theta-Defensins: Cyclic Peptides with Endless Potential

theta-Defensins, the only cyclic peptides of animal origin, have been isolated from the leukocytes of rhesus macaques and baboons. Their biogenesis is unusual because each peptide is an 18-residue chimera formed by the head-to-tail splicing of nonapeptides derived from two separate precursors. theta-Defensins have multiple arginines and a ladder-like tridisulfide array spanning their two antiparallel beta-strands. Human theta-defensin genes contain a premature stop codon that prevents effective translation of the needed precursors; consequently, these peptides are not present in human leukocytes. Synthetic theta-defensins with sequences that correspond to those encoded within the human pseudogenes are called retrocyclins. Retrocyclin-1 inhibits the cellular entry of HIV-1, HSV, and influenza A virus. The rhesus theta-defensin RTD-1 protects mice from an experimental severe acute respiratory syndrome coronavirus infection, and retrocyclin- 1 protects mice from infection by Bacillus anthracis spores. The small size, unique structure, and multiple host defense activities of theta-defensins make them intriguing potential therapeutic agents

High Fidelity Processing and Activation of the Human α-Defensin HNP1 Precursor by Neutrophil Elastase and Proteinase 3

The azurophilic granules of human neutrophils contain four α-defensins called human neutrophil peptides (HNPs 1–4). HNPs are tridisulfide-linked antimicrobial peptides involved in the intracellular killing of organisms phagocytosed by neutrophils. The peptides are produced as inactive precursors (proHNPs) which are processed to active microbicides by as yet unidentified convertases. ProHNP1 was expressed in E. coli and the affinity-purified propeptide isolated as two species, one containing mature HNP1 sequence with native disulfide linkages (“folded proHNP1”) and the other containing non-native disulfide linked proHNP1 conformers (misfolded proHNP1). Native HNP1, liberated by CNBr treatment of folded proHNP1, was microbicidal against Staphylococcus aureus, but the peptide derived from misfolded proHNP1 was inactive. We hypothesized that neutrophil elastase (NE), proteinase 3 (PR3) or cathepsin G (CG), serine proteases that co-localize with HNPs in azurophil granules, are proHNP1 activating convertases. Folded proHNP1 was converted to mature HNP1 by both NE and PR3, but CG generated an HNP1 variant with an N-terminal dipeptide extension. NE and PR3 cleaved folded proHNP1 to produce a peptide indistinguishable from native HNP1 purified from neutrophils, and the microbicidal activities of in vitro derived and natural HNP1 peptides were equivalent. In contrast, misfolded proHNP1 conformers were degraded extensively under the same conditions. Thus, NE and PR3 possess proHNP1 convertase activity that requires the presence of the native HNP1 disulfide motif for high fidelity activation of the precursor in vitro

Peptides and Peptidomimetics for Antimicrobial Drug Design

The purpose of this paper is to introduce and highlight a few classes of traditional antimicrobial peptides with a focus on structure-activity relationship studies. After first dissecting the important physiochemical properties that influence the antimicrobial and toxic properties of antimicrobial peptides, the contributions of individual amino acids with respect to the peptides antibacterial properties are presented. A brief discussion of the mechanisms of action of different antimicrobials as well as the development of bacterial resistance towards antimicrobial peptides follows. Finally, current efforts on novel design strategies and peptidomimetics are introduced to illustrate the importance of antimicrobial peptide research in the development of future antibiotics

Bacteria and fungi respond differently to multifactorial climate change in a temperate heathland, traced with ¹³C-Glycine and FACE CO₂

It is vital to understand responses of soil microorganisms to predicted climate changes, as these directly control soil carbon (C) dynamics. The rate of turnover of soil organic carbon is mediated by soil microorganisms whose activity may be affected by climate change. After one year of multifactorial climate change treatments, at an undisturbed temperate heathland, soil microbial community dynamics were investigated by injection of a very small concentration (5.12 µg C g(-1) soil) of (13)C-labeled glycine ((13)C2, 99 atom %) to soils in situ. Plots were treated with elevated temperature (+1°C, T), summer drought (D) and elevated atmospheric carbon dioxide (510 ppm [CO2]), as well as combined treatments (TD, TCO2, DCO2 and TDCO2). The (13)C enrichment of respired CO2 and of phospholipid fatty acids (PLFAs) was determined after 24 h. (13)C-glycine incorporation into the biomarker PLFAs for specific microbial groups (Gram positive bacteria, Gram negative bacteria, actinobacteria and fungi) was quantified using gas chromatography-combustion-stable isotope ratio mass spectrometry (GC-C-IRMS). Gram positive bacteria opportunistically utilized the freshly added glycine substrate, i.e. incorporated (13)C in all treatments, whereas fungi had minor or no glycine derived (13)C-enrichment, hence slowly reacting to a new substrate. The effects of elevated CO2 did suggest increased direct incorporation of glycine in microbial biomass, in particular in G(+) bacteria, in an ecosystem subjected to elevated CO2. Warming decreased the concentration of PLFAs in general. The FACE CO2 was (13)C-depleted (δ(13)C = 12.2‰) compared to ambient (δ(13)C = ∼-8‰), and this enabled observation of the integrated longer term responses of soil microorganisms to the FACE over one year. All together, the bacterial (and not fungal) utilization of glycine indicates substrate preference and resource partitioning in the microbial community, and therefore suggests a diversified response pattern to future changes in substrate availability and climatic factors

