Abstract
Phthalates are a class of plasticizers that have been characterized as endocrine disrupters, and are associated with genital diseases, cardiotoxicity, hepatotoxicity, and nephrotoxicity in the GeneOntology gene/protein database. In this study, we synthesized phthalic acid chemical probes and demonstrated differing protein–protein interactions between MCF-7 cells and MDA-MB-231 breast cancer cell lines. Phthalic acid chemical probes were synthesized using silicon dioxide particle carriers, which were modified using the silanized linker 3-aminopropyl triethoxyslane (APTES). Incubation with cell lysates from breast cancer cell lines revealed interactions between phthalic acid and cellular proteins in MCF-7 and MDA-MB-231 cells. Subsequent proteomics analyses indicated 22 phthalic acid-binding proteins in both cell types, including heat shock cognate 71-kDa protein, ATP synthase subunit beta, and heat shock protein HSP 90-beta. In addition, 21 MCF-7-specific and 32 MDA-MB-231 specific phthalic acid-binding proteins were identified, including related proteasome proteins, heat shock 70-kDa protein, and NADPH dehydrogenase and ribosomal correlated proteins, ras-related proteins, and members of the heat shock protein family, respectively.
Highlights
Phthalates such as di-(2-ethylhexyl) phthalate (DEHP), butyl benzyl phthalate (BBP), di-isononyl phthalate (DINP), di-isodecyl phthalate (DIDP), di-ethyl phthalate (DEP), di-isobutyl phthalate (DIBP), and di-n-butyl phthalate (DBP) are globally used to increase the plasticity of plastic products, and are detected in bags, infants’ toys, bottles, flooring, and cosmetics
After reaction of carboxylic groups with 1-Ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC)/N-hydroxysuccinimide (NHS), phthalic acid was generated for 12 h and synthetic probes were characterized using infrared spectroscopy (IR) [31]
Chemical probes were individually incubated with MCF-7 and MDA-MB-231 cell lysates, and phthalic acid-bound proteins were identified using LC-mass spectrometry (MS)/MS (Figure 1)
Summary
Phthalates such as di-(2-ethylhexyl) phthalate (DEHP), butyl benzyl phthalate (BBP), di-isononyl phthalate (DINP), di-isodecyl phthalate (DIDP), di-ethyl phthalate (DEP), di-isobutyl phthalate (DIBP), and di-n-butyl phthalate (DBP) are globally used to increase the plasticity of plastic products, and are detected in bags, infants’ toys, bottles, flooring, and cosmetics. An increased concentration of phthalates’ metabolites reportedly induced asthma and allergic diseases, and they were detected in urine [7,8]. Urine samples from >75% of Americans contain DEHP and dimethyl phthalate (DMP). Phthalic acid was observed as a secondary metabolite from phthalate derivation, which is observed in dialysis patients [2]. We used phthalic acid as a phthalate precursor to synthesize esterified phthalic acid chemical probes and detect protein–protein interactions. We used chemical probes to characterize phthalic acid-binding proteins in MCF-7 and MDA-MB-231 cells. Quantitative proteomics analyses identified 22 binding proteins that were common to both cell types, including heat shock cognate 71-kDa protein, ATP synthase subunit beta, and heat shock protein HSP 90-beta. ATP synthase subunit beta, heat shock protein HSP 90-beta, and heat shock cognate 71-kDa protein-linked proteasome protein were identified as exclusive MCF-7 proteins, and connected ribosomal correlated proteins were identified as specific to MDA-MB-231 cells
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