Mass-balanced 1H/2H isotope dipeptide tag for simultaneous protein quantitation and identification.
Mass-balanced (1)H/(2)H isotope dipeptide tags (MBITs) are presented for simultaneous protein quantitation and identification. MBIT is derived from N-acetyl-Ala-Ala dipeptide and conjugated to primary amines of target peptides. (1)H/(2)H isotopes are encoded in the methyl groups of N-acetylated dipeptide: one tag deuterated on the N-acetyl group and another on the C-terminal alanine. MBIT-linked peptides comigrate in reversed-phase liquid chromatography without significant (1)H/(2)H isotope effects and provide 2-plex quantitation signals at 114 and 117 Th as well as peptide sequence information upon MS/MS analysis with MALDI TOF/TOF. MBIT shows good quantitation linearity in a concentration range of 20-250 fmol. The performance of MBIT on protein quantitation and identification is further tested with yeast heat-shock protein (Hsp82p) obtained from three different physiological states. MBIT using nanogram-scale samples produces the relative abundance ratios comparable to those obtained from optical imaging of microgram-scale samples visualized with SYPRO Ruby stain. The MBIT strategy is a simple and low-cost alternative for 2-plex quantitation of proteins and offers possibilities of tuning the 2-plex signal mass window by replacing the N-terminal alanine with other amino acid residues.
Seo
Jongcheol
J
Bio-Nanotechnology Center, Department of Chemistry, Pohang University of Science and Technology, San31 Hyojadong Namgu, Pohang, Korea 790-784.
Suh
Min-Soo
MS
Thangadurai
T Daniel
TD
Kim
Jinhee
J
Rhee
Young Ho
YH
Yoon
Hye-Joo
HJ
Shin
Seung Koo
SK
D000804:Angiotensin II / Q000737:chemistry*; D000818:Animals; D002417:Cattle; D002853:Chromatography, Liquid; D004151:Dipeptides / Q000737:chemistry*; D018841:HSP90 Heat-Shock Proteins / Q000737:chemistry*; D006801:Humans; D006859:Hydrogen / Q000032:analysis*; D007553:Isotope Labeling; D010449:Peptide Mapping; D012441:Saccharomyces cerevisiae / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry*; D012710:Serum Albumin, Bovine / Q000737:chemistry*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
D004151:Dipeptides; C074510:HSP82 protein, S cerevisiae; D018841:HSP90 Heat-Shock Proteins; D029701:Saccharomyces cerevisiae Proteins; D000804:Angiotensin II; D012710:Serum Albumin, Bovine; D006859:Hydrogen