ACIDIC PEPTIDE-MEDIATED EXPRESSION OF THE ANTIMICROBIAL PEPTIDE BUFORIN-II AS TANDEM REPEATS IN ESCHERICHIA-COLI

Antimicrobial peptides have received increasing attention as a new pha rmaceutical substance, because of their broad spectrum of antimicrobia l activities and the rapid development of multidrug-resistant pathogen ic microorganisms, The main obstacle to the wide application of antimi crobial peptides has been the lack of a cost-effective, mass-productio n method. A novel mass-production method for an antimicrobial peptide of 21 amino acids, buforin II, which was isolated from the stomach of the amphibian Bufo bufo gargarizans, has been developed. This method i s based on the neutralization of the positive charges of buforin II by fusing to an acidic peptide to avoid the lethal effect of the express ed antimicrobial peptide on the host cells, The fusion peptide was exp ressed in Escherichia coli as tandem repeats to increase the product y ield. Multimers of the acidic peptide-buforin II fusion peptide were e xpressed at high levels without causing damage to the cells. The prese nce of cysteine residues in the acidic peptide was critical for the hi gh level expression of the fusion peptide multimers. Multimers of this fusion peptide were expressed as inclusion bodies, and about 107 mg o f pure buforin II was obtained from 1 L of E. coli culture by cleaving the multimers with CNBr, Recombinant buforin II had an antimicrobial activity identical to that of natural buforin II. These results may le ad to a general, cost-effective solution to the mass production of ant imicrobial peptides and other basic peptides which are lethal to the h ost strain. (C) 1998 Academic Press.