Distinguishing Aspartic and Isoaspartic Acids in Peptides by Several Mass Spectrometric Fragmentation Methods

DeGraan-Weber, Nick; Zhang, Jun; Reilly, James P.

doi:10.1007/s13361-016-1487-9

Cited by 28 publications

(43 citation statements)

References 51 publications

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“…2b,d. To distinguish between the possible products of Asp and iso-Asp, we noted the difference in their fragmentation patterns of MS/MS spectra, expecting that iso-Asp residues show the enhanced b-or y-type ion on the N-terminal side 10 . As shown in Fig.…”

Section: Resultsmentioning

confidence: 99%

Effect of UVC Irradiation on the Oxidation of Histidine in Monoclonal Antibodies

Miyahara

Shintani

Hayashihara-Kakuhou

et al. 2020

Sci Rep

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We oxidized histidine residues in monoclonal antibody drugs of immunoglobulin gamma 1 (IgG1) using ultraviolet C irradiation (UVC: 200-280 nm), which is known to be potent for sterilization or disinfection. Among the reaction products, we identified asparagine and aspartic acid by mass spectrometry. In the photo-induced oxidation of histidine in angiotensin II, 18 O atoms from H 2 18 O in the solvent were incorporated only into aspartic acid but not into asparagine. This suggests that UVC irradiation generates singlet oxygen and induces [2 + 2] cycloaddition to form a dioxetane involving the imidazole c γ − C δ2 bond of histidine, followed by ring-opening in the manner of further photo-induced retro [2 + 2] cycloaddition. This yields an equilibrium mixture of two keto-imines, which can be the precursors to aspartic acid and asparagine. The photo-oxidation appears to occur preferentially for histidine residues with lower pK a values in IgG1. We thus conclude that the damage due to UVC photo-oxidation of histidine residues can be avoided in acidic conditions where the imidazole ring is protonated.

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Section: Resultsmentioning

confidence: 99%

Effect of UVC Irradiation on the Oxidation of Histidine in Monoclonal Antibodies

Miyahara

Shintani

Hayashihara-Kakuhou

et al. 2020

Sci Rep

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“…A challenge in primary structure elucidation is the biochemical instability inherent in proteins. Deamidation of Asn and the isomerization of Asp is one of the most common non-enzymatic PTMs [80][81][82]. isoAsp has been implicated in several high-profile pathologies, e.g., Alzheimer's disease and metastatic breast cancer, due to its aggregation or loss of functionality [67].…”

Section: Discussionmentioning

confidence: 99%

Aspartic Acid Isomerization Characterized by High Definition Mass Spectrometry Significantly Alters the Bioactivity of a Novel Toxin from Poecilotheria

Johnson

Rikli

2020

Toxins

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Research in toxinology has created a pharmacological paradox. With an estimated 220,000 venomous animals worldwide, the study of peptidyl toxins provides a vast number of effector molecules. However, due to the complexity of the protein-protein interactions, there are fewer than ten venom-derived molecules on the market. Structural characterization and identification of post-translational modifications are essential to develop biological lead structures into pharmaceuticals. Utilizing advancements in mass spectrometry, we have created a high definition approach that fuses conventional high-resolution MS-MS with ion mobility spectrometry (HDMSE) to elucidate these primary structure characteristics. We investigated venom from ten species of “tiger” spider (Genus: Poecilotheria) and discovered they contain isobaric conformers originating from non-enzymatic Asp isomerization. One conformer pair conserved in five of ten species examined, denominated PcaTX-1a and PcaTX-1b, was found to be a 36-residue peptide with a cysteine knot, an amidated C-terminus, and isoAsp33Asp substitution. Although the isomerization of Asp has been implicated in many pathologies, this is the first characterization of Asp isomerization in a toxin and demonstrates the isomerized product’s diminished physiological effects. This study establishes the value of a HDMSE approach to toxin screening and characterization.

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“…Interestingly, this fragmentation channel allows discriminating leucine from isoleucine residues based on the loss of an ethyl‐ or 2‐propyl radical, respectively. Similarly, aspartic and isoaspartic acids can be also distinguished . In contrast, H‐atom abstraction from the C γ ‐methylene group induces the homolytic cleavage of a C α −C β bond and the formation of an even‐electron side chain fragment, while the radical is retained at the C α ‐position (Figure E).…”

Section: Fripsmentioning

confidence: 99%

“…66 Hydrogen abstraction from the C α -position (Figure 2A) can be also distinguished. 96 In contrast, H-atom abstraction from the C γmethylene group induces the homolytic cleavage of a C α −C β bond and the formation of an even-electron side chain fragment, while the radical is retained at the C α -position ( Figure 2E). It has been also reported that acetyl radicals, formed by TEMPObased FRIPS reagent dissociations, homolytically cleave the S─S bond.…”

Section: Proposed Mechanismsmentioning

confidence: 99%

Free radical‐initiated peptide sequencing (FRIPS)‐based cross‐linkers for improved peptide and protein structure analysis

Iacobucci

Schäfer

Sinz

2018

Mass Spectrometry Reviews

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Free radical-initiated peptide sequencing (FRIPS) has recently been introduced as an analytical strategy to create peptide radical ions in a predictable and effective way by collisional activation of specifically modified peptides ions. FRIPS is based on the unimolecular dissociation of open-shell ions and yields fragments that resemble those obtained by electron capture dissociation (ECD) or electron transfer dissociation (ETD). In this review article, we describe the fundamentals of FRIPS and highlight its fruitful combination with chemical cross-linking/mass spectrometry (MS) as a highly promising option to derive complementary structural information of peptides and proteins. FRIPS does not only yield exhaustive sequence information of cross-linked peptides, but also defines the exact cross-linking sites of the connected peptides. The development of more advanced FRIPS cross-linkers that extend the FRIPS-based cross-linking/MS approach to the study of large protein assemblies and protein interaction networks can be eagerly anticipated.

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Distinguishing Aspartic and Isoaspartic Acids in Peptides by Several Mass Spectrometric Fragmentation Methods

Cited by 28 publications

References 51 publications

Effect of UVC Irradiation on the Oxidation of Histidine in Monoclonal Antibodies

Effect of UVC Irradiation on the Oxidation of Histidine in Monoclonal Antibodies

Aspartic Acid Isomerization Characterized by High Definition Mass Spectrometry Significantly Alters the Bioactivity of a Novel Toxin from Poecilotheria

Free radical‐initiated peptide sequencing (FRIPS)‐based cross‐linkers for improved peptide and protein structure analysis

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