Significance of the cyclic structure and of arginine residues for the antibacterial activity of arenicin-1 and its interaction with phospholipid and lipopolysaccharide model membranes

Abstract

Arenicin-1 (Ar-1) is a beta-sheeted antimicrobial peptide from the marine lugworm Arenicola marina. To elucidate the significance of its unique 18-residue cyclic structure and of six cationic arginines for its biological activity and its interaction with biomembranes, we synthesized one linear peptide in which the two cysteines were exchanged for serines (C/S-Ar-1) and a cyclic peptide in which all arginines were replaced by lysines (R/K-Ar-1). We addressed antibacterial and hemolytic activities, the impact of the peptides on bacterial morphology, and their binding to, intercalation into, and permeabilization of model membranes composed of phospholipids or lipopolysaccharide (LPS). In accordance with high salt concentration in sea water, the antibacterial activity of Ar-1 was almost insensitive to high NaCl concentrations. In contrast, the linear derivative lost activity under these conditions against polymyxin B-resistant Proteus mirabilis. Ar-1 intercalated into phospholipid and LPS membranes and formed heterogeneous and short-lived lesions. However, when the peptide was present in both membrane leaflets, it formed defined pores. This characteristic was not observed for the linear derivative C/S-Ar-1. Apparently, the disulfide bond provides conforma-tional stability, which has an impact on salt tolerance, prevents fast degradation by trypsin, and is a prerequisite for the formation of structurally defined pores.

Multiple Peptide Resistance Factor (MprF)-mediated Resistance of Staphylococcus aureus against Antimicrobial Peptides Coincides with a Modulated Peptide Interaction with Artificial Membranes Comprising Lysyl-Phosphatidylglycerol

“…It is non-hemolytic and very stable against tryptic digest. Moreover, it is particularly insensitive toward high salt concentration (10,28). In addition to the natural peptide, we used a linear form where two terminal cysteine residues have been replaced by serine (C/S-Ar-1) (28).…”

Conformational Properties of Arenicins: From the Bulk to the Air–Water Interface

“…teristics in planar lipid bilayer experiments, when compared with the natural peptide Ar-1. [11] The linear peptide displayed also lower cytotoxicity in experiments with human erythrocytes. Studies performed using a Langmuir trough coupled with other techniques can complete the existing information on arenicins, and give the necessary introduction into an understanding of processes accompanying the peptide-membrane interaction.…”

“…To determine the role of this disulfide bond in their antimicrobial activity, peptides with two disulfide bonds were also synthesized by substitution of valines by cysteines [9] and a linear derivative by substitution of cysteines by serins (C/S-Ar-1). [10,11] Positively charged amino acid residues were also considered as playing significant role in biological activity, but the replacement of arginine residues by lysines does not support the necessity of arginine. [11] The tertiary structures of the two natural cyclic arenicin isoforms were determined in solution by NMR spectroscopy.…”

“…[10,11] Positively charged amino acid residues were also considered as playing significant role in biological activity, but the replacement of arginine residues by lysines does not support the necessity of arginine. [11] The tertiary structures of the two natural cyclic arenicin isoforms were determined in solution by NMR spectroscopy. Both exhibit prolonged b-hairpins, stabilized by several non-covalent interactions such as hydrogen bonds, hydrophobic interactions, and p-cation interactions between tyrosine and arginine side chains.…”

“…Biological studies [7,10,11] have shown that both peptides exhibit high antibacterial activity against gram-negative bacteria (Escherichia coli, Pseudomonas aeruginosa) and gram-positive bacteria (Staphylococcus aureus, Staphylococcus epidermidis) with the minimal inhibitory concentrations (MIC) values ranging from 2.0 to 8.0 mm for Ar-1 and 4.0-16.0 mm for C/S-Ar-1. The linear peptide exhibited a higher structural flexibility, [10] which results in the slightly impaired antibacterial activity, higher susceptibility to protease digest, and significantly different pore characThe structures of two antimicrobial peptides (arenicin Ar-1 and its linear derivative C/S-Ar-1) are studied in different solutions and at the air-water interface using spectroscopic methods such as circular dichroism (CD) and infrared reflection absorption spectroscopy (IRRAS) as well as grazing incidence X-ray diffraction (GIXD) and specular X-ray reflectivity (XR).…”