) spectra have been used to quantitatively determine the orientation of alpha -helic al polypeptides. Transmittance and polarized ATR spectra of five monolayers of poly-gamma -benzyl-L-glutamate (PBG) transferred by the Langmuir-Blodge tt techniques on germanium crystal have been recorded, and the dichroic rat ios of the amide I and amide II bands have been calculated. Simulations wer e performed using anisotropic optical constants of PEG in the molecular coo rdinate system and using the 4 x 4 matrix formalism of Berreman. Assuming t hat the alpha -helices are parallel to the plane of the ATR crystal, an ave rage angle of 38 degrees +/- 1 degrees between the helix axes and the withd rawing direction has been found. Simulated spectra for various orientations of the alpha -helices are then given and influence of the azimuthal and ti lt angles on the dichroic ratios of the amide bands has been investigated.