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Iodoacetamide‐alkylated methionine can mimic neutral loss of phosphoric acid from phosphopeptides as exemplified by nano‐electrospray ionization quadrupole time‐of‐flight parent ion scanning
Author(s) -
Krüger Ralf,
Hung ChienWen,
EdelsonAverbukh Marina,
Lehmann Wolf D.
Publication year - 2005
Publication title -
rapid communications in mass spectrometry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.528
H-Index - 136
eISSN - 1097-0231
pISSN - 0951-4198
DOI - 10.1002/rcm.1976
Subject(s) - chemistry , phosphoserine , electrospray ionization , collision induced dissociation , tandem mass spectrometry , mass spectrometry , serine , chromatography , biochemistry , phosphorylation
Formation of S ‐carbamidomethylmethionine (camMet) occurs as a side reaction during cysteine alkylation with iodoacetamide (IAA). In collision‐induced dissociation, peptides with camMet show an abundant neutral loss of 2‐(methylthio)acetamide (C 3 H 7 NOS = 105.025 Da) at moderate collision offset values which are similar to those optimal for loss of phosphoric acid (H 3 PO 4 = 97.977 Da). Neutral loss analysis is used for spotting of phosphopeptides which contain phosphoserine (pSer) or phosphothreonine (pThr) residues. In the case where precursor ions cannot be accurately assigned in the survey spectrum (e.g. due to low ion abundance or signal overlap), the mass accuracy of a neutral loss tandem mass spectrometry (MS/MS) analysis depends on the precursor ion isolation window. For the charge states 2+, 3+ or 4+, a typical 3.5 Da precursor isolation window results in neutral loss windows of 7, 10.5 or 14 Da, respectively. Consequently, neutral loss of 105 Da from alkylated methionine residues can mimic the phosphoserine/phosphothreonine‐specific neutral loss of 98 Da. In the evaluation of quadrupole time‐of‐flight (QTOF) parent ion scan data for neutral loss of H 3 PO 4 , this interference was frequently observed. It is illustrated in this study using the analysis of ovalbumin phosphorylation as an example. The +80 Da molecular weight shift connected with phosphorylation at serine or threonine may also be mimicked by carbamidomethylation of methionine through a combination with sodium adduction (+57 Da +22 Da = +79 Da). For highly sensitive neutral loss analysis of serine and threonine phosphorylation, careful data inspection is recommended if reduction and alkylation by IAA is employed. Copyright © 2005 John Wiley & Sons, Ltd.