A Fourier Transform Infrared Spectroscopy Technique to Study Peptide Self-Assembly

Abstract

Source: Neves, R., et al. Synthesis and Characterization of 1,2-Dithiolane Modified Self-Assembling Peptides. J. Vis. Exp. (2018)

This video demonstrates the self-assembly of amyloid peptides into a beta sheet-rich supramolecular structure using Fourier transform infrared spectroscopy. An infrared beam is passed through a crystal with a high refractive index, and energy absorption across the interface of the crystal and the sample is recorded. The secondary structures of the sample proteins are identified by analyzing their characteristic absorption peaks at specific regions of the infrared spectrum, which aids in identifying the formation of the supramolecular assembly.

Protocol

1. Characterization of Supramolecular Self-Assembly Structures Formation of Amyloid Fibers To prepare a self-assembly solution, weigh out 1 mg of the peptide powder using an analytical balance. Dissolve into a mixture (pH 7.5) of 20% acetonitrile and 10 mM (4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid (HEPES) in a 1.5 mL microcentrifuge tube, to a final concentration of 1 mg/mL peptide assembly mixture. Vortex the assembly solution and leave it to assemble at room temperatur…

Representative Results

Figure 1. Supramolecular characterization of 1,2-dithiolane modified peptide. (A) FT-IR of 1 mg/mL 1,2-dithiolane-KLVFFAQ-NH2 fibers assembled in 10mM HEPES, pH 7.5 in 20% CH3CN. The peak at 1627 cm-1 is consistent with the peptides assembled in a β-sheet conformation. (B) CD of 1 mg/mL 1,2-dithiolane-KLVFFAQ-NH2 fibers assembled i…

Materials

Acetonitrile	Fisher Scientific	A998-4	Flammable; irritating to eyes; Use personal protective equipment; Use only under a fume hood; keep away from open flame or hot surface; if contacted rinse wiith water for at least 15 minutes and obtain medical attention
4-(2-Hydroxyethyl)piperazine-1-ethanesulfonic acid	Acros Organics	AC172571000	Do not inhale; use outdoors or in well-ventilated area
Fourier-Transform Infrared Spectrometer	Alpha Tensor, Bruker