Conformer selection of protein ions by ion mobility in a triple quadrupole mass spectrometer

Abstract

Electrospray mass spectra of multiply charged protein molecules show two distinct charge state distributions proposed to correspond to a more highly charged, open conformational form and a lower charged, folded form. Elastic collisions carried out in the radiofrequency-only collision cell of a triple quadrupole mass spectrometer have dramatic effects on the appearance of the mass spectra. The different cross sectional areas of the conformers allow preferential selection of one charge state distribution over the other on the basis of ion mobility. Preferential selection is dependent on the nature and pressure of the target gas as well as the nature of the protein. In the case of positively charged horse heart apomyoglobin (MW 16,951 da), a high charge state distribution centered around (M + 20H)(20+) predominates at low target gas pressures and a second distribution centered around (M + 10H)(10+) predominates at high target gas pressures. Bimodal distributions are observed at intermediate pressures and, remarkably, charge states between the two distributions are not effectively populated under most of the conditions examined. Hard sphere collision calculations show large differences in collision frequencies and in the corresponding kinetic energy losses for the two conformational states and they demonstrate that the observed charge state selectivity can be explained through elastic collisions.

Applying a dynamic method to the measurement of ion mobility

“…A quick comparison of the method we used with the experiments for determination of collision cross sections in the collision cell of tandem quadrupole mass spectrometers [17][18][19] shows the following: In the stationary gas of the collision cell the ions with smallest cross sections pass the stopping potential (a low pass filter in terms of cross sections) and are detected, while the collisions with ions of larger cross sections reduce their translational energy so that they cannot reach the analyzer quadrupole. However, in our experiments, where the ions are transported by the gas flow, ions with the largest cross sections pass the barrier (high pass filter) and are detected.…”

“…Interesting experiments for measuring collision cross sections of ions were also reported by Covey and Douglas, [17], by Cox et al [18], and by Gill et al [19], who used tandem quadrupole mass spectrometers. In these experiments, ions traveling through the collision cell lost a certain portion of their kinetic energy depending on their cross sections.…”

“…Larger ions suffered a more significant loss of energy in the collision cell. They were unable to enter the third quadrupole due to its DCoffset voltage which was set higher than the voltage of the collision cell [17,18]. This effect was used for checking the translational energy and obtaining information about collision cross section of ions.…”

Conformations of biopolymers in the gas phase: a new mass spectrometric method

“…This truism has led to the development of several collisional methods for extracting structural information. The first of these involves determination of collisional cross sections for mass-selected ions under relatively low pressure collision conditions [7,8]. Typically, ions are formed by electrospray, mass selected, impacted on neutral gas in a collision cell, and then detected with a second mass spectrometer.…”

Conformations and folding of lysozyme ions in vacuo.

“…The development of new ionization techniques, including matrix-assisted laser desorption/ionization (2,3) and electrospray ionization (ESI) (4), has made possible the formation of intact molecular ions of large biomolecules, the structures of which can be investigated both in solution as well as in the complete absence of solvent using an impressive array of mass spectrometric techniques (5)(6)(7)(8)(9)(10)(11)(12)(13)(14)(15)(16)(17)(18)(19)(20)(21)(22). In solution, information about protein conformation can be inferred from charge state distributions observed in electrospray mass spectra (9)(10)(11)(12)(13), as well as from shifts in mass upon solution-phase deuterium exchange (14,15).…”