ABSTRACT
The widespread resistance to antibiotics in pathogenic bacteria makes the development of a new generation of antimicrobials an urgent task. The development of new antibiotics must be accompanied by a comprehensive study of all of their biological activities in order to avoid adverse side-effects from their application. Some promising antibiotic prototypes derived from the structures of arenicins, antimicrobial peptides from the lugworm Arenicola marina, have been developed. Previously, we described the ability of natural arenicins -1 and -2 to modulate the human complement system activation in vitro. In this regard, it seems important to evaluate the effect of therapeutically promising arenicin analogues on complement activation. Here, we describe the complement-modulating activity of three such analogues, Ar-1[V8R], ALP1, and AA139. We found that the mode of action of Ar-1[V8R] and ALP1 on the complement was similar to that of natural arenicins, which can both activate and inhibit the complement, depending on the concentration. However, Ar-1[V8R] behaved predominantly as an inhibitor, showing only a moderate increase in C3a production in the alternative pathway model and no enhancement at all of the classical pathway of complement activation. In contrast, the action of ALP1 was characterized by a marked increase in the complement activation through the classical pathway in the concentration range of 2.5-20 µg/mL. At the same time, at higher concentrations (80-160 µg/mL), this peptide exhibited a complement inhibitory effect characteristic of the other arenicins. Peptide AA139, like other arenicins, exhibited an inhibitory effect on complement at a concentration of 160 µg/mL, but was much less pronounced. Overall, our results suggest that the effect on the complement system should be taken into account in the development of antibiotics based on arenicins.
Subject(s)
Polychaeta , Animals , Humans , Polychaeta/chemistry , Antimicrobial Cationic Peptides/pharmacology , Prospective Studies , Helminth Proteins/pharmacology , Anti-Bacterial Agents/pharmacology , Anti-Bacterial Agents/chemistry , Complement ActivationABSTRACT
Two cyclic theta-defensin peptides were isolated from leukocytes of the hamadryas baboon, Papio hamadryas, and purified to homogeneity by gel electrophoresis and reversed-phase high-performance liquid chromatography. Both peptides had high in vitro activity against Escherichia coli, Listeria monocytogenes, methicillin-resistant Staphylococcus aureus (MRSA) and Candida albicans. Here, we report their de novo sequencing by matrix-assisted laser desorption/ionization tandem time-of-flight mass spectrometry (MALDI-TOF/TOF-MS). This was accomplished by combining conventional enzymatic digestion with N-terminal derivatization by 2-sulfobenzoic acid cyclic anhydride (SACA) or 4-sulfophenylisothiocyanate (SPITC) to facilitate the interpretation of fragment ion spectra. In addition to the two cyclic theta-defensins (PhTDs) we also sequenced a novel Papio hamadryas alpha-defensin, PhD-4, which showed high sequence homology to rhesus alpha-defensin RMAD-1 and human alpha-defensin HNP-1.
Subject(s)
Defensins/chemistry , Leukocytes/chemistry , Papio hamadryas/blood , Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization/methods , Animals , Bacteria/drug effects , Benzenesulfonates/chemistry , Benzoates/chemistry , Candida albicans/drug effects , Computer Simulation , Defensins/genetics , Defensins/metabolism , Defensins/pharmacology , Isothiocyanates/chemistry , Microbial Sensitivity Tests , Papio hamadryas/genetics , Peptide Fragments/chemistry , Peptide Fragments/genetics , Peptide Fragments/metabolism , Peptide Mapping , Sequence Homology, Amino Acid , alpha-Defensins/chemistry , alpha-Defensins/geneticsABSTRACT
A novel 40-residue antimicrobial peptide, aurelin, exhibiting activity against Gram-positive and Gram-negative bacteria, was purified from the mesoglea of a scyphoid jellyfish Aurelia aurita by preparative gel electrophoresis and RP-HPLC. Molecular mass (4296.95 Da) and complete amino acid sequence of aurelin (AACSDRAHGHICESFKSFCKDSGRNGVKLRANCKKTCGLC) were determined. Aurelin has six cysteines forming three disulfide bonds. The total RNA was isolated from the jellyfish mesoglea, RT-PCR and cloning were performed, and cDNA was sequenced. A 84-residue preproaurelin contains a putative signal peptide (22 amino acids) and a propiece of the same size (22 amino acids). Aurelin has no structural homology with any previously identified antimicrobial peptides but reveals partial similarity both with defensins and K+ channel-blocking toxins of sea anemones and belongs to ShKT domain family.
Subject(s)
Anti-Infective Agents/chemistry , Antimicrobial Cationic Peptides/chemistry , Amino Acid Sequence , Animals , Anti-Infective Agents/isolation & purification , Anti-Infective Agents/pharmacology , Antimicrobial Cationic Peptides/isolation & purification , Antimicrobial Cationic Peptides/pharmacology , Base Sequence , Cloning, Molecular , Defensins/chemistry , Escherichia coli/drug effects , Listeria monocytogenes/drug effects , Marine Toxins/chemistry , Molecular Sequence Data , Scyphozoa/chemistryABSTRACT
Two novel 21-residue antimicrobial peptides, arenicin-1 and arenicin-2, exhibiting activity against Gram-positive and Gram-negative bacteria and fungi, were purified from coelomocytes of marine polychaeta Arenicola marina (lugworm) by preparative gel electrophoresis and RP-HPLC. Molecular masses (2758.3 and 2772.3 Da) and complete amino acid sequences (RWCVYAYVRVRGVLVRYRRCW and RWCVYAYVRIRGVLVRYRRCW) were determined for each isoform. Each arenicin has one disulfide bond (Cys3-Cys20). The total RNA was isolated from the lugworm coelomocytes, RT-PCR and cloning were performed, and cDNA was sequenced. A 202-residue preproarenicin contains a putative signal peptide (25 amino acids) and a long prodomain. Arenicins have no structure similarity to any previously identified antimicrobial peptides.
