title: Quantification and visualization of glutathione S-transferase omega 1 in cells using inductively coupled plasma mass spectrometry (ICP-MS) and fluorescence microscopy.
abstract: We report a novel activity-based and Cu-free click chemistry (CC) mediated methodology for glutathione S-transferase omega 1 (GSTO1) quantification using species-unspecific isotope dilution inductively coupled plasma mass spectrometry (SUID ICP-MS), in which dibenzylcyclooctyne-modified 2-chloroacetamide (DBCO-ChAcA) was designed and synthesized, meanwhile, as a navigator towards GSTO1 for subsequent N3-DOTA-Eu-tagging via Cu-free CC. Using (153)Eu-SUID ICP-MS coupled with size exclusion chromatography (SEC), the LOD (3σ) of GSTO1 reached 6.9 fmol with an RSD of 2.4% at the 0.1 μM level (n = 5) considering the recovery of GSTO1 on the SEC was 96.5 ± 2.4%. The GSTO1 contents in the cells of human hepatocellular carcinoma C7721 and breast carcinoma MCF-7 as well as normal hepatic C7701 without or with cis-platin administration were quantified to be from 1.2 μg/10,000 cells (n = 3, RSD = 4.5%) corresponding to 1.2 × 10(-2) ng per cell to 4.76 μg/10,000 cells (n = 3, RSD = 2.9%) corresponding to 4.76 × 10(-2) ng per cell. For a comparative study, DBCO-ChAcA-fluor 488-based fluorescence microscopy could not alone visualize GSTO1 in the cells but could together with those from the small SH-containing molecules such as GSH and that from extra N3-fluor 488 in the cells. This activity-based CC-mediated tagging/labeling strategy provided an opportunity for ICP-MS-based targeted protein quantification, and is very much expected to find its applications in biological mechanism study and the subsequent drug design.
authors.affiliation: Department of Chemistry and the MOE Key Lab of Spectrochemical Analysis and Instrumentation, College of Chemistry and Chemical Engineering, State Key Lab of Marine Environmental Science, Xiamen University, Xiamen, 361005, China.
authors.forename: Yong
authors.initials: Y
authors.lastname: Liang
authors.forename: Xin
authors.initials: X
authors.lastname: Jiang
authors.forename: Nannan
authors.initials: N
authors.lastname: Tang
authors.forename: Limin
authors.initials: L
authors.lastname: Yang
authors.forename: Haifeng
authors.initials: H
authors.lastname: Chen
authors.forename: Qiuquan
authors.initials: Q
authors.lastname: Wang
mesh_terms: D045744:Cell Line, Tumor; D002477:Cells / Q000737:chemistry* / Q000201:enzymology*; D057930:Click Chemistry; D057075:Enzyme Assays / Q000379:methods*; D005982:Glutathione Transferase / Q000737:chemistry*; D006801:Humans; D013058:Mass Spectrometry / Q000379:methods*; D008856:Microscopy, Fluorescence / Q000379:methods*
chemical_list: C490476:GSTO1 protein, human; D005982:Glutathione Transferase