Protein structure information from mass spectrometry? Selective titration of arginine residues by sulfonates

166

This work experimentally verifies and proves the two long since postulated matrix-assisted laser desorption/ionization (MALDI) analyte protonation pathways known as the Lucky Survivor and the gas phase protonation model. Experimental differentiation between the predicted mechanisms becomes possible by the use of deuterated matrix esters as MALDI matrices, which are stable under typical sample preparation conditions and generate deuteronated reagent ions, including the deuterated and deuteronated free matrix acid, only upon laser irradiation in the MALDI process. While the generation of deuteronated analyte ions proves the gas phase protonation model, the detection of protonated analytes by application of deuterated matrix compounds without acidic hydrogens proves the survival of analytes precharged from solution in accordance with the predictions from the Lucky Survivor model. The observed ratio of the two analyte ionization processes depends on the applied experimental parameters as well as the nature of analyte and matrix. Increasing laser fluences and lower matrix proton affinities favor gas phase protonation, whereas more quantitative analyte protonation in solution and intramolecular ion stabilization leads to more Lucky Survivors. The presented results allow for a deeper understanding of the fundamental processes causing analyte ionization in MALDI and may alleviate future efforts for increasing the analyte ion yield.

Section: The Lucky Survivor Modelmentioning

confidence: 99%

Section: Medium Basic Analytes A/[a + H] +mentioning

confidence: 99%

Compelling Evidence for Lucky Survivor and Gas Phase Protonation: The Unified MALDI Analyte Protonation Mechanism

Jaskolla

Karas

2011

166

“…To leave no doubt about the possible gas-phase aggregation between DNA and the polybasic compound within our experimental conditions, a second experiment was carried out. This experiment was previously described by Friess and Zenobi [30]. Two solutions were prepared separately, one containing DAB8 and DHAP, the other containing DNAi and DHAP, just as for two classical MALDI deposits.…”

Section: Control Experimentsmentioning

confidence: 99%

Noncovalent complexes between DNA and basic polypeptides or polyamines by MALDI-TOF

Terrier

Tortajada

Zin

et al. 2007

MALDI-MS was evaluated as a method for the study of noncovalent complexes involving DNA oligonucleotides and various polybasic compounds (basic polypeptides and polyamines). Complexes involving single-stranded DNA were successfully detected using DHAP matrix in the presence of an ammonium salt. Control experiments confirmed that the interactions involved basic sites of the polybasic compounds and that the complexes were not formed in the gas phase but were pre-existing in the matrix crystals. Moreover, the pre-existence in solution was probed by isothermal titration calorimetry at concentration and ionic strength similar to those used for mass spectrometry. Spectra showed no important difference between negative and positive ion modes. The influence of nature and size of DNA and polybasic compound on the relative intensities and stoichiometries of the complexes was investigated. Despite the fact that relative intensities can be affected by ionization yields and the gas-phase stabilities of the different species, numerous trends observed in the MALDI study were consistent with the expected in-solution behaviors. Experimental conditions related to sample preparation were investigated also. Complex abundance generally decreased when increasing the ammonium acetate concentration. It was dramatically decreased when using ATT instead of DHAP. Penta-L-arginine is an exception to these observations. Lastly, in the case of complexes involving DNA duplex, the ATT matrix was shown to favor the observation of specific DNA duplex but not that of its complex with polybasic compounds. Inversely, DHAP was appropriate for the conservation of DNA-polybasic compound interaction but not for the transfer of intact

“…There are mass spectrometric methods available for obtaining conformational information of molecules in the gas phase: blackbody infrared radiative dissociation where ions undergo unimolecular dissociation by exchanging their energy with the surroundings by absorption and emission of infrared photons [3], collision-induced dissociation where ions are dissociated as a result of interaction with a target neutral species [4], hydrogendeuterium exchange where hydrogen atoms of the protein are exchanged with the deuterium atoms from a solution [5], covalent or noncovalent tagging of biomolecules based on the surface accessibility of specific moieties [6,7], and ion mobility spectrometry where the measured cross section of the molecule is compared with a theoretically calculated cross section [8]. However, generally speaking, these methods yield only indirect information about the gas-phase conformation, which is often derived from either the overall cross section of the molecule, from a fragmentation pattern, or from a dissociation rate.…”

mentioning

confidence: 99%

Clear evidence of fluorescence resonance energy transfer in gas-phase ions

Dashtiev

Azov

Frankevich

et al. 2005

Self Cite

Fluorescence resonance energy transfer (FRET) is a distance-sensitive method that correlates changes in fluorescence intensity with conformational changes, for example, of biomolecules in the cellular environment. Applied to the gas phase in combination with Fourier transform ion cyclotron resonance mass spectrometry, it opens up possibilities to define structural/ conformational properties of molecular ions, in the absence of solvent, and without the need for purification of the sample. For successfully observing FRET in the gas phase it is important to find suitable fluorophores. In this study several fluorescent dyes were examined, and the correlation between solution-phase and gas-phase fluorescence data were studied. he study of biomolecular conformation in the gas phase has attracted great attention because it opens possibilities to compare gas-phase and solution-phase structures and to understand the effect of the solvent on a molecule. Because matrix-assisted laser desorption/ionization (MALDI) [1] and electrospray ionization (ESI) [2] typically generate unsolvated ions without any adducts, their structure also provides an ideal model system for theoretical calculations of conformations. A major question today is whether singly or multiply charged biomolecular ions produced by soft ionization methods retain their native (or at least a "native-like"), active conformation in the gas phase. This is often assumed but needs to be proven rigorously.Mass spectrometry has many attractive features for studying biomolecules in the gas phase, including the capability of the isolation of ions and elimination of unwanted species before detection and a positive identification of the molecule from the exact mass or from tandem mass spectrometry (MS/MS) data. There are mass spectrometric methods available for obtaining conformational information of molecules in the gas phase: blackbody infrared radiative dissociation where ions undergo unimolecular dissociation by exchanging their energy with the surroundings by absorption and emission of infrared photons [3], collision-induced dissociation where ions are dissociated as a result of interaction with a target neutral species [4], hydrogendeuterium exchange where hydrogen atoms of the protein are exchanged with the deuterium atoms from a solution [5], covalent or noncovalent tagging of biomolecules based on the surface accessibility of specific moieties [6,7], and ion mobility spectrometry where the measured cross section of the molecule is compared with a theoretically calculated cross section [8]. However, generally speaking, these methods yield only indirect information about the gas-phase conformation, which is often derived from either the overall cross section of the molecule, from a fragmentation pattern, or from a dissociation rate.Recently, the efficient trapping capabilities of mass spectrometry have been coupled with the high sensitivity of fluorescence detection to provide structural and spectroscopic information of molecules in the gas phase. The general idea...