Article ID Journal Published Year Pages File Type
2020707 Protein Expression and Purification 2013 11 Pages PDF
Abstract

Five variants of human β-defensins (HBDs) were expressed in Escherichia coli using two vector systems (pET28a(+) and pQE30) with inducible expression by IPTG. The last vector has not been previously reported as an expression system for HBDs. The recombinant peptides were different in their lengths and overall charge. The HBDs were expressed as soluble or insoluble proteins depending on the expression system used, and the final protein yields ranged from 0.5 to 1.6 mg of peptide/g of wet weight cells, with purities higher than 90%. The recombinant HBDs demonstrated a direct correlation between antimicrobial activity and the number of basic charged residues; that is, their antimicrobial activity was as follows: HBD3-M-HBD2 > HBD3 = HBD3-M = HB2-KLK > HBD2 when assayed against E. coli, Staphylococcus aureus and Pseudomonas aeruginosa. Interestingly, HBD2 had the best antimicrobial activity against the Mycobacterium tuberculosis strain H37Rv (1.5 μM) and the heterologous tandem peptide, HBD3-M-HBD2, had the best minimal inhibitory concentration (MIC) value (2.7 μM) against a multidrug resistance strain (MDR) of M. tuberculosis, demonstrating the feasibility of the use of HBDs against pathogenic M. tuberculosis reported to be resistant to commercial antibiotics.

► Human β-defensins (HBDs) were expressed in the plasmid vectors pET28(a+) and pQE30. ► HBDs were biologically active even with a short peptide fused at their N-terminus. ► The chimera HBD3-HBD2 had similar protein yield and activity to single expressed HBDs. ► Recombinant human β-defensins were active against Mycobacterium tuberculosis.

Related Topics
Life Sciences Biochemistry, Genetics and Molecular Biology Biochemistry
Authors
, , , , ,