Comparison of laser‐induced dissociation and high‐energy collision‐induced dissociation using matrix‐assisted laser desorption/ionization tandem time‐of‐flight (MALDI‐TOF/TOF) for peptide and protein identification

Macht, Marcus; Asperger, Arndt; Deininger, Sören‐Oliver

doi:10.1002/rcm.1589

Cited by 36 publications

(28 citation statements)

References 43 publications

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“…These gentle conditions for desorption generated precursor ions possessing insufficient internal energy to support metastable fragmentation. Accordingly nearly pure high energy CID spectra without PSD fragments can be recorded under such conditions (38,39). Unfortunately the yield of the "cold" precursor ions is very low compared with the yield of "hotter" precursors ions obtained at higher laser intensities.…”

Section: Resultsmentioning

confidence: 99%

Collisional Activation by MALDI Tandem Time-of-flight Mass Spectrometry Induces Intramolecular Migration of Amide Hydrogens in Protonated Peptides

Jørgensen

Bache

Roepstorff

et al. 2005

Molecular & Cellular Proteomics

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Mass spectrometry-based proteomics is currently one of the key technologies for the identification of protein interaction networks in the cell (1-3). This technology has provided a wealth of information unraveling the complexity of these biological processes at the protein level. A detailed understanding of the function of these protein networks at the molecular level requires, however, characterization of the dynamics and structural properties of the interacting proteins. Structural analyses by NMR spectroscopy or x-ray crystallography require relatively large amounts of pure proteins (typically milligram quantities), and many proteins are furthermore not inherently amenable to analysis by these traditional techniques (e.g. modular proteins or proteins with heterogeneous glycosylation patterns are often reluctant to yield well diffracting crystals).An alternative approach for the characterization of protein complexes without these limitations is provided by carefully monitoring by mass spectrometry changes in the rates of amide hydrogen ( 1 H/ 2 H) exchange upon complex formation (4, 5). In a typical experimental set-up for investigation of conformational properties of a protein complex, the unligated proteins as well as the preformed protein complex are incubated separately in deuterated buffer under physiological conditions. Global deuterium incorporation kinetics are subsequently established by monitoring deuterium contents in these protein samples withdrawn at appropriate intervals with their amide hydrogen isotopic exchange quenched by acidification and cooling. Information about local deuterium incorporation is subsequently collected after pepsin digestion followed by reversed-phase LC-MS. The chromatography is carried out with cold protiated solvents allowing effective back-exchange at all exchangeable sites with protium ( 1 H) with the sole exception of the amide groups forming the polypeptide chain. The actual number of individual amide hydrogens that can be resolved by this method, however, is limited by the number and sizes of peptides generated by pepsin. Overlapping sequence coverage is often provided by the broad specificity of pepsin, and although this promiscuity in substrate recognition increases the resolution in the local assignment of deuterium incorporation, single residue information can generally be obtained for only a few positions (6). For a first hand view, gas-phase fragmentation in the mass spectrometer may appear to be the logical choice for an auxiliary method providing the desired site-specific informaFrom the ‡Department

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Section: Resultsmentioning

confidence: 99%

Collisional Activation by MALDI Tandem Time-of-flight Mass Spectrometry Induces Intramolecular Migration of Amide Hydrogens in Protonated Peptides

Jørgensen

Bache

Roepstorff

et al. 2005

Molecular & Cellular Proteomics

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“…PSD spectra are composed of post-source decay fragment ions resulting from metastable ion and CID processes [35]. We previously showed that it is important to minimize the contributions of PSD ions to the photofragment ion spectrum.…”

Section: Methodsmentioning

confidence: 99%

Comparative studies of 193-nm photodissociation and TOF-TOFMS analysis of bradykinin analogues: The effects of charge site(s) and fragmentation timescales

