Neue Targets fuer Antibiotika-Therapie. Wirkungsmechanismus und Optimierung des Lantibiotikums Mersacidin zum Einsatz gegen Methicillin-resistente Staphylokokken Schlussbericht

There is an urgent need for new antibiotics with novel targets to avoid future straits in treatment of infections with multi-resistent bacteria such as staphylococci. Identification of the mode of action and of the molecular target of a new antibiotic of the family of lantibiotics (mersacidin). A rich spectrum of biochemical and molecular genetics methods was used. Mersacidin inhibits the biosynthesis of bacterial peptidoglycan by binding to the membrane-bound precursor lipid II. It interacts with lipid II (and prevents it polymerization into macromolecular peptidoglycan) via a novel target binding site; consequently there are no known cross-resistances among bacteria. Mersacidin can be regarded as a prototyp antibiotic of a new class and could be developed for use against multi-resistent bacteria. (orig.)SIGLEAvailable from TIB Hannover: DtF QN1(62,55) / FIZ - Fachinformationszzentrum Karlsruhe / TIB - Technische InformationsbibliothekBundesministerium fuer Bildung, Wissenschaft, Forschung und Technologie, Bonn (Germany)DEGerman

Model membrane studies for characterization of different antibiotic activities of lipopeptides from Pseudomonas

Lipopeptides (LPs) are a structurally diverse class of amphipathic natural products that were in the past mainly known for their surfactant properties. However, the recent discovery of their antimicrobial and cytotoxic bioactivities have fueled and renewed the interest in this compound class. Propelled by the antimicrobial potential of this compound class, in this study a range of six underinvestigated LPs from Pseudomonads were examined with respect to their antibiotic activities towards bacteria. The assays revealed that only the glycosylated lipodipeptide SB-253514, produced by Pseudomonas strain SH-C52, showed significant antibacterial activity. Since the bioactivity of LPs is commonly attributed to membrane interactions, we analyzed the molecular interactions between the LPs and bacteria-like lipid model membranes in more detail via complementary biophysical approaches. Application of the quartz crystal microbalance (QCM) showed that all LPs possess a high binding affinity towards the model membranes. Despite their similar membrane affinity, monolayer studies displayed different tendencies of LPs to incorporate into the membrane. The degree of membrane incorporation could be correlated with specific structural features of the investigated LPs, such as distance between the peptidic macrocycle and the fatty acid, but did not fully reflect their respective antibacterial activity. Cyclic voltammetry (CV) experiments further demonstrated that SB-253514 showed no membrane permeabilization effects at inhibitory concentrations. Collectively, these results suggests that the antibacterial activity of SB-253514 cannot be explained by an unspecific detergent-like mechanism generally proposed for amphiphilic molecules but instead appears to occur via a defined structural target

Proteomic response of Bacillus subtilis to lantibiotics reflects differences in interaction with the cytoplasmic membrane

Mersacidin, gallidermin, and nisin are lantibiotics, antimicrobial peptides containing lanthionine. They show potent antibacterial activity. All three interfere with cell wall biosynthesis by binding lipid II, but they display different levels of interaction with the cytoplasmic membrane. On one end of the spectrum, mersacidin interferes with cell wall biosynthesis by binding lipid II without integrating into bacterial membranes. On the other end of the spectrum, nisin readily integrates into membranes, where it forms large pores. It destroys the membrane potential and causes leakage of nutrients and ions. Gallidermin, in an intermediate position, also readily integrates into membranes. However, pore formation occurs only in some bacteria and depends on membrane composition. In this study, we investigated the impact of nisin, gallidermin, and mersacidin on cell wall integrity, membrane pore formation, and membrane depolarization in Bacillus subtilis. The impact of the lantibiotics on the cell envelope was correlated to the proteomic response they elicit in B. subtilis. By drawing on a proteomic response library, including other envelope-targeting antibiotics such as bacitracin, vancomycin, gramicidin S, or valinomycin, YtrE could be identified as the most reliable marker protein for interfering with membrane-bound steps of cell wall biosynthesis. NadE and PspA were identified as markers for antibiotics interacting with the cytoplasmic membrane

Genome mining of the rhizosphere bacterium Pseudomonas sp. SHC52

Soil ecosystems represent an enormous untapped resource for discovering novel microorganisms, traits and bioactive genes. Natural disease suppressive soils are particularly interesting as they have a relatively higher abundance of beneficial microorganisms that guard plants against infections by soil-borne pathogens. By using both culture-independent and culture-dependent approaches, we recently discovered a novel group of Pseudomonas species in the rhizosphere of sugar beet seedlings grown in a soil that is suppressive to the fungal pathogen Rhizoctonia solani. Representative strain Pseudomonas sp. SHC52 was shown to inhibit hyphal growth of R. solani and various other fungal and oomycete pathogens. Sequencing of Pseudomonas sp. SH-C52 revealed a genome size of 6.7 Mb with approximately 4% of the genome dedicated to secondary metabolism. In detail studies of the genome, will help in determining the genetic factors involved in the activity of this strain

