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  • Hashimoto S et al a Serial analysis of gene

    2022-09-14

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    Contents lists available at ScienceDirect
    International Journal of Biological Macromolecules
    Cloning, expression of the truncation of recombinant peroxidase derived from millet bran and its reversal effects on 5-Fu resistance in colorectal cancer
    a Key Laboratory of Chemical Biology and Molecular Engineering of National Ministry of Education, Institute of Biotechnology, Shanxi University, Taiyuan, China
    b School of Life Sciences, Shanxi University, Taiyuan, China
    Article history:
    Keywords:
    Millet bran peroxidase (FMBP)
    Truncated protein
    5-Fu resistance of colorectal cancer 
    A novel peroxidase (FMBP) was extracted and purified from foxtail millet bran in our previous study and it pos-sessed excellent anti-colon cancer activity both in vitro and in vivo. However, the active fragment of FMBP respon-sible for the anti-colon cancer effects remains unclear. In present, three different truncated sequences of FMBP were designed and cloned into a plasmid vector (pMal-s). Three recombinant segments were successfully expressed in host strain Escherichia coli DH5α induced by IPTG (0.3 mM) at 37 °C for 4 h, respectively named MBP-FMBP-1, MBP-FMBP-2 and MBP-FMBP-3. MTT assay showed that only MBP-FMBP-2 possessed significant anti-colon cancer activity, and its anti-colon cancer activity was equivalent to FMBP. Further, the results showed that MBP-FMBP-2 significantly reversed the 5-Fu resistance in human colorectal cancer HCT-8/Fu cell through inhibiting cell proliferation, promoting cell apoptosis and increasing the intracellular accumulation of 5-Fu. RT-PCR and western blot assays showed that MBP-FMBP-2 also decreased the expression levels of multi-drug resis-tance protein 1 (MRP1), P-glycoprotein (P-gp) and breast cancer resistance protein (BCRP). These results indicate that MBP-FMBP-2 is the effective segment of FMBP which exhibits anti-colon cancer activity and has potential as an outstanding chemotherapeutic agent against colon cancer.
    1. Introduction
    Colorectal cancer (CRC) is a common malignant tumor of the diges-tive system in clinical practice. According to 2018 global cancer data sta-tistics, the incidence and mortality of CRC accounted for 10.2% and 9.2% of the total cancer cases [1]. At present, 5-Fluorouracil (5-Fu) remains a commonly used chemotherapeutic drug in CRC treatment [2–4]. Though 5-Fu has improved the survival rate of CRC patients, long-term use of 5-Fu not only leads to drug resistance of CRC but also causes tremendous side effects to patients [5–7]. Recent studies showed that the functional molecules of nature products possessed the activity of overcoming 5-Fu resistance of tumor. Wen et al. found that curcumin reversed 5-Fu resistance of human colon cancer HCT-8/Fu cell through promoting cell apoptosis and down-regulating Heat Shock Protein 27 and P-gp expression [8]. Ravindranathan et al. study showed that oligo-meric proanthocyanidins (OPCs) from grape seed extract overcame the