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Expression, Purification, and Therapeutic Implications of Recombinant sFRP1

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Abstract

Secreted frizzled-related proteins (sFRPs) constitute a family of proteins, which impede the Wnt signaling pathway. Upregulation of the Wnt cascade is one of the multiple facets of carcinogenesis. Herein, we report the expression, solubilization, purification, characterization, and anti-cell proliferative activity of a novel recombinant GST-tagged sFRP1 of human origin. sFRP1 was cloned into pGEX-4T2 bacterial expression vector, and the recombinant protein was overexpressed in Escherichia coli BL21 (DE3). It was solubilized from inclusion bodies with N-lauroylsarcosine and Triton X-100, before being purified to homogeneity using glutathione agarose affinity chromatography column. The purified protein was characterized using Western blotting, MALDI TOF-TOF, and circular dichroism spectroscopy analysis. Homology modeling and docking studies revealed that tagging GST with sFRP1 does not change the binding conformation of the cysteine-rich domain and hence, possibly does not alter its function. The novel anti-proliferative activity of GST-sFRP1 was demonstrated in a dose-dependent manner on two cancer cell lines, viz., HeLa (cervical cancer) and MCF-7 (breast cancer). Also, combination therapy of the protein with chemotherapeutic drugs resulted in enhanced anti-cancer activity. This opens up a new avenue in the application of recombinant sFRP1 for cancer therapeutics.

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Acknowledgments

This work was supported by the Department of Biotechnology (No. BT/01/NE/PS/08) and the Department of Electronics and Information Technology, Government of India for financial support (No. 5(9)/2012-NANO (Vol. II)). The authors acknowledge the aid of the Central Instruments Facility (CIF) and the Centre for Nanotechnology, IIT Guwahati.

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  1. Department of Biotechnology, Indian Institute of Technology Guwahati, Guwahati-39, Assam, India

    Archita Ghoshal & Siddhartha Sankar Ghosh

  2. Centre for Nanotechnology, Indian Institute of Technology Guwahati, Guwahati-39, Assam, India

    Siddhartha Sankar Ghosh

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  1. Archita Ghoshal
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Correspondence to Siddhartha Sankar Ghosh.

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Ghoshal, A., Ghosh, S.S. Expression, Purification, and Therapeutic Implications of Recombinant sFRP1. Appl Biochem Biotechnol 175, 2087–2103 (2015). https://doi.org/10.1007/s12010-014-1354-8

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