Publication details

A comparative study of synthetic winged peptides for absolute protein quantification

Authors

BENEŠOVÁ Eliška VIDOVÁ Veronika SPÁČIL Zdeněk

Year of publication 2021
Type Article in Periodical
Magazine / Source Nature Scientific Reports
MU Faculty or unit

Faculty of Science

Citation
Web https://www.nature.com/articles/s41598-021-90087-9
Doi http://dx.doi.org/10.1038/s41598-021-90087-9
Keywords INTERNAL STANDARD SELECTIONT; ANDEM MASS-SPECTROMETRY; SIGNATURE PEPTIDE; TRYPTIC DIGESTION; PLASMA; IDENTIFICATION; QUANTITATION; OSTEOPONTIN; ACCURACY; IMPACT
Attached files
Description A proper internal standard choice is critical for accurate, precise, and reproducible mass spectrometry-based proteomics assays. Synthetic isotopically labeled (SIL) proteins are currently considered the gold standard. However, they are costly and challenging to obtain. An alternative approach uses SIL peptides or SIL "winged" peptides extended at C- or/and N-terminus with an amino acid sequence or a tag cleaved during enzymatic proteolysis. However, a consensus on the design of a winged peptide for absolute quantification is missing. In this study, we used human serum albumin as a model system to compare the quantitative performance of reference SIL protein with four different designs of SIL winged peptides: (i) commercially available SIL peptides with a proprietary trypsin cleavable tag at C-terminus, (ii) SIL peptides extended with five amino acid residues at C-terminus, (iii) SIL peptides extended with three and (iv) with five amino acid residues at both C- and N-termini. Our results demonstrate properties of various SIL extended peptides designs, e.g., water solubility and efficiency of trypsin enzymatic cleavage with primary influence on quantitative performance. SIL winged peptides extended with three amino acids at both C- and N-termini demonstrated optimal quantitative performance, equivalent to the SIL protein.
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