title: Bioengineering and functional characterization of arenicin shortened analogs with enhanced antibacterial activity and cell selectivity.
abstract: New bioengineering approaches are required for development of more active and less toxic antimicrobial peptides. In this study we used β-hairpin antimicrobial peptide arenicin-1 as a template for design of more potent antimicrobials. In particular, six shortened 17-residue analogs were obtained by recombinant expression in Escherichia coli. Besides, we have introduced the second disulfide bridge by analogy with the structure of tachyplesins. As a result, a number of analogs with enhanced activity and cell selectivity were developed. In comparison with arenicin-1, which acts on cell membranes with low selectivity, the most potent and promising its analog termed ALP1 possessed two-fold higher antibacterial activity and did not affect viability of mammalian cells at concentration up to 50 μM. The therapeutic index of ALP1 against both Gram-positive and Gram-negative bacteria was significantly increased compared with that of arenicin-1 while the mechanism of action remained the same. Like arenicin-1, the analog rapidly disrupt membranes of both stationary and exponential phase bacterial cells and effectively kills multidrug-resistant Gram-negative bacteria. Furthermore, ALP1 was shown to bind DNA in vitro at a ratio of 1:1 (w/w). The circular dichroism spectra demonstrated that secondary structures of the shortened analogs were similar to that of arenicin-1 in water solution, but significantly differed in membrane-mimicking environments. This work shows that a strand length is one of the key parameters affecting cell selectivity of β-hairpin antimicrobial peptides.
authors.affiliation: Shemyakin and Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, Miklukho-Maklaya str., 16/10, 117997, Moscow, Russia.
authors.forename: Pavel V
authors.initials: PV
authors.lastname: Panteleev
authors.affiliation: Shemyakin and Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, Miklukho-Maklaya str., 16/10, 117997, Moscow, Russia.
authors.forename: Ilia A
authors.initials: IA
authors.lastname: Bolosov
authors.affiliation: Shemyakin and Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, Miklukho-Maklaya str., 16/10, 117997, Moscow, Russia.
authors.forename: Tatiana V
authors.initials: TV
authors.lastname: Ovchinnikova
mesh_terms: D000595:Amino Acid Sequence; D000900:Anti-Bacterial Agents / Q000737:chemistry / Q000494:pharmacology*; D023181:Antimicrobial Cationic Peptides / Q000737:chemistry / Q000235:genetics / Q000494:pharmacology*; D004912:Erythrocytes / Q000187:drug effects; D004926:Escherichia coli / Q000187:drug effects; D015801:Helminth Proteins / Q000737:chemistry / Q000235:genetics / Q000494:pharmacology*; D006461:Hemolysis; D006801:Humans; D008826:Microbial Sensitivity Tests; D008969:Molecular Sequence Data; D010539:Permeability; D010957:Plasmids / Q000737:chemistry; D011485:Protein Binding; D015202:Protein Engineering; D011550:Pseudomonas aeruginosa / Q000187:drug effects; D013211:Staphylococcus aureus / Q000187:drug effects
chemical_list: D000900:Anti-Bacterial Agents; D023181:Antimicrobial Cationic Peptides; D015801:Helminth Proteins; C496888:arenicin-1, Arenicola marina
keywords: antimicrobial peptide; arenicin; bioengineering; heterological expression; therapeutic index; β-hairpin