Hydrogen-deuterium exchange analyzed by matrix-assisted laser desorption-ionization mass spectrometry and the HET-s prion model.
Hydrogen/deuterium (H/D) exchange analyzed by mass spectrometry (HXMS) is a valuable tool for the investigation of protein conformation and dynamics. After exchange, the sample is generally submitted to electrospray ionization for mass analysis. Matrix-assisted laser desorption ionization (MALDI) has been used in a limited number of studies but has several significant advantages that include simplification of the spectra attributable to a predominance of singly charged ions, speed of analysis, sensitivity, and low H/D back-exchange level. MALDI-HXMS has been used to study amyloid aggregates from the HET-s prion protein. Our results underline the ability of this method to determine solvent accessibility within the amyloid aggregates, reaching a resolution of one to four amino acids. To achieve a complete peptide mass fingerprint of the protein, we have taken benefits of an ion trap operating in liquid chromatography-MS/MS mode. MALDI time-of-flight-MS was then used to determine deuterium incorporation within each peptide along the sequence of HET-s. The combined advantages of these two instruments yield a suitable solution for HXMS experiments that require highly resolved peptide mass fingerprints, high sensitivity, and speed of analysis for deuterium incorporation measurements.
Nazabal
Alexis
A
Swiss Federal Institute of Technology, ETH, Department of Chemistry and Applied Biosciences, Zurich, Switzerland.
Schmitter
Jean-Marie
JM
D041961:Deuterium Exchange Measurement / Q000379:methods*; D005656:Fungal Proteins / Q000737:chemistry*; D010434:Pepsin A / Q000378:metabolism; D010446:Peptide Fragments / Q000737:chemistry; D010449:Peptide Mapping / Q000379:methods; D011328:Prions / Q000737:chemistry*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods*
D005656:Fungal Proteins; C083891:HET-S protein, Podospora anserina; D010446:Peptide Fragments; D011328:Prions; D010434:Pepsin A