Ferrisilicalite‐1 Zeolite Monolith‐Based Affinitive Recognition of Intracellular Phosphorylated Protein Alteration in Tetrahymena thermophila Exposed to Photodegradates of Roxarsone and p‐Arsanilic Acid

Abstract

Research ArticleFerrisilicalite-1 Zeolite Monolith-Based Affinitive Recognition of Intracellular Phosphorylated Protein Alteration in Tetrahymena thermophila Exposed to Photodegradates of Roxarsone and p-Arsanilic AcidIn this study, heteroatomic ferrisilicalite-1 zeolite monolith (FS-1 ZM) was developed for affinitive recognition of intracellular phosphorylated protein in Tetrahymena thermophila after exposure to organoarsenic feed additives (roxarsone, ROX; p-arsanilic acid, PASA) and their photodegradates. ROX or PASA can be photodegradated into toxic inorganic arsenicals, which has become a public concern. After T. thermophila being exposed to ROX or PASA and their photodegradates, it is of significance to address intracellular protein phosphorylation alteration. However, it is challenging for effective capture of phosphorylated protein under the interference of arsenicals uptake. Followed by ferrisilicalite-1 zeolite monolith based immobilization process, sodium dodecyl sulfatepolyacrylamide electrophoresis, in-gel enzymatic digestion, and subsequent LC-MS/MS methodology, the integrated protocol was firstly attempted for identification of arsenicinduced phosphorylated protein alteration. It was found there was little impact of ROX or PASA on intracellular phosphorylated protein distribution. Alternatively, phosphorylated 14-3-3e was down-regulated after the cells were exposed to photodegradates of ROX or PASA, which were similar to those following exposure to inorganic arsenicals. Overall, inorganic arsenicals present during the degradative pathways of ROX or PASA affect the signal transduction system of T. thermophila cells. The down-regulation of phosphorylated 14-3-3e would be further studied as potential biomarker for organoarsenicals released into the environment.Abbreviations: ABS, acetate buffer solution; BSA, bovine serum albumin; FS-1 ZM, ferrisilicalite-1 zeolite monolith; HG-AFS, hydride generationatomic fluorescence spectrometry method; HSP, heat shock protein; iAs III , inorganic arsenite; iAs V , inorganic arsenate; IMAC, immobilized metal-affinity chromatography; MAPK, mitogen-activated protein kinases; PASA, p-arsanilic acid; PBS, phosphate buffered saline; PTM, post-translational modification; ROX, roxarsone; SDS-PAGE, sodium dodecyl sulfate-polyacrylamide electrophoresis; SEM, scanning electron microscopy