Fusion expression of recombinant human beta-defensin-3 and analysis of its biological activity

Abstract

Human beta-defensins (hBDs) are small cationic antimicrobial peptides with multiple biologic activities. The aim of the study was cloning, expression in E.coli, purification and in vitro analysis of biological activity of recombinant human beta-defensin-3 (rec-hBD-3). hBD-3 cDNA was cloned into pGEX-2T vector, and recombinant plasmid was transformed into E.coli BL21(DE3) cells. Rec-hBD-3 was expressed in bacterial cells as GST-hBD-3 fusion protein, and purified by 3-step procedure via affine chromatography on glutathione-agarose, cleavage of fusion protein by thrombin, and reverse phase chromatography on Sep-Pack C18. Analysis of biological activity of rec-hBD-3 has shown that the peptide is active against Pseudomonas aeruginosa in micromolar concentrations in radial diffusion test. Rec-hBD-3 did not affect proliferation and viability of cultured human cancer cells of A431, A549, and TPC-1 lines, but was capable to potentiate cytotoxic effects of rec-hBD-2 and docetaxel in vitro.

Keywords

• human beta-defensin-3, cancer cell, proliferation, viability, antimicrobial activity