Protein Conformation in Amorphous Solids by FTIR and by Hydrogen/Deuterium Exchange with Mass Spectrometry
| Autor: | Elizabeth M. Topp, Sandipan Sinha, Todd D. Williams, Yunsong Li |
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| Jazyk: | angličtina |
| Předmět: |
Resolution (mass spectrometry)
Protein Conformation Electrospray ionization Biophysics Analytical chemistry Lactoglobulins Mass spectrometry Mass Spectrometry Spectroscopy Fourier Transform Infrared Concanavalin A Fourier transform infrared spectroscopy Myoglobin Chemistry technology industry and agriculture Proteins Deuterium Exchange Measurement Ribonuclease Pancreatic Cadherins Protein Structure Tertiary Amorphous solid Attenuated total reflection Muramidase Hydrogen–deuterium exchange lipids (amino acids peptides and proteins) Nuclear chemistry |
| Zdroj: | Biophysical Journal. (12):5951-5961 |
| ISSN: | 0006-3495 |
| DOI: | 10.1529/biophysj.108.139899 |
| Popis: | Solid-state hydrogen/deuterium exchange (ssHDX) with electrospray ionization mass spectrometry (ESI-MS) and Fourier transform infrared (FTIR) spectroscopy were used to assess protein conformation in amorphous solids. Myoglobin, lysozyme, beta-lactoglobulin, ribonuclease A, E-cadherin 5, and concanavalin A were co-lyophilized with carbohydrates (trehalose, raffinose, and dextran 5000), linear polymers (polyvinyl alcohol and polyvinyl pyrrolidone) or guanidine hydrochloride (negative control). For ssHDX, samples were exposed to D2O vapor at 33% relative humidity and room temperature, and then reconstituted at low temperature (4 degrees C) and pH 2.5 and analyzed by ESI-MS. Peptic digestion of selected proteins was used to provide region-specific information on exchange. FTIR spectra were acquired using attenuated total reflectance. FTIR and ssHDX of intact proteins showed preservation of structure by raffinose and trehalose, as indicated by FTIR band intensity and protection from exchange. ssHDX of peptic digests further indicated that these protective effects were not exerted uniformly along the protein sequence but were observed primarily in alpha-helical regions, a level of structural resolution not afforded by FTIR. The results thus demonstrate the utility of HDX with ESI-MS for analyzing protein conformation in amorphous solid samples. |
| Databáze: | OpenAIRE |
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