Dansylation of tryptic peptides for increased sequence coverage in protein identification by matrix‐assisted laser desorption/ionization time‐of‐flight mass spectrometric peptide mass fingerprinting

Abstract

A database search using peptide mass fingerprints obtained by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry leads to protein identification with incomplete sequence coverage, because certain peptides are preferentially desorbed/ionized and some are not detected at all. We show that certain tryptic peptides mainly with C-terminal arginine not detected before derivatization become detectable upon dansylation. Others, mainly with C-terminal lysine, are suppressed. An increase in protein sequence coverage and protein identification score by combined data from underivatized and dansylated peptides in database search is demonstrated using human amnion proteins (human serum albumin precursor, calmodulin, collagen alpha 2(VI) chain precursor, galectin-3) separated by two-dimensional gel electrophoresis as well as femtomole amounts of BSA in solution.

Chemically enhanced liquid chromatography/tandem mass spectrometry determination of glutamic acid in the diffusion medium of retinal cells

“…Thus, different temperature and reaction times were examined to find the optimum conditions for derivatization. The temperature, for example, ranged from room temperature to 80°C (Park et al, 2005;Quirke et al, 1994;Minocha and Long, 2004). Different temperatures and reaction times were examined to find the optimum conditions for derivatization of glutamic acid.…”

Selective detection and identification of phosphopeptides by dansyl MS/MS/MS fragmentation

“…In an attempt to improve and simplify the plethora of methods so far suggested11 we have set up a new approach to label selectively phospho‐Ser/‐Thr residues by exploiting the features of a novel linear ion trap mass spectrometer 12. Dansyl chloride is known to react with the ε‐amino group of lysine as well as with the N‐terminal amino group 10. Here, using chemical manipulations coupled with dansyl chloride labelling, we show a methodology capable of large‐scale proteomic profiling of phosphorylation sites.…”

Recent Advancement of Chemical Derivatization and Its Applications to High Sensitive Analysis of Peptide in Mass Spectrometry

“…Therefore, hydrophobicity and gas-phase basicity of peptides could also be improved via fluorescence derivatization. Recently, several of fluorescent reagents have been used for peptide derivatization to enhance the detection sensitivity of peptides by MS [22] . Karger and co-workers reported that peptides derivatized with coumarin for ionization enhancement.…”