Genome mining of the rhizosphere bacterium Pseudomonas sp. SHC52

Soil ecosystems represent an enormous untapped resource for discovering novel microorganisms, traits and bioactive genes. Natural disease suppressive soils are particularly interesting as they have a relatively higher abundance of beneficial microorganisms that guard plants against infections by soil-borne pathogens. By using both culture-independent and culture-dependent approaches, we recently discovered a novel group of Pseudomonas species in the rhizosphere of sugar beet seedlings grown in a soil that is suppressive to the fungal pathogen Rhizoctonia solani. Representative strain Pseudomonas sp. SHC52 was shown to inhibit hyphal growth of R. solani and various other fungal and oomycete pathogens. Sequencing of Pseudomonas sp. SH-C52 revealed a genome size of 6.7 Mb with approximately 4% of the genome dedicated to secondary metabolism. In detail studies of the genome, will help in determining the genetic factors involved in the activity of this strain

Biomedical applications of nisin

Nisin is a bacteriocin produced by a group of Gram-positive bacteria that belongs to Lactococcus and Streptococcus species. Nisin is classified as a Type A (I) lantibiotic that is synthesized from mRNA and the translated peptide contains several unusual amino acids due to post-translational modifications. Over the past few decades, nisin has been used widely as a food biopreservative. Since then, many natural and genetically modified variants of nisin have been identified and studied for their unique antimicrobial properties. Nisin is an FDA approved and GRAS (generally regarded as safe) peptide with recognized potential for clinical use. Over the past two decades the application of nisin has been extended to biomedical fields. Studies have reported that nisin can prevent the growth of drug-resistant bacterial strains, such as methicillin resistant Staphylococcus aureus, Streptococcus pneumoniae, Enterococci and Clostridium difficile. Nisin has now been shown to have antimicrobial activity against both Gram-positive and Gram-negative disease-associated pathogens. Nisin has been reported to have anti-biofilm properties and can work synergistically in combination with conventional therapeutic drugs. In addition, like host defense peptides, nisin may activate the adaptive immune response and have an immunomodulatory role. Increasing evidence indicates that nisin can influence the growth of tumors and exhibit selective cytotoxicity towards cancer cells. Collectively, the application of nisin has advanced beyond its role as a food biopreservative. Thus, this review will describe and compare studies on nisin and provide insight into its future biomedical applications

Genome mining of the rhizosphere bacterium Pseudomonas sp. SHC52

Soil ecosystems represent an enormous untapped resource for discovering novel microorganisms, traits and bioactive genes. Natural disease suppressive soils are particularly interesting as they have a relatively higher abundance of beneficial microorganisms that guard plants against infections by soil-borne pathogens. By using both culture-independent and culture-dependent approaches, we recently discovered a novel group of Pseudomonas species in the rhizosphere of sugar beet seedlings grown in a soil that is suppressive to the fungal pathogen Rhizoctonia solani. Representative strain Pseudomonas sp. SHC52 was shown to inhibit hyphal growth of R. solani and various other fungal and oomycete pathogens. Sequencing of Pseudomonas sp. SH-C52 revealed a genome size of 6.7 Mb with approximately 4% of the genome dedicated to secondary metabolism. In detail studies of the genome, will help in determining the genetic factors involved in the activity of this strain

Identification of antibacterial peptides from endophytic microbiome

Endophytes, microorganisms living inside plant tissues, are promising producers of lead compounds for the pharmaceutical industry. However, the majority of endophytes are unculturable and therefore inaccessible for functional studies. To evaluate genetic resources of endophytes, we analyzed the biodiversity of fungal microbiome of black crowberry (Empetrum nigrum L.) by next-generation sequencing and found that it consists mainly of unknown taxa. We then separated the host and the endophyte genomes and constructed a fosmid expression library from the endophytic DNA. This library was screened for antibacterial activity against Staphylococcus aureus. A unique antibacterial clone was selected for further analysis, and a gene En-AP1 was identified with no similarity to known sequences. The expressed, folded protein En-AP1 was not active against S. aureus, while tryptic digests exhibited antimicrobial activity. Seven out of twelve synthesized peptides, predicted antibacterial in silico, exhibited in vitro activity towards both S. aureus and Escherichia coli. We propose that the En-AP1 protein is degraded in the library host E. coli and antimicrobial fragments are released from the cell, explaining the in vitro antibacterial activity of the clone. This is the first report of a novel gene expressed in vitro derived from an endophytic microbiome, demonstrating the potential of finding novel genes and compounds from unculturable endophytes