Use of Internal Reflectance Infrared Spectroscopy for the in Situ Study of Supported Lipid Monolayers

Abstract

An instrument has been developed to acquire polarized attenuated total internal reflection (ATIR) mid-IR spectra of supported monomolecular films in situ at the lipid/water (L/W) interface. The monolayer film is applied to the treated surface of a trapezoidal germanium crystal as it is immersed into the subphase of a specially designed Langmuir-Blodgett trough. This approach offers several advantages over the study of monolayer films at the air/water (A/W) interface by external reflection techniques, including (1) an improved signal throughput and multiple reflections, both contributing to an improved signal-to-noise (S/N) ratio; (2) versatility with respect to the chemical nature of the surface; and (3) the ability to use standard spectral subtraction methods to compensate for absorption of the aqueous subphase. The enhanced sensitivity of this approach facilitates the study of in situ interactions of peptides with fully hydrated lipid membrane surfaces.

Phospholipid flip-flop modulated by transmembrane peptides WALP and melittin

“…The amide I absorbance visible in the spectrum (1650 cm -1 ) indicates the presence of the peptide in the bilayer film and is consistent with previous ATR-FTIR results. (Axelsen et al, 1995; Frey and Tamm, 1991; Sharon et al, 1999) As with WALP 23 , the frequency of the amide-I vibrational mode for melittin is consistent with an α-helical secondary structure. (Bellamy, 1975; Chen et al, 2007b; Flach et al, 1996; Sharon et al, 1999) For comparison, the ATR-FTIR spectrum of a DSPC bilayer without peptide is shown in Figure 2A, in which case the amide band at 1650 cm -1 is clearly absent.…”

The Structure of Human Lipoprotein A-I

“…The concentrations of DMPS used in these experiments are well below the threshold at which nonliquid lipid phases would be induced by 1 mM Ca 2ϩ (29). Polarized attenuated total internal reflection Fourier-transform (PATIR-FTIR) spectroscopy was performed on a previously described instrument (19), modified such that the internal reflection crystal is applied flat onto a monolayer at the air-water interface in a Langmuir trough (23). The angle of incidence between the IR beam and the crystal surface was 30°.…”

“…The angle of incidence between the IR beam and the crystal surface was 30°. The internal reflection crystal was treated with octadecyltrichlorosilane to render the surface hydrophobic (19,30). All spectra (1024 co-added interferograms) were collected at room temperature, using a Bio-Rad FTS-60A spectrometer in rapid-scanning mode, a liquidnitrogen-cooled MCT detector, a resolution of 2 cm…”

Promotion of Amyloid β Protein Misfolding and Fibrillogenesis by a Lipid Oxidation Product

“…39,40,43,[69][70][71] The subphase buffer in the trough consisted of 2.5 mL of 30 mM Hepes-NaOD buffer in D 2 O at pD 7.4. HNE in Hepes buffer was added where indicated.…”