Increased chemical acetylation of peptides and proteins in rats after daily ingestion of diacetyl analyzed by Nano-LC-MS/MS

Background. Acetylation alters several protein properties including molecular weight, stability, enzymatic activity, protein protein interactions, and other biological functions. Our previous findings demonstrating that diacetyl/peroxynitrite can acetylate L-lysine, L-histidine, and albumin in vitro led us to investigate whether diacetyl-treated rats suffer protein acetylation as well. Methods. Wistar rats were administered diacetyl daily for four weeks, after which they were sacrificed, and their lung proteins were extracted to be analysed by Nano-LC-MS/MS (Q-TOF). A C18 reversed-phase colurnn and gradient elution with formic acid/acetonitrile solutions from 2 to 50% over 150 min were used to separate the proteins. Protein detection was performed using a microTOE-Q II (QTOF) equipped with captive source and an electrospray-ionization source. The data frommass spectrometry were processed using a Compass 1.7 and analyzed using Protein Scape, software that uses Mascot algorithms to perform protein searches. Results. A set of 3,162 acetylated peptides derived from 351 acetylated proteins in the diacetyl-treated group was identified. Among them, 23 targeted proteins were significantly more acetylated in the diacetyl-treated group than in the PBS control. Protein acetylation of the group treated with 540 mg/kg/day of diacetyl was corroborated by Western blotting analysis. Conclusions. These data support our hypothesis that diacetyl exposure in animals may lead to the generation of acetyl radicals, compounds that attach to proteins, affecting their functions and triggering adverse health problems.Sao Paulo Research Foundation (FAPESP)Brazilian Innovation Agency (FINEP)Univ Fed Sao Paulo, Inst Environm Chem & Pharmaceut Sci, Diadema, SP, BrazilUniv Fed Sul & Sudeste Para, Inst Studies Hlth & Biol, Collect Hlth, Maraba, PA, BrazilFundacao Univ Fed Rondonia, Dept Chem, Porto Velho, RO, BrazilUniv Sao Paulo, Sao Carlos Inst Chem, Sao Carlos, SP, BrazilUniv Sao Paulo, Inst Chem, Dept Fundamental Chem, Sao Paulo, SP, BrazilUniv Fed Sao Paulo, Inst Environm Chem & Pharmaceut Sci, Diadema, SP, BrazilFAPESP: 2012/02514-9FAPESP: 2013/07763-0FAPESP: 2010/01404-0Web of Scienc