title: Identification of new binding sites of human transferrin incubated with organophosphorus agents via Q Exactive LC-MS/MS.
abstract: Organophosphorus agents (OPs) like sarin, VX, or soman could inhibit acetylcholinesterase activity and cause poisoning. OPs could bind many proteins, such as butyrylcholinesterase and albumin, and the adducts formed could identify the exposure. In this paper, we studied human transferrin, which was one of the proteins that could be labeled by OPs. Pure human transferrin was incubated with an overdose of organophosphorus agents, including sarin, soman, VX, tabun, cyclosarin, ethyl tabun, and propyl tabun, and then additional OPs was removed through dialysis. Trypsin was used to cleave the OP-treated proteins and Q Exactive liquid chromatography tandem mass spectrometry (Q Exactive LC-MS/MS) was used to identify them. The present study set out to accomplish two goals. The first goal was to find a good method for identifying multiple binding sites on a given protein through Q Exactive LC-MS/MS. The second goal was to investigate the labeled peptides when transferrin was incubated with a numerous molar excess of OPs. Results showed that tyrosine, lysine, and serine formed covalent bonds with OPs. Twenty OP-labeled sites were found: ten tyrosine sites (including two reported sites), seven lysine sites, and three serine sites. Characteristic fragments for labeled-tyrosine and labeled-lysine adducts were summarized in detail. In conclusion, the method by Q Exactive LC-MS/MS using in this present work is a good way to diagnose exposure to OPs accurately when the binding sites of OPs are uncertain. Novel modified peptides and the characteristic ions found in this work could help investigators assess exposure to OPs.
authors.affiliation: State Key Laboratory of NBC Protection for Civilian, Changping District, Beijing 102205 China. Electronic address: fengjuan_sun@163.com.
authors.forename: Fengjuan
authors.initials: F
authors.lastname: Sun
authors.affiliation: State Key Laboratory of NBC Protection for Civilian, Changping District, Beijing 102205 China. Electronic address: djj224@163.com.
authors.forename: Junjie
authors.initials: J
authors.lastname: Ding
authors.affiliation: State Key Laboratory of NBC Protection for Civilian, Changping District, Beijing 102205 China. Electronic address: yuhuilan1@163.com.
authors.forename: Huilan
authors.initials: H
authors.lastname: Yu
authors.affiliation: State Key Laboratory of NBC Protection for Civilian, Changping District, Beijing 102205 China. Electronic address: gaorunli@163.com.
authors.forename: Runli
authors.initials: R
authors.lastname: Gao
authors.affiliation: State Key Laboratory of NBC Protection for Civilian, Changping District, Beijing 102205 China. Electronic address: wanghongmei@dicp.ac.cn.
authors.forename: Hongmei
authors.initials: H
authors.lastname: Wang
authors.affiliation: State Key Laboratory of NBC Protection for Civilian, Changping District, Beijing 102205 China. Electronic address: chengxin.ricd@yahoo.com.cn.
authors.forename: Chengxin
authors.initials: C
authors.lastname: Pei
mesh_terms: D001665:Binding Sites; D002853:Chromatography, Liquid / Q000379:methods*; D006801:Humans; D008958:Models, Molecular; D009943:Organophosphorus Compounds / Q000737:chemistry / Q000378:metabolism*; D010446:Peptide Fragments / Q000032:analysis / Q000737:chemistry / Q000378:metabolism; D010449:Peptide Mapping; D020539:Sequence Analysis, Protein / Q000379:methods; D053719:Tandem Mass Spectrometry / Q000379:methods*; D014168:Transferrin / Q000032:analysis / Q000737:chemistry / Q000378:metabolism
chemical_list: D009943:Organophosphorus Compounds; D010446:Peptide Fragments; D014168:Transferrin
keywords: Human transferrin; Lysine adducts; Organophosphorus agents; Serine adducts; Tyrosine adducts