cDNA sequences of three sheep myeloid cathelicidins

AbstractSeveral myeloid antimicrobial peptide precursors have been shown to consist of a N-terminal proregion similar to a protein named cathelin and a structurally varied C-terminal antimicrobial domain. Proteins with these features have been named cathelicidins. In this paper we report the cDNA sequences of three ovine cathelicidins of 155, 160 and 190 residues, respectively, with cationic C-terminal sequences corresponding to putative antimicrobial domains. These are structurally varied and include a Cys-rich sequence of 12 residues, which is similar to the bovine antimicrobial cyclic dodecapeptide, a novel 29 residue sequence named SMAP-29 with a possible α-helical conformation, and a 60 residue sequence named Bac7.5, which appears to be a new member of the Pro- and Arg-rich group of mammalian antimicrobial peptides

Characteristic structural features of indolicidin: effects of the cis-trans isomerism on its conformation

Indolicidin is an antimicrobial peptide showing a broad spectrum of antibacterial and antifungal activities, and according to the cis-trans isomerism of three Xaa-Pro peptide bonds, eight different stereoisomers could be distinguished for this peptide. As the cis-trans isomerism about the Xaa-Pro peptide bonds was not considered in previous studies, the structural features of distinct stereoisomeric forms were not characterized in detail, so far. In this theoretical study, the influences of cis-trans isomerism on the conformation of indolicidin were investigated, as well as the typical structural properties of each stereoisomer were determined, focusing on the secondary structures and intramolecular interactions. Based on the results derived from the molecular dynamics simulations, it could be concluded that the eight different stereoisomeric forms of indolicidin adopted characteristic conformational features. Nevertheless, the appearance of various turn structures and intramolecular interactions proved to be dependent on the cis or trans nature of Xaa-Pro peptide bonds, indicating the relevant role of Pro amino acids in determining the three-dimensional structure of this peptide

Microbicidal Effects of α- and θ-Defensins Against Antibiotic-Resistant \u3cem\u3eStaphylococcus aureus\u3c/em\u3e and \u3cem\u3ePseudomonas aeruginosa\u3c/em\u3e

Antibiotic-resistant bacterial pathogens threaten public health. Because many antibiotics target specific bacterial enzymes or reactions, corresponding genes may mutate under selection and lead to antibiotic resistance. Accordingly, antimicrobials that selectively target overall microbial cell integrity may offer alternative approaches to therapeutic design. Naturally occurring mammalian α- and θ-defensins are potent, non-toxic microbicides that may be useful for treating infections by antibiotic-resistant pathogens because certain defensin peptides disrupt bacterial, but not mammalian, cell membranes. To test this concept, clinical isolates of methicillin-resistant Staphylococcus aureus (MRSA), including vancomycin heteroresistant strains, and ciprofloxacin-resistant Pseudomonas aeruginosa (CipR-PA) were tested for sensitivity to α-defensins Crp-4, RMAD-4 and HNPs 1-3, and to RTD-1, macaque θ-defensin-1. In vitro, 3 μM Crp-4, RMAD-4 and RTD-1 reduced MRSA cell survival by 99%, regardless of vancomycin susceptibility. For PA clinical isolates that differ in fluoroquinolone resistance and virulence phenotype, peptide efficacy was independent of strain ciprofloxacin resistance, site of isolation or virulence factor expression. Thus, Crp-4, RMAD-4 and RTD-1 are effective in vitro antimicrobials against clinical isolates of MRSA and CipR-PA, perhaps providing templates for development of α- and θ-defensin-based microbicides against antibiotic resistant or virulent infectious agents