Enhanced Electron Transfer Dissociation of Peptides Modified at C-terminus with Fixed Charges

Ko, Byoung Joon; Brodbelt, Jennifer S.

doi:10.1007/s13361-012-0458-z

Cited by 25 publications

(37 citation statements)

References 53 publications

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“…Notice that the peak intensities for the precursor and charge-reduced precursors are similar to those of the c -and • z-type fragment ions, indicating that nearly all of the peptide ions undergo fragmentation. The fragmentation efficiency is calculated by summing all c - and • z-type product ion intensities and then dividing by the sum of all fragment and precursor ion intensities in the ETD spectrum [42]. The ETD efficiencies for the DAFLGSFLYEYSR peptide in its unmodified, C4Quat-modified, and C5-methylTert-modified forms are 2%, 80%, and 90%, respectively.…”

Section: Resultsmentioning

confidence: 99%

“…A few researchers have derivatized carboxyl groups on peptides to increase charge states during LC-MS [38–42]. All have employed essentially the same derivatization strategy, first developed by Lu and co-workers, namely using the carbodiimide EDC along with HOAt in DMF solvent (EDC: 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide, HOAt: 1-hydroxy-7-azabenzotriazole, and DMF: dimethylformamide) [38].…”

Section: Introductionmentioning

confidence: 99%

“…However, fragmentation was still not particularly efficient for these dication precursors for the reasons described above. Ko and Brodbelt derivatized model peptides, as well as those from a tryptic digest of the small protein cytochrome c, by attaching basic or fixed-charge groups to the peptide carboxyls [42]. They thoroughly analyzed the charge states and ETD fragmentation efficiencies, both of which increased substantially for the modified peptides.…”

Section: Introductionmentioning

confidence: 99%

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Chemical Derivatization of Peptide Carboxyl Groups for Highly Efficient Electron Transfer Dissociation

Frey

Ladror

Sondalle

et al. 2013

J. Am. Soc. Mass Spectrom.

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The carboxyl groups of tryptic peptides were derivatized with a tertiary or quaternary amine labeling reagent to generate more highly charged peptide ions that fragment efficiently by electron transfer dissociation (ETD). All peptide carboxyl groups—aspartic and glutamic acid side-chains as well as C-termini—were derivatized with an average reaction efficiency of 99%. This nearly complete labeling avoids making complex peptide mixtures even more complex due to partially-labeled products, and it allows the use of static modifications during database searching. Alkyl tertiary amines were found to be the optimal labeling reagent among the four types tested. Charge states are substantially higher for derivatized peptides: a modified tryptic digest of bovine serum albumin (BSA) generates ∼90% of its precursor ions with z > 2, compared to less than 40% for the unmodified sample. The increased charge density of modified peptide ions yields highly efficient ETD fragmentation, leading to many additional peptide identifications and higher sequence coverage (e.g. 70% for modified versus only 43% for unmodified BSA). The utility of this labeling strategy was demonstrated on a tryptic digest of ribosomal proteins isolated from yeast cells. Peptide derivatization of this sample produced an increase in the number of identified proteins, a >50% increase in the sequence coverage of these proteins, and a doubling of the number of peptide spectral matches. This carboxyl derivatization strategy greatly improves proteome coverage obtained from ETD-MS/MS of tryptic digests, and we anticipate that it will also enhance identification and localization of post-translational modifications.

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Section: Resultsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 1 more Smart Citation

Chemical Derivatization of Peptide Carboxyl Groups for Highly Efficient Electron Transfer Dissociation

Frey

Ladror

Sondalle

et al. 2013

J. Am. Soc. Mass Spectrom.

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“…Another way to enhance ionization efficiency of peptides is their derivatization which results in a formation of fixed charged peptide derivative and their increased intensity in ESI1112. Several derivatization strategies leading to the formation of charge-tagged peptides are reported including quaternary ammonium (QAS)13 or phosphonium salt (QPS)14.…”

mentioning

confidence: 99%

Peptides Labeled with Pyridinium Salts for Sensitive Detection and Sequencing by Electrospray Tandem Mass Spectrometry

Waliczek

Kijewska

Setner

et al. 2016

Sci Rep

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Mass spectrometric analysis of trace amounts of peptides may be problematic due to the insufficient ionization efficiency resulting in limited sensitivity. One of the possible ways to overcome this problem is the application of ionization enhancers. Herein we developed new ionization markers based on 2,4,6-triphenylpyridinium and 2,4,6-trimethylpyridinium salts. Using of inexpensive and commercially available pyrylium salt allows selective derivatization of primary amino groups, especially those sterically unhindered, such as ε-amino group of lysine. The 2,4,6-triphenylpyridinium modified peptides generate in MS/MS experiments an abundant protonated 2,4,6-triphenylpyridinium ion. This fragment is a promising reporter ion for the multiple reactions monitoring (MRM) analysis. In addition, the fixed positive charge of the pyridinium group enhances the ionization efficiency. Other advantages of the proposed ionization enhancers are the simplicity of derivatization of peptides and the possibility of convenient incorporation of isotopic labels into derivatized peptides.

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“…Amidation has been used for derivatization of aspartate, glutamate, and the C-terminus. 16 Unlike the modification of lysine and the N-terminus, distinguishing among these target side chains was not possible. Also, because there were three sites for reaction, an efficient modification approach was necessary.…”

Section: Resultsmentioning

confidence: 99%

Solution-phase and solid-phase sequential, selective modification of side chains in KDYWEC and KDYWE as models for usage in single-molecule protein sequencing

et al. 2017

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Single-molecule protein sequencing is regarded as a promising new method in the field of proteomics. It potentially offers orders of magnitude improvements in sensitivitiy and throughput for protein detection when compared to mass spectrometry. However, the development of such a technology faces significant barriers, especially in the need to chemically derivatize specific amino-acid types with unique labels. For example, fluorescent dyes would be suitable for single-molecule microscopy or nanopore-based sequencing. These emerging single-molecule protein-sequencing technologies suggests a need to develop an amino acid side chain-selective modification scheme that could target several side chains of interest. Current work for modifying residues focuses mainly on one or two side chains. The need to label many side chains, as recent computational modeling suggests, is required for high protein, sequencing coverage of the human proteome. Herein, we report our stragety for modifying two model peptides KYDWEC and KDYWE containing the most reactive residues, using highly opitmized mass labels in a sequential and selective fashion both using solution-phase and solid-phase chemistries, respectively. This will serve as a step towards a modification scheme appropriate for single-molecule studies.

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Enhanced Electron Transfer Dissociation of Peptides Modified at C-terminus with Fixed Charges

Cited by 25 publications

References 53 publications

Chemical Derivatization of Peptide Carboxyl Groups for Highly Efficient Electron Transfer Dissociation

Chemical Derivatization of Peptide Carboxyl Groups for Highly Efficient Electron Transfer Dissociation

Peptides Labeled with Pyridinium Salts for Sensitive Detection and Sequencing by Electrospray Tandem Mass Spectrometry

Solution-phase and solid-phase sequential, selective modification of side chains in KDYWEC and KDYWE as models for usage in single-molecule protein sequencing

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