False EX1 signatures caused by sample carryover during HX MS analyses

TY - JOUR

T1 - False EX1 signatures caused by sample carryover during HX MS analyses

AU - Fang, Jing

AU - Rand, Kasper Dyrberg

AU - Beuning, Penny J

AU - Engen, John R

PY - 2011/4/30

Y1 - 2011/4/30

N2 - Discovery of EX1 kinetics in hydrogen exchange (HX) mass spectrometry (MS) experiments is rare. Proteins follow the EX1 kinetic regime when cooperative unfolding events simultaneously expose multiple residues to solvent such that they all become deuterated together before the region is able to refold. A number of factors can contribute to what we call "false EX1" in which it appears as though EX1 occurs in a protein when it probably does not. One of the contributors to false EX1 is peptide carryover between chromatographic runs. In this work, we explore the origins of peptide carryover in HX MS, describe how carryover causes mass spectra to indicate false EX1 kinetics and then describe an optimized washing protocol that can be used to eliminate peptide carryover. A series of solvent injections was developed and found to efficiently eliminate carryover signatures such that analysis of deuterium incorporation could be reliably followed for two proteins prone to high carryover.

AB - Discovery of EX1 kinetics in hydrogen exchange (HX) mass spectrometry (MS) experiments is rare. Proteins follow the EX1 kinetic regime when cooperative unfolding events simultaneously expose multiple residues to solvent such that they all become deuterated together before the region is able to refold. A number of factors can contribute to what we call "false EX1" in which it appears as though EX1 occurs in a protein when it probably does not. One of the contributors to false EX1 is peptide carryover between chromatographic runs. In this work, we explore the origins of peptide carryover in HX MS, describe how carryover causes mass spectra to indicate false EX1 kinetics and then describe an optimized washing protocol that can be used to eliminate peptide carryover. A series of solvent injections was developed and found to efficiently eliminate carryover signatures such that analysis of deuterium incorporation could be reliably followed for two proteins prone to high carryover.

U2 - 10.1016/j.ijms.2010.06.039

DO - 10.1016/j.ijms.2010.06.039

M3 - Journal article

C2 - 21643454

SN - 1387-3806

VL - 302

SP - 19

EP - 25

JO - International Journal of Mass Spectrometry

JF - International Journal of Mass Spectrometry

IS - 1-3

ER -