Morgan

Russell

2006

J. Am. Soc. Mass Spectrom.

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The dissociation reactions of [M ϩ H] ϩ , [M ϩ Na] ϩ , and [M ϩ Cu] ϩ ions of bradykinin (amino acid sequence RPPGFSPFR) and three bradykinin analogues (RPPGF, RPPGFSPF, PPGF-SPFR) are examined by using 193-nm photodissociation and post-source decay (PSD) TOF-TOF-MS techniques. The photodissociation apparatus is equipped with a biased activation cell, which allows us to detect fragment ions that are formed by dissociation of short-lived (Ͻ1 s) photo-excited ions. In our previously reported photodissociation studies, the fragment ions were formed from ions dissociating with lifetimes that exceeded 10 s; thus these earlier photofragment ion spectra and post-source decay (PSD) spectra [composite of both metastable ion (MI) and collision-induced dissociation (CID)] were quite similar. On the other hand, short-lived photo-excited ions dissociate by simple bond cleavage reactions and other high-energy dissociation channels. We also show that product ion types and abundances vary with the location of the charge on the peptide ion. For example, H ϩ and Na ϩ cations can bind to multiple polar functional groups (basic amino acid side chains) of the peptide, whereas Cu ϩ ions preferentially bind to the guanidino group of the arginine side-chain and the N-terminal amine group. Furthermore, when Cu ϩ is the charge carrier, the abundances of non-sequence informative ions, especially loss of small neutral molecules (H 2 O and NH 3 ) is decreased for both photofragment ion and PSD spectra relative to that observed for [ [3] have revolutionized the analysis of biomolecules by mass spectrometry (MS) techniques. Identification of proteins by using "bottom-up" MS techniques, i.e., enzymatic digestion of proteins or protein mixtures followed by peptide mass fingerprinting and database searching [4,5], is now routine, and de novo sequencing [6] and determination of post-translational modifications using tandem mass spectrometry techniques is now commonly practiced in many laboratories. The major factor that complicates peptide structure determination is that the types of fragment ions observed in a tandem mass spectrum are related to the intrinsic properties of the gas-phase peptide ion [7], the activation method used, and the timeframe for probing unimolecular dissociation products [8,9]. For example, utilizing only the internal energy acquired during desorption/ionization and a relatively long fragmentation timescale [ϳ10 s, as in post-source decay (PSD) experiments] favors product ions formed via the lowest energy channels [10], i.e., b i -and y i -type fragments [11,12]. As we showed previously, the observed fragment ions and their relative abundances change significantly if the timescale of the experiment is altered [13].Collisional activation of gas-phase peptide ions with a neutral gas (eV to several keV collision energies) can be employed to increase the internal energy of the ions and, thereby, enhance the abundances of sequence informative fragment ions; however, the resulting spectra also contain large numbers of non-se...

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“…Large peptides and proteins prepared in hot matrices such as α-cyano-4-hydroxycinnaminic acid (HCCA) suffer extensive metastable fragmentation in reflector mode MALDI spectra, which complicates further analysis. 48,61 This effect was not significant in the 5 – 6 kDa range, but prevented the analysis of 8 – 9 kDa hinge peptides. The B isoform digests were therefore crystallized in super DHB (sDHB) matrix instead of HCCA to reduce the effect of metastable fragmentation on the molecular ion.…”

Section: Resultsmentioning

confidence: 96%

Rapid, automated characterization of disulfide bond scrambling and IgG2 isoform determination

Resemann

Liu-Shin

Tremintin

et al. 2018

mAbs

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Human antibodies of the IgG2 subclass exhibit complex inter-chain disulfide bonding patterns that result in three structures, namely A, A/B, and B. In therapeutic applications, the distribution of disulfide isoforms is a critical product quality attribute because each configuration affects higher order structure, stability, isoelectric point, and antigen binding. The current standard for quantification of IgG2 disulfide isoform distribution is based on chromatographic or electrophoretic techniques that require additional characterization using mass spectrometry (MS)-based methods to confirm disulfide linkages. Detailed characterization of the IgG2 disulfide linkages often involve MS/MS approaches that include electrospray ionization or electron-transfer dissociation, and method optimization is often cumbersome due to the large size and heterogeneity of the disulfide-bonded peptides. As reported here, we developed a rapid LC-MALDI-TOF/TOF workflow that can both identify the IgG2 disulfide linkages and provide a semi-quantitative assessment of the distribution of the disulfide isoforms. We established signature disulfide-bonded IgG2 hinge peptides that correspond to the A, A/B, and B disulfide isoforms and can be applied to the fast classification of IgG2 isoforms in heterogeneous mixtures.

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Comparison of laser‐induced dissociation and high‐energy collision‐induced dissociation using matrix‐assisted laser desorption/ionization tandem time‐of‐flight (MALDI‐TOF/TOF) for peptide and protein identification

Cited by 36 publications

References 43 publications

Collisional Activation by MALDI Tandem Time-of-flight Mass Spectrometry Induces Intramolecular Migration of Amide Hydrogens in Protonated Peptides

Collisional Activation by MALDI Tandem Time-of-flight Mass Spectrometry Induces Intramolecular Migration of Amide Hydrogens in Protonated Peptides

Comparative studies of 193-nm photodissociation and TOF-TOFMS analysis of bradykinin analogues: The effects of charge site(s) and fragmentation timescales

Rapid, automated characterization of disulfide bond scrambling and IgG2 isoform determination

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