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Recombinant FIP-gat, a Fungal Immunomodulatory Protein from Ganoderma atrum, Induces Growth Inhibition and Cell Death in Breast Cancer Cells

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Key Laboratory of Urban Agriculture (South) Ministry of Agriculture, and Engineering Research Center of Cell & Therapeutic Antibody, Ministry of Education, School of Agriculture and Biology, Shanghai Jiaotong University, Shanghai 200240, People’s Republic of China
Key Laboratory of Systems Biomedicine (Ministry of Education), Shanghai Center for Systems Biomedicine, Shanghai Jiao Tong University, Shanghai 200240, People’s Republic of China
§ Department of Immunology, University of Connecticut Health Center, Farmington, Connecticut 06032, United States
# State Key Laboratory of Microbial Metabolism, School of Life Sciences and Biotechnology, Shanghai Jiao Tong University, Shanghai 200240, People’s Republic of China
*(X.-W.Z.) E-mail: [email protected]. Phone: +86-21-34205778. Fax: +86-21-34205778.
*(W.L.) E-mail: [email protected]. Phone: +86-21-34205885. Fax: +86-21-34206059.
Cite this: J. Agric. Food Chem. 2016, 64, 13, 2690–2698
Publication Date (Web):March 20, 2016
https://doi.org/10.1021/acs.jafc.6b00539
Copyright © 2016 American Chemical Society

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    Abstract

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    FIP-gat, an immunomodulatory protein isolated from Ganoderma atrum, is a new member of the FIP family. Little is known, however, about its expressional properties and antitumor activities. It was availably expressed in Escherichia coli with a total yield of 29.75 mg/L. The migration of recombinant FIP-gat (rFIP-gat) on SDS-PAGE corresponded to the predicted molecular mass, and the band was correctly detected by a specific antibody. To characterize the direct effects of rFIP-gat on MDA-MB-231 breast cancer cells, MDA-MB-231 cells were treated with different concentrations of rFIP-gat in vitro; the results showed that this protein could reduce cell viability dose-dependently with a median inhibitory concentration (IC50) of 9.96 μg/mL and agglutinate the MDA-MB-231 cells at a concentration as low as 5 μg/mL. Furthermore, FIP-gat at a concentration of 10 μg/mL can induce significant growth inhibition and cell death in MDA-MB-231 cells. Notably, FIP-gat treatment triggers significant cell cycle arrest at the G1/S transition and pronounced increase in apoptotic cell population. Molecular assays based on microarray and real-time PCR further revealed the potential mechanisms encompassing growth arrest, apoptosis, and autophagy underlying the phenotypic effects.

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    The Supporting Information is available free of charge on the ACS Publications website at DOI: 10.1021/acs.jafc.6b00539.

    • Primers and amplicons for Cyber Green-based real-time PCR (Table S1) (PDF)

    • Characterization of pET30a::FIP-gat plasmid by PCR and restriction enzyme digestion (Figure S1) (PDF)

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