title: Mass-tag technology responding to intracellular signals as a novel assay system for the diagnosis of tumor.
abstract: A novel mass spectrometry-based assay system for determining protein kinase activity employing mass-tagged substrate peptide probes was used for the diagnosis of tumors. Two peptide probes (H-type and D-type) were synthesized containing the same substrate peptide sequence for protein kinase C (PKC). The molecular weights of the two probes differ because of the incorporation of deuterium into the acetyl groups of the D-type probe. The lysates of the normal and tumor tissue were prepared and reacted with the H- and D-type peptide probes, respectively. The PKC activities of the normal and tumor tissues can be compared simply and directly by calculating the phosphorylated ratio to each peptide probe, obtained from the peak intensity of the mass spectrum after mixing of the two reaction solutions. The phosphorylation ratio for the reaction of the H-type peptide probe with the tumor tissue lysate (B16 melanoma) was more than three times higher than that of the D type peptide probe with the normal skin tissue lysate. These results show that the novel assay system for detecting protein kinase activity using mass-tag technology can be a simple and useful means to profile protein kinase activity for cell or tissue lysate samples, and can be applied to the diagnosis of tumors.
authors.affiliation: CREST, Japan Science and Technology Corporation, Kawaguchi, Japan.
authors.forename: Jeong-Hun
authors.initials: JH
authors.lastname: Kang
authors.forename: Yoshiki
authors.initials: Y
authors.lastname: Katayama
authors.forename: Aishan
authors.initials: A
authors.lastname: Han
authors.forename: Shuhei
authors.initials: S
authors.lastname: Shigaki
authors.forename: Jun
authors.initials: J
authors.lastname: Oishi
authors.forename: Kenji
authors.initials: K
authors.lastname: Kawamura
authors.forename: Riki
authors.initials: R
authors.lastname: Toita
authors.forename: Xiao Ming
authors.initials: XM
authors.lastname: Han
authors.forename: Takeshi
authors.initials: T
authors.lastname: Mori
authors.forename: Takuro
authors.initials: T
authors.lastname: Niidome
mesh_terms: D000818:Animals; D045744:Cell Line, Tumor; D004791:Enzyme Inhibitors / Q000494:pharmacology; D007211:Indoles / Q000494:pharmacology; D008297:Male; D008301:Maleimides / Q000494:pharmacology; D008546:Melanoma, Experimental / Q000175:diagnosis* / Q000188:drug therapy / Q000201:enzymology; D051379:Mice; D008815:Mice, Inbred Strains; D009368:Neoplasm Transplantation; D010455:Peptides / Q000737:chemistry* / Q000378:metabolism; D010766:Phosphorylation; D011493:Protein Kinase C / Q000037:antagonists & inhibitors; D011494:Protein Kinases / Q000032:analysis* / Q000378:metabolism; D012867:Skin / Q000737:chemistry / Q000201:enzymology; D012878:Skin Neoplasms / Q000175:diagnosis* / Q000188:drug therapy / Q000201:enzymology; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods*
chemical_list: D004791:Enzyme Inhibitors; D007211:Indoles; D008301:Maleimides; D010455:Peptides; D011494:Protein Kinases; D011493:Protein Kinase C; C064755:Ro 31-7549