Identification of endogenous site-specific covalent binding of catechol estrogens to serum proteins in human blood

Chieh Ming Fang, Ming Chun Ku, Che Kai Chang, Huei Chen Liang, Tzu Fan Wang, Chih Hsing Wu, Shu Hui Chen

Research output: Contribution to journalArticlepeer-review

12 Citations (Scopus)

Abstract

Protein adducts covalently modified by catechol estrogens (CEs), referred to as estrogenized proteins, are potential biomarkers for estrogen homeostasis or exposure to environmental toxicants. However, serum proteins endogenously modified by CEs and the modification sites remain elusive. In this study, liquid chromatography-mass spectrometry (LC-MS)-based shotgun proteomics is applied to identify site-specific protein estrogenization in human blood via a systematic approach and stringent validation. We showed CEs, namely 2- and 4-hydroxyl estrogens which are regarded as biomarkers for estrogen homeostasis, form covalent bonds with proteins, mainly via side chain Cys, Lys, or His residue. Estrogenization of purified human serum albumin (HSA) and immunoglobulin G (IgG) at specific sites was achieved by co-incubation and used as the standards to confirm the identified estrogenization in serum proteins. Based on a database search, estrogenized peptides derived from serum proteins in patient blood were identified; endogenous estrogenization of HSA and IgG-1 at multiple sites were confirmed as compared to the standards. Based on a test using Ellman's reagent, estrogenization produced stable products and irreversibly abolished the reactivity of Cys34-HSA, which is the most important antioxidant and nitric oxide carrier in blood. Given the importance of estrogen metabolism in environmental toxicology, further exploration of estrogenized proteins is warranted for biomarker discovery and/or new mechanisms in disease process.

Original languageEnglish
Pages (from-to)433-442
Number of pages10
JournalToxicological Sciences
Volume148
Issue number2
DOIs
Publication statusPublished - 2015 Dec 1

All Science Journal Classification (ASJC) codes

  • Toxicology

Fingerprint Dive into the research topics of 'Identification of endogenous site-specific covalent binding of catechol estrogens to serum proteins in human blood'. Together they form a unique fingerprint.

Cite this