Authors
Zhihong Wu, Bjoern O. Schroeder, Jens-M Schroeder, and Jan Wehkamp
Summary
Antimicrobial peptides represent the first-line host defence against microbial pathogens and an essential component of innate immunity. They have received growing interest because of their potential use as therapeutic antibiotics. Due to the fact that most antimicrobial peptides are toxic to prokaryotic host cells, they are currently often produced by chemical synthesis. However, this is too costly for them to be used when large quantities of antimicrobial peptides are required for investigations and clinical trials. Thus, the convenience and cost efficiencies of bacterial production of antimicrobial peptides have become a bottleneck problem. As an important group of antimicrobial peptides human ß-defensins are cationic peptides with 38-47 amino acid residues showing three strands of anti-parallel β-sheets that provide a compact small structure [1,2]. We describe an optimized strategy for recombinant expression of hBD-1 and its mutants in Escherichia coli, to efficiently produce milligram quantities of pure oxidized and correctly folded hBD-1, which was converted into its fully reduced form in a previous study [3]. Recombinant hBD-1 and its mutants were used in its linearized form for bactericidal testing, nuclear magnetic resonance spectroscopy as well as generation of specific polyclonal antibodies. Here we present a step-by-step protocol, with minor modifications, which might be applied also for production of other cysteine-rich and linearized antimicrobial peptides in E.coli and their specific polyclonal antibodies.Further details
The protocol was published on Protocol Exchange on 20 January 2011. To see the entire protocol, click on the source link.Advertisement
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