Electron capture dissociation mass spectrometry of tyrosine nitrated peptides

Hydrogen-deficient peptide radical cations exhibit fascinating gas phase chemistry, which is governed by radical driven dissociation and, in many cases, by a combination of radical and charge driven fragmentation. Here we examine electron capture dissociation (ECD) of doubly, • precursor ions resulted in efficient electron capture by the hydrogen-deficient species. However, the intensities of c-and z-type product ions were reduced, compared with those observed for the even electron species, indicating suppression of N-C α backbone bond cleavages. We postulate that radical recombination occurs after the initial electron capture event leading to a stable even electron intermediate, which does not trigger N-C α bond dissociations. Although the intensities of c-and z-type product ions were reduced, the number of backbone bond cleavages remained largely unaffected between the ECD spectra of the even electron and hydrogen-deficient species. We hypothesize that a small ion population exist as a biradical, which can trigger N-C α bond cleavages. Alternatively, radical recombination and N-C α bond cleavages can be in competition, with radical recombination being the dominant pathway and N-C α cleavages occurring to a lesser degree. Formation of b-and y-type ions observed for two of the hydrogendeficient peptides examined is also discussed.

Section: Discussionsupporting

confidence: 71%

Section: Dissociation Of Doubly [M + H] 2+• and Triply [M + 2h] 3+mentioning

confidence: 76%

Electron Capture Dissociation of Hydrogen-Deficient Peptide Radical Cations

Kalli

Hess

2012

“…Electron capture dissociation (ECD) [11,12] mass spectrometry, and its sister technique electron transfer dissociation (ETD) [13], have emerged as useful methods for the analysis of protein PTMs, including phosphorylation [14,15], and glycosylation [16,17]. Nevertheless, ECD is not universally appropriate for the characterization of PTMs (e.g., tyrosine nitration [18]). In peptide/protein ECD, low energy electrons are captured by multiply-charged cations with subsequent cleavage of disulfide S-S and peptide backbone N-C α bonds i.e., cleavage is radically driven.…”

mentioning

confidence: 99%

The Radical Ion Chemistry of S-Nitrosylated Peptides

Jones

Winn

Cooper

2012

Self Cite

The radical ion chemistry of a suite of S-nitrosopeptides has been investigated. Doubly and triply-protonated ions of peptides NYCGLPGEYWLGNDK, NYCGLPGEYWLGNDR, NYCGLP-GERWLGNDR, NACGAPGEKWAGNDK, NYCGLPGEKYLGNDK, NYGLPGCEKWYGNDK and NYGLPGEKWYGCNDK were subjected to electron capture dissociation (ECD), and collisioninduced dissociation (CID). The peptide sequences were selected such that the effect of the site of S-nitrosylation, the nature and position of the basic amino acid residues, and the nature of the other amino acid side chains, could be interrogated. The ECD mass spectra were dominated by a peak corresponding to loss of• NO from the charge-reduced precursor, which can be explained by a modified Utah-Washington mechanism. Some backbone fragmentation in which the nitrosyl modification was preserved was also observed in the ECD of some peptides. Molecular dynamics simulations of peptide ion structure suggest that the ECD behavior was dependent on the surface accessibility of the protonated residue. CID of the S-nitrosylated peptides resulted in homolysis of the S-N bond to form a long-lived radical with loss of • NO. The radical peptide ions were isolated and subjected to ECD and CID. ECD of the radical peptide ions provided an interesting comparison to ECD of the unmodified peptides. The dominant process was electron capture without further dissociation (ECnoD). CID of the radical peptide ions resulted in cysteine, leucine, and asparagine side chain losses, and radical-induced backbone fragmentation at tryptophan, tyrosine, and asparagine residues, in addition to charge-directed backbone fragmentation.

“…All of the following peptides were provided by Protea Biosciences Inc. (Morgantown, WV, USA): the phosphorylated and non-phosphorylated forms of angiotensin II, cholecystokin (10)(11)(12)(13)(14)(15)(16)(17)(18)(19)(20) and calcitonin (15)(16)(17)(18)(19)(20)(21)(22)(23)(24)(25)(26)(27)(28)(29); the sulfated and non-sulfated forms of cholecystokinin (26)(27)(28)(29)(30)(31)(32)(33), leuenkephalin, and hirudin. Methanol (HPLC grade), ammonium hydroxide, and glacial acetic acid were purchased from Sigma-Aldrich (St. Louis, MO, USA).…”

Section: Preparation Of Peptidesmentioning

confidence: 99%

“…To date, both methods have been successfully employed to determine phosphorylation [13,14], glycosylation [15][16][17], and sulfonation [9,10] modifications, among others. On the other hand, it has been recently shown that certain modifications, such as phosphorylation [18] and nitration [19][20][21], have hindered ECD and ETD sequence coverage and modification identification. Liu and Håkansson showed that highly acidic peptides with no basic amino acids produced complete loss of the sulfonated modification, thereby rendering site determination impossible by ECD [22].…”

mentioning

confidence: 99%

Metastable Atom-Activated Dissociation Mass Spectrometry of Phosphorylated and Sulfonated Peptides in Negative Ion Mode

Cook

Jackson

2011

The dissociation behavior of phosphorylated and sulfonated peptide anions was explored using metastable atom-activated dissociation mass spectrometry (MAD-MS) and collision-induced dissociation (CID). A beam of high kinetic energy helium (He) metastable atoms was exposed to isolated phosphorylated and sulfonated peptides in the 3-and 2-charge states. Unlike CID, where phosphate losses are dominant, the major dissociation channels observed using MAD were C α -C peptide backbone cleavages and neutral losses of CO 2 , H 2 O, and [CO 2 +H 2 O] from the charge reduced (oxidized) product ion, consistent with an electron detachment dissociation (EDD) mechanism such as Penning ionization. Regardless of charge state or modification, MAD provides ample backbone cleavages with little modification loss, which allows for unambiguous PTM site determination. The relative abundance of certain fragment ions in MAD is also demonstrated to be somewhat sensitive to the number and location of deprotonation sites, with backbone cleavage somewhat favored adjacent to deprotonated sites like aspartic acid residues. MAD provides a complementary dissociation technique to CID, ECD, ETD, and EDD for peptide sequencing and modification identification. MAD offers the unique ability to analyze highly acidic peptides that contain few to no basic amino acids in either negative or positive ion mode.