Isobaric peptide termini labeling for MS/MS-based quantitative proteomics.
Since its introduction, isobaric peptide labeling has played an important role in relative quantitative comparisons of proteomes. This paper describes isobaric peptide termini labeling (IPTL), a novel approach for the identification and quantification of two differentially labeled states using MS/MS spectra. After endoproteinase Lys-C digestion, peptides were labeled at C-terminal lysine residues with either 2-methoxy-4,5-dihydro-1H-imidazole (MDHI) or with tetradeuterated MDHI-d(4). Subsequently, their N-termini were derivatized either with tetradeuterated succinic anhydride (SA-d(4)) or with SA. The mixed isotopic labeling results in isobaric masses and provided several quantification data points per peptide. The suitability of this approach is demonstrated with MS and MS/MS analyses of Lys-C digests of standard proteins. A conceptually simple quantification strategy with a dynamic range of 25 is achieved through the use of Mascot score ratios. The utility of IPTL for the analysis of proteomes was verified by comparing the well-characterized effect of the antimitotic inhibitor S-Trityl-l-Cysteine (STLC) on HeLa cells that were treated for either 24 or 48 h with the inhibitor. Many apoptosis-linked proteins were identified as being differentially regulated, confirming the suitability of IPTL for the analysis of complex proteomes.
Koehler
Christian J
CJ
The Biotechnology Centre of Oslo, University of Oslo, Oslo, Norway.
Strozynski
Margarita
M
Kozielski
Frank
F
Treumann
Achim
A
Thiede
Bernd
B
D006367:HeLa Cells; D006801:Humans; D007553:Isotope Labeling / Q000379:methods*; D009211:Myoglobin / Q000737:chemistry / Q000378:metabolism; D010449:Peptide Mapping; D010455:Peptides / Q000737:chemistry* / Q000378:metabolism; D040901:Proteomics / Q000379:methods*; D012710:Serum Albumin, Bovine / Q000737:chemistry / Q000378:metabolism; D053719:Tandem Mass Spectrometry / Q000379:methods*
D009211:Myoglobin; D010455:Peptides; D012710:Serum Albumin, Bovine