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A versatile peptide p I calculator for phosphorylated and N‐terminal acetylated peptides experimentally tested using peptide isoelectric focusing
Author(s) -
Gauci Sharon,
van Breukelen Bas,
Lemeer Simone M.,
Krijgsveld Jeroen,
Heck Albert J. R.
Publication year - 2008
Publication title -
proteomics
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.26
H-Index - 167
eISSN - 1615-9861
pISSN - 1615-9853
DOI - 10.1002/pmic.200800295
Subject(s) - peptide , calculator , acetylation , isoelectric focusing , chemistry , chromatography , phosphorylation , biochemistry , computer science , gene , enzyme , operating system
We experimentally demonstrate the use of an in‐house developed p I calculator which takes into account peptide PTM such as phosphorylation and N‐terminal acetylation. The p I calculator was utilized for a large set of peptides derived from a complex zebrafish lysate fractionated using peptide IEF, whereby a good correlation between the calculated (theoretical) p I and the experimental p I could be established. This p I calculator permits the implementation of optimal p K values depending on the experimental conditions and a reliable calculation of peptide p I which can be utilized as a filtering technique in validating peptide identifications. Our data reveal that the shift due to a phosphorylation or N‐terminal acetylation is highly dependent on the presence of acidic or basic residues in the peptide. Furthermore, using this p I calculator, we revealed previously unknown position‐specific p K s of asparagine and carbamidomethylated cysteine depending on their location in the peptide. Collectively, this peptide p I calculator is a welcome addition to the versatility and robustness of IEF for the separation and confident identification of (post‐translationally modified) peptides.