Collisionally activated decomposition of modified peptides using a tandem hybrid instrument

Abstract

Analyses are described of small peptides and related compounds using a tandem hybrid mass spectrometer of BEQQ geometry. Collisionally activated decomposition of [M + H]+ ions, generated by fast atom bombardment, was performed in the radio frequency (rf)-only quadrupole. Interpretation of fragmentation was greatly facilitated by analysis of labeled analogs, obtained by 18O exchange of carboxyl oxygens. N-Acetylation was also valuable although significant changes in fragmentation resulted from derivatization. Daughter ion spectra of [M + H]+ ions of angiotensin III showed diagnostic fragmentations throughout the peptide chain.

Tandem mass spectrometry of peptides using hybrid and four-sector instruments: A comparative study

“…Similar tendency toward side-chain cleavages (v and w series) can be observed in high-energy CAD data for peptides with charge localization on the C-terminus (3). Such cleavages are not only useful for confirmation of amino acid residues but are essential for distinguishing isobaric amino acid residues such as Leu/lie (62,63) or -/ß-Asp (80). This type of distinction is not possible under the low-energy collision regimes used for the present analyses.…”

Global Differential Non-Gel Proteomics by Quantitative and Stable Labeling of Tryptic Peptides with Oxygen-18

“…Sequencing-grade modified trypsin was from Promega Corporation (Madison, WI). 18 O-rich water (93.7% pure) was purchased from ARC Laboratories (Amsterdam, The Netherlands). P10 ZipTips (U-C18) were from Millipore Corporation (Billerica, MA).…”

“…[10][11][12][13][14][15][16][17] This enzymatic approach has also been used to label a peptide's carboxylic end, thereby creating a recognizable signature for peptide fragments containing this group and thus helping in assigning the y-type of peptide fragments in spectra leading to more reliable protein identification. [18][19][20][21] Until recently, this labeling method was little used in quantitative studies of complex mixtures of peptides, probably because of variable degrees of labeling due to inefficient isotope incorporation and oxygen back-exchange. However, this tagging procedure, when quantitative, can be extremely useful for differential analysis since it has a number of advantages over metabolic or chemical tagging; (1) the reaction is simple and extra handling steps for the removal of excess reagents are not necessary, (2) oxygen-18 rich water is readily available and the volumes needed are economically justified, (3) every tryptic peptide (except those containing the proteins' C-termini that do not end on a lysine or an arginine) is labeled with the same mass tag, (4) this mass difference allows sufficient separation between the isotope envelopes of the light and heavy peptide variants, while maintaining identical ionization behavior as well as highly similar chromatographic properties, and (5) this procedure is not biased toward the type of sample (e.g., peptides from digested proteomes from body fluids, tissues and biopsies can be labeled as efficiently as those from cultured cells).…”

Stable isotope labeling for matrix‐assisted laser desorption/ionization mass spectrometry and post‐source decay analysis of ribonucleic acids

“…The incorporation of 18 O during enzymatic digestion of proteins followed by MS analysis was first reported by Desiderio and Kai in 1983 34. Later reports followed in which the stable isotope labeling was used for the identification of the C‐terminal peptide of a protein,35, 36 disulfide‐linked peptides37 and N ‐glycosylation site,38 for the assignment of ions produced during collision‐induced dissociation (CID),39–42 for peptide quantitation43–47 and for the identification of marker proteins 48…”