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Characterization of γ-carboxylated tryptic peptides by collision-induced dissociation and electron transfer dissociation mass spectrometry.
Octapharma AB.
Octapharma AB.
2011 (Engelska)Ingår i: European journal of mass spectrometry, ISSN 1469-0667, E-ISSN 1751-6838, Vol. 17, nr 5, s. 497-506Artikel i tidskrift (Refereegranskat) Published
Abstract [en]

Vitamin K-dependent carboxylation of glutamic acid (Glu) residues into γ-carboxyglutamic acid (Gla) is a post-translational modification essential for normal protein activity of, for example, proteins involved in the blood coagulation system. These proteins may contain as many as 12 sites for γ-carboxylation within a protein sequence of 45 amino acid residues. In the biopharmaceutical industry, powerful analytical techniques are required for identification and localization of modified sites. We here present comparatively easy and rapid methods for studies of Gla-containing proteins using recent technology. The performances of two mass spectrometric fragmentation techniques, collision-induced dissociation (CID) and electron transfer dissociation (ETD), were evaluated with respect to γ-carboxylated peptides, applying on-line LC-ion trap MS. ETD MS has so far not been reported for Gla-containing peptides and the applicability of CID for heavily γ-carboxylated proteins has not been evaluated. The anticoagulant protein, protein C, containing nine Gla-sites, was chosen as a model protein. After tryptic digestion, three peptides containing Gla-residues were detected by MS; a 1.2 kDa fragment containing two Gla-residues, a 4.5 kDa peptide containing seven residues and also the 5.6 kDa tryptic peptides containing all nine Gla-residues. Regarding the shortest peptide, both CID and ETD provided extensive peptide sequencing. For the larger peptides, fragmentation by CID resulted in loss of the 44 Da CO(2)-group, while little additional fragmentation of the peptide chain was observed. In contrast, ETD resulted in comprehensive fragmentation of the peptide backbone. The study demonstrates that the combination of both techniques would be beneficial and complementary for investigation of γ-carboxylated proteins and peptides.

Ort, förlag, år, upplaga, sidor
2011. Vol. 17, nr 5, s. 497-506
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Naturvetenskap
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URN: urn:nbn:se:uu:diva-172150DOI: 10.1255/ejms.1149PubMedID: 22173536OAI: oai:DiVA.org:uu-172150DiVA, id: diva2:513437
Tillgänglig från: 2012-04-02 Skapad: 2012-04-02 Senast uppdaterad: 2017-12-07

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