title: Recombinant expression and purification of a MAP30-cell penetrating peptide fusion protein with higher anti-tumor bioactivity.
abstract: MAP30 (Momordica Antiviral Protein 30 Kd), a single-stranded type-I ribosome inactivating protein, possesses versatile biological activities including anti-tumor abilities. However, the low efficiency penetrating into tumor cells hampers the tumoricidal effect of MAP30. This paper describes MAP30 fused with a human-derived cell penetrating peptide HBD which overcome the low uptake efficiency by tumor cells and exhibits higher anti-tumor bioactivity. MAP30 gene was cloned from the genomic DNA of Momordica charantia and the recombinant plasmid pET28b-MAP30-HBD was established and transferred into Escherichia coli BL21 (DE3). The recombinant MAP30-HBD protein (rMAP30-HBD) was expressed in a soluble form after being induced by 0.5mM IPTG for 14h at 15°C. The recombinant protein was purified to greater than 95% purity with Ni-NTA affinity chromatography. The rMAP30-HBD protein not only has topological inactivation and protein translation inhibition activity but also showed significant improvements in cytotoxic activity compared to that of the rMAP30 protein without HBD in the tested tumor cell lines, and induced higher apoptosis rates in HeLa cells analyzed by Annexin V-FITC with FACS. This paper demonstrated a new method for improving MAP30 protein anti-tumor activity and might have potential applications in cancer therapy area.
authors: State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai 200237, China.
authors: Qiang
authors: Q
authors: Lv
authors: State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai 200237, China; Zhejiang Reachall Pharmaceutical Co. Ltd., Zhejiang 322100, China.
authors: Xu-Zhong
authors: XZ
authors: Yang
authors: Zhejiang Reachall Pharmaceutical Co. Ltd., Zhejiang 322100, China.
authors: Long-Yun
authors: LY
authors: Fu
authors: Zhejiang Reachall Pharmaceutical Co. Ltd., Zhejiang 322100, China.
authors: Yv-Ting
authors: YT
authors: Lu
authors: State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai 200237, China.
authors: Yan-Hua
authors: YH
authors: Lu
authors: State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai 200237, China. Electronic address: zhaojian@ecust.edu.cn.
authors: Jian
authors: J
authors: Zhao
authors: Zhejiang Reachall Pharmaceutical Co. Ltd., Zhejiang 322100, China; Shanghai University of Chinese Traditional Medicine, Shanghai 201203, China. Electronic address: wfj@shutcm.edu.cn.
authors: Fu-Jun
authors: FJ
authors: Wang
mesh_terms: D000970:Antineoplastic Agents / Q000737:chemistry / Q000302:isolation & purification / Q000494:pharmacology; D057846:Cell-Penetrating Peptides / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000494:pharmacology; D006367:HeLa Cells; D006801:Humans; D009369:Neoplasms / Q000188:drug therapy* / Q000378:metabolism / Q000473:pathology; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000494:pharmacology; D054790:Ribosome Inactivating Proteins, Type 2 / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000494:pharmacology
chemical_list: D000970:Antineoplastic Agents; D057846:Cell-Penetrating Peptides; C066347:MAP30 protein, Momordica charantia; D011993:Recombinant Fusion Proteins; D054790:Ribosome Inactivating Proteins, Type 2
keywords: Anti-tumor activity; Cell Penetrating Peptide; Ni–NTA purification; Recombinant Momordica Antiviral Protein 30 Kd (rMAP30); Ribosome inactivating protein