Mass spectral analysis of acetylated peptides: Implications in proteomics

Abstract

Sequence determination of peptides using mass spectrometry plays a crucial role in the bottom-up approaches for the identification of proteins. It is crucially important to minimise false detection and validate sequence of the peptides in order to correctly identify a protein. Chemical modification of peptides followed by mass spectrometry is an option for improving the spectral quality. In silico-derived tryptic peptides with different N-terminal amino acids were designed from human proteins and synthesized. The effect of acetylation on the fragmentation of peptides was studied. N-terminal acetylation of the tryptic peptides was shown to form b1-ions, improve the abundance and occurrence of b-ions. In some cases, the intensity and occurrence of some y-ions also varied. Thus, it is demonstrated that acetylation plays an important role in improving the de novo sequencing efficiency of the peptides. The acetylation method was extended to tryptic peptides generated from the proteome of an Antarctic bacterium Pseudomonas syringae Lz4W using the proteomics work flow and mass spectra of the peptides were analysed. Comparison of the MS/MS spectra of the acetylated and unacetylated peptides revealed that acetylation helped in improving the spectral quality and validated the peptide sequences. Using this method, 673 proteins of the 1070 proteins identified were validated.

Positive Effect of Acetylation on Proteomic Analysis Based on Liquid Chromatography with Atmospheric Pressure Chemical Ionization and Photoionization Mass Spectrometry

“…Protonated and sodiated precursors provided abundant b, y, and a ions. In agreement with previous reports [60], MS 2 spectra of acetylated peptides showed higher numbers and significantly more abundant b ions compared to non-acetylated peptides. Protonated peptides in low-energy CID produce b and y ion series.…”

Mass spectrometry insights into interactions of selenoprotein P with auranofin and cisplatin

“…Acetylation of tryptic peptides by a reaction with acetic anhydride is a means of improving the quality of mass spectra by increasing the occurrence and abundance of fragment ions. 27 Although no intentional acetylation was carried out, the presence of these modifications can be attributed to a prolonged contact of the sample with high concentrations of acetate during the SelP purification step followed by acidification with ( ca. 1% formic acid just after tryptic digestion) which could catalyze the observed reaction.…”

Rapid and Easy Enrichment Strategy for Naturally Acetylated N Termini Based on LysN Digestion and Amine-Reactive Resin Capture

“…We found that the identified 203 protein N termini with the initiator methionine were corresponding to 248 peptide identification, among which 215 peptides (86.7%) were not found to contain oxidized methionine, while two forms of some peptides both with and without oxidized methionine were identified at the same time. It was reported that N-terminal acetylation of the tryptic peptides could improve the abundance and occurrence of b-ions including b1 ions . Therefore, the % of b1 ions detected from N-terminal acetylated residues was also counted to provide additional information for the N-terminal acetylated peptides.…”

“…It was reported that N-terminal acetylation of the tryptic peptides could improve the abundance and occurrence of b-ions including b1 ions. 32 Therefore, the % of b1 ions detected from N-terminal acetylated residues was also counted to provide additional information for the N-terminal acetylated peptides. Their molecular functions and subcellular locations were analyzed using IPA (Ingenuity Pathway Analysis), and we found that these identified proteins were involved in various molecular functions such as cell cycle, gene expression, cell death and survival, molecular transport and cell signaling, and so on.…”