Abstract
P-113, which was originally derived from the human saliva protein histatin 5, is a histidine-rich antimicrobial peptide with the sequence AKRHHGYKRKFH. P-113 is currently undergoing phase II clinical trial as a pharmaceutical agent to fight against fungal infections in HIV patients with oral candidiasis. Previously, we developed a new procedure for the high-yield expression and purification of hG31P, an analogue and antagonist of human CXCL8. Moreover, we have successfully removed lipopolysaccharide (LPS, endotoxin) associated with hG31P in the expression with Escherichia coli. In this paper, we have used hG31P as a novel fusion protein for the expression and purification of P-113. The purity of the expressed P-113 is more than 95% and the yield is 4 mg P-113 per liter of E. coli cell culture in Luria-Bertani (LB) medium. The antimicrobial activity of the purified P-113 was tested. Furthermore, we used circular dichroism (CD) and nuclear magnetic resonance (NMR) spectroscopy to study the structural properties of P-113. Our results indicate that using hG31P as a fusion protein to obtain large quantities of P-113 is feasible and is easy to scale up for commercial production. An effective way of producing enough P-113 for future clinical studies is evident in this study.
Highlights
Antimicrobial peptides normally consist of 12 to 50 amino acids in length and fold into α-helices, β-sheets, extended, and looped peptides [1,2]
P-113 is in phase II clinical trial for HIV patients with oral candidiasis [4]
We have used hG31P as a fusion protein to successfully express hG31P-P-113 in E. coli
Summary
Antimicrobial peptides normally consist of 12 to 50 amino acids in length and fold into α-helices, β-sheets, extended, and looped peptides [1,2]. The main microbicidal mechanism of antimicrobial peptides to fight against bacteria and fungi is to incorporate and permeabilize the microbial membranes, causing the destruction of the microbes [3]. Because of this special mechanism, antimicrobial peptides may serve as promising therapeutic agents in the fight against antibiotic resistances [4]. It was reported that P-113 is safe and effective in the clinical trials for HIV patients with oral candidiasis [6]. The results showed that P-113 reduces gingivitis, gingival bleeding, and plaque in humans, and it is safe. The advantages of P-113 over the Molecules 2018, 23, 800; doi:10.3390/molecules23040800 www.mdpi.com/journal/molecules
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