Single peptide-based protein identification in XXXX proteome through @entity215 coupled with amino acids coded mass tagging.

@entity535 @entity3928 @entity1 @entity215
Identification of proteins with low sequence coverage using @entity3928 ( @entity3928 ) requires tandem @entity3928 / @entity3928 peptide sequencing. It is very challenging to obtain a complete or to interpret an incomplete tandem @entity3928 / @entity3928 spectrum from fragmentation of a weak peptide ion signal for sequence assignment. Here, we have developed an effective and high-throughput MALDI-TOF-based method for the identification of membrane and other low-abundance proteins with a simple, one-dimensional separation step. In this approach, several stable isotope-labeled @entity535 precursors were selected to mass-tag, in parallel, the @entity1 proteome of @entity1 skin fibroblast cells in a residue-specific manner during in vivo cell culturing. These labeled residues can be recognized by their characteristic isotope patterns in @entity215 spectra. The isotope pattern of particular peptides induced by the different labeled precursors provides information about their @entity535 compositions. The specificity of peptide signals in a peptide mass mapping is thus greatly enhanced, resolving a high degree of mass degeneracy of proteolytic peptides derived from the complex @entity1 proteome. Further, false positive matches in database searching can be eliminated. More importantly, proteins can be accurately identified through a single peptide with its m/z value and partial @entity535 composition. With the increased solubility of hydrophobic proteins in SDS, we have demonstrated that our approach is effective for the identification of membrane and low-abundant proteins with low sequence coverage and weak signal intensity, which are often difficult for obtaining informative fragment patterns in tandem @entity3928 / @entity3928 peptide sequencing analysis.
