Loss of ammonia during electron-transfer dissociation of deuterated peptides as an inherent gauge of gas-phase hydrogen scrambling

TY - JOUR

T1 - Loss of ammonia during electron-transfer dissociation of deuterated peptides as an inherent gauge of gas-phase hydrogen scrambling

AU - Rand, Kasper Dyrberg

AU - Zehl, Martin

AU - Jensen, Ole

AU - Jørgensen, Thomas

PY - 2010/12/1

Y1 - 2010/12/1

N2 - The application of electron-transfer dissociation (ETD) to obtain single-residue resolution in hydrogen exchange - mass spectrometry (HX-MS) experiments has recently been demonstrated. For such measurements, it is critical to ensure that the level of gas-phase hydrogen scrambling is negligible. Here we utilize the abundant loss of ammonia upon ETD of peptide ions as a universal reporter of positional randomization of the exchangeable hydrogens (hydrogen scrambling) during HX-ETD experiments. We show that the loss of ammonia from peptide ions proceeds without depletion of deuterium when employing optimized mild electrospray ion source settings for the HX-ETD analysis of a selectively labeled model peptide and peptides derived from fully labeled β2-microglobulin. Hydrogen scrambling, as induced by excessive vibrational excitation of peptide ions during harsh declustering conditions, is easily detected by a depletion of deuterium when deuterated ammonia is lost from peptides during ETD. This straightforward method requires no modifications to the experimental workflow and has the great advantage that the occurrence of hydrogen scrambling can be directly detected in the actual peptides analyzed in the HX-ETD experiment.

AB - The application of electron-transfer dissociation (ETD) to obtain single-residue resolution in hydrogen exchange - mass spectrometry (HX-MS) experiments has recently been demonstrated. For such measurements, it is critical to ensure that the level of gas-phase hydrogen scrambling is negligible. Here we utilize the abundant loss of ammonia upon ETD of peptide ions as a universal reporter of positional randomization of the exchangeable hydrogens (hydrogen scrambling) during HX-ETD experiments. We show that the loss of ammonia from peptide ions proceeds without depletion of deuterium when employing optimized mild electrospray ion source settings for the HX-ETD analysis of a selectively labeled model peptide and peptides derived from fully labeled β2-microglobulin. Hydrogen scrambling, as induced by excessive vibrational excitation of peptide ions during harsh declustering conditions, is easily detected by a depletion of deuterium when deuterated ammonia is lost from peptides during ETD. This straightforward method requires no modifications to the experimental workflow and has the great advantage that the occurrence of hydrogen scrambling can be directly detected in the actual peptides analyzed in the HX-ETD experiment.

U2 - 10.1021/ac101889b

DO - 10.1021/ac101889b

M3 - Journal article

C2 - 21033729

SN - 0003-2700

VL - 82

SP - 9755

EP - 9762

JO - Industrial And Engineering Chemistry Analytical Edition

JF - Industrial And Engineering Chemistry Analytical Edition

IS - 23

ER -