De novo sequencing of tryptic peptides sulfonated by 4‐sulfophenyl isothiocyanate for unambiguous protein identification using post‐source decay matrix‐assisted laser desorption/ionization mass spectrometry
peer-reviewed · Rapid Communications in Mass Spectrometry · 2004
| Date | 2004-01-30 |
| Type | peer-reviewed |
| Venue | Rapid Communications in Mass Spectrometry |
| Publisher | Wiley |
| Contribution | algorithm |
| DOI | 10.1002/rcm.1280 |
| Citations (OpenAlex) | 57 |
| Venue 2-year citedness | 1.80 |
Abstract
A simple method of solid-phase derivatization and sequencing of tryptic peptides has been developed for rapid and unambiguous identification of spots on two-dimensional gels using post-source decay (PSD) matrix-assisted laser desorption/ionization (MALDI) mass spectrometry. The proteolytic digests of proteins are chemically modified by 4-sulfophenyl isothiocyanate. The derivatization reaction introduces a negative sulfonic acid group at the N-terminus of a peptide, which can increase the efficiency of PSD fragmentation and enable the selective detection of only a single series of fragment ions (y-ions). This chemically assisted method avoids the limitation of high background normally observed in MALDI-PSD spectra, and makes the spectra easier to interpret and facilitates de novo sequencing of internal fragment. The modification reaction is conducted in C(18) microZipTips to decrease the background and to enhance the signal/noise. Derivatization procedures were optimized for MALDI-PSD to increase the structural information and to obtain a complete peptide sequence even in critical cases. The MALDI-PSD mass spectra of two model peptides and their sulfonated derivatives are compared. For some proteins unambiguous identification could be achieved by MALDI-PSD sequencing of derivatized peptides obtained from in-gel digests of phosphorylase B and proteins of hepatic satellite cells (HSC).
Methods and tools
- ZipTip SPITC derivatization for MALDI-PSD de novo sequencing: Sulfonates tryptic peptides with 4-sulfophenyl isothiocyanate on C18 microZipTips so MALDI post-source decay spectra show only y ions, which are then read de novo to identify gel spots.
Cites (2)
- Derivatization procedures to facilitate de novo sequencing of lysine-terminated tryptic peptides using postsource decay matrix-assisted laser desorption/ionization mass spectrometry (2000) crossref
- A method for high-sensitivity peptide sequencing using postsource decay matrix-assisted laser desorption ionization mass spectrometry (1999) crossref
Cited by (3)
- The effect of starvation stress on Lactobacillus brevis L62 protein profile determined by de novo sequencing in positive and negative mass spectrometry ion mode (2013) crossref
- Rapid and Reliable Peptide de Novo Sequencing Facilitated by Microfluidic Chip-Based Edman Degradation (2008) crossref
- Detection of hypoxia-related proteins in medaka (Oryzias latipes) brain tissue by difference gel electrophoresis and de novo sequencing of 4-sulfophenyl isothiocyanate-derivatized peptides by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (2007) crossref