Obtaining information on protein dynamics using FT-IR spectroscopy
In biological systems, protein function is dependent on spatial and temporal changes known as protein dynamics, which can be probed by amide hydrogen/deuterium (H/D) exchange. Fourier-transform infrared (FT-IR) spectroscopy is a convenient and efficient tool for determining the H/D exchange rate of proteins on a global scale. The H/D exchange process is monitored by following the apparent changes in FT-IR intensity at the amide II band maxima near 1550 cm−1. This protocol covers the principles underlying the determination of protein dynamics by FT-IR, as well as the basic steps involved in protein sample preparation and FT-IR spectra collection and detailed methods for analyzing spectra to determine the H/D exchange rate of proteins. This is a semi-quantitative method and can only be used for comparing protein dynamics under certain condition. Applications include the effects of protein mutation or protein and metal ion or ligand interactions on the protein H/D exchange rate. Typically, the procedure can be completed in 2-3 days.
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Table2 Troubleshooting table
Posted 04 Jun, 2018
Obtaining information on protein dynamics using FT-IR spectroscopy
Posted 04 Jun, 2018
In biological systems, protein function is dependent on spatial and temporal changes known as protein dynamics, which can be probed by amide hydrogen/deuterium (H/D) exchange. Fourier-transform infrared (FT-IR) spectroscopy is a convenient and efficient tool for determining the H/D exchange rate of proteins on a global scale. The H/D exchange process is monitored by following the apparent changes in FT-IR intensity at the amide II band maxima near 1550 cm−1. This protocol covers the principles underlying the determination of protein dynamics by FT-IR, as well as the basic steps involved in protein sample preparation and FT-IR spectra collection and detailed methods for analyzing spectra to determine the H/D exchange rate of proteins. This is a semi-quantitative method and can only be used for comparing protein dynamics under certain condition. Applications include the effects of protein mutation or protein and metal ion or ligand interactions on the protein H/D exchange rate. Typically, the procedure can be completed in 2-3 days.
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