Arsenic-induced protein phosphorylation changes in HeLa cells.
Arsenic is well documented as a chemotherapeutic agent capable of inducing cell death while at the same time is considered a human carcinogen and an environmental contaminant. Although arsenic toxicity is well known and has formed an impressive literature over the time, little is known about how its effects are exerted at the proteome level. Protein phosphorylation is an important post-translational modification involved in the regulation of cell signaling and likely is altered by arsenic treatment. Despite the importance of phosphorylation for many regulatory processes in cells, the identification and characterization of phosphorylation, as effected by arsenic through mass spectrometric detection, are not fully studied. Here, we identify phosphorylated proteins, which are related to post-translational modifications after phenylarsine oxide (PAO) inoculation to HeLa cells. PAO was chosen because of its high cytotoxicity, measured earlier in these labs. In this study, size exclusion chromatography coupled to inductively coupled plasma mass spectrometry (SEC-ICP-MS) is used to establish several molecular weight fractions with phosphorylated proteins by monitoring (31)P signal vs. time via ICP-MS. SEC-ICP-MS fractions are collected and then separated by the nano-LC-CHIP/ITMS system for peptide determination. Spectrum Mill and MASCOT protein database search engines are used for protein identification. Several phosphorylation sites and proteins related to post-translational modifications are also identified.
Alp
Orkun
O
Analytical Chemistry Department, Faculty of Pharmacy, Gazi University, 06330 Ankara, Turkey.
Merino
Edward J
EJ
Caruso
Joseph A
JA
D000595:Amino Acid Sequence; D001152:Arsenicals / Q000494:pharmacology*; D002851:Chromatography, High Pressure Liquid; D004305:Dose-Response Relationship, Drug; D006367:HeLa Cells; D006801:Humans; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D010766:Phosphorylation; D011499:Protein Processing, Post-Translational / Q000187:drug effects*; D011506:Proteins / Q000032:analysis* / Q000378:metabolism*
D001152:Arsenicals; D011506:Proteins; C029341:oxophenylarsine