Simulating the antimicrobial mechanism of human β-defensin-3 with coarse-grained molecular dynamics

Human β-defensin-3 (HβD-3) is an endogenous antimicrobial peptide with potent and broad killing activity against various microorganisms, and thus, it is an attractive candidate for the development of novel peptide antibiotics, but its antimicrobial mechanism remains elusive. To characterize the mechanism, we used multi-microsecond coarse-grained simulations with the MARTINI force field. These simulations show HβD-3 peptides can form oligomers on the surface of bacterial membrane and make anionic lipids (POPG) clustered. Furthermore, two kinds of regions (one is composed of pure POPG lipids, and the other is enriched in POPE lipids) are formed in the membrane; on the border of them, there are some obvious defects, which result in the membrane disruption. By contrast, the simulations also reveal that the contacts between the HβD-3 peptides and mammalian membrane are not stable. These results provide biophysical insights into HβD-3 selectivity and suggest a possible antimicrobial mechanism.     

Isolation and Some Properties of the Antimicrobial Peptide (Pa-AMP) from the Seeds of Pokeweed (Phytolacca americana).

An antimicrobial peptide, designated Pa-AMP, was purified by gel filtration on Sephadex G-75 followed by S-Sepharose, Cosmosil-SP, and reverse-phase HPLC from the seeds of pokeweed (Phytolacca americana). Pa-AMP is a basic peptide having an isoelectric point of over 10 and its extinction coefficient at 280 nm of 1% aqueous solution was 7.7. Pa-AMP has a molecular mass of 4 kDa and 3.4 kDa on tricine SDS-PAGE under nonreducing and reducing conditions, respectively. The N-terminal amino acid of Pa-AMP was blocked. The concentrations of peptide required for 50% inhibition (IC₅₀) of the growth of plant pathogenic fungi, Gram-positive, and Gram-negative bacteria were 3 to 41 μg/ml. Differing from other peptides, Pa-AMP inhibited the growth of some Gram-negative bacteria.     

Methyl Jasmonate and Lactones Including Jasmine Lactone in Ceylon Tea

The ester and lactone fraction possessing the most attractive aroma was separated from the aroma concentrate of Ceylon flavory tea by silica-gel column chromatography and analyzed by GC-MS.

Methyl 2-(cis-2′-pentenyl)-cyclopentanone-3-acetate(methyl jasmonate), 5-(cis-2′-pentenyl)-5-pentanolide (jasmine lactone), 2,3-dimethyl-2-nonen-4-olide, 4-octanolide, 4-nonanolide and 5-decanolide were newly identified as the constituents of tes aroma. Former two compounds seemed to carry a major share of aroma character of Ceylon flavory tea.     

Selective binding of antimicrobial porphyrins to the heme‐receptor IsdH‐NEAT3 of Staphylococcus aureus

The Isd (iron‐regulated surface determinant) system of the human pathogen Staphylococcus aureus is responsible for the acquisition of heme from the host organism. We recently reported that the extracellular heme receptor IsdH‐NEAT3 captures and transfers noniron antimicrobial porphyrins containing metals in oxidation state (III). However, it is unclear if geometric factors such as the size of the metal (ionic radius) affect binding and transfer of metalloporphyrins. We carried out an ample structural, functional, and thermodynamic analysis of the binding properties of antimicrobial indium(III)‐porphyrin, which bears a much larger metal ion than the iron(III) of the natural ligand heme. The results demonstrate that the NEAT3 receptor recognizes the In(III)‐containing PPIX in a manner very similar to that of heme. Site‐directed mutagenesis identifies Tyr642 as the central element in the recognition mechanism as suggested from the crystal structures. Importantly, the NEAT3 receptor possesses the remarkable ability to capture dimers of metalloporphyrin. Molecular dynamics simulations reveal that IsdH‐NEAT3 does not require conformational changes, or large rearrangements of the residues within its binding site, to accommodate the much larger (heme)₂ ligand. We discuss the implications of these findings for the design of potent inhibitors against this family of key receptors of S. aureus.     

抗植物病原真菌紫叶小檗内生真菌的筛选和鉴定

对紫叶小檗根、茎、叶和果实中分离纯化得到的28株内生真菌进行液体培养,培养物烘干研磨后用丙酮浸提,测定各提取物对5种供试植物病原真菌的抑菌活性,对抗菌谱较广的菌株进行分子鉴定,并分别测定其胞内、胞外代谢产物的抑菌活性.结果显示:(1)紫叶小檗25株内生真菌中有5株菌(R3、S4、L6、F2、F6)的抗菌谱较广,其中,茎中的内生菌S4抑菌活性最强,对5种植物病原真菌的抑菌活性均大于90％,对马铃薯干腐病菌的抑菌率最高达93.06％.(2)经鉴定S4为子囊菌门的Para phaeos phaeria属真菌.(3)S4菌丝体丙酮提取液对马铃薯干腐病菌的抑菌率随其浓度(加入体积)的增加而增大,呈正相关线性关系(y=15.334x+14.618,r=0.99),而S4发酵液对马铃薯干腐病菌的抑菌率随其浓度(加入体积)的增加而减小,呈负相关线性关系(y=-20.196x+64.984,r=0.99),即在较高浓度时促进病原菌生长.研究表明,紫叶小檗内生真菌S4菌株的抑菌活性成分主要为胞内代谢产物.     

The acute-phase protein serum amyloid A3 is expressed in the bovine mammary gland and plays a role in host defence

The serum amyloid A protein is one of the major reactants in the acute-phase response. Using representational difference analysis comparing RNA from normal and involuting quarters of a dairy cow mammary gland, we found an mRNA encoding the SAA3 protein (M-SAA3). The M-SAA3 mRNA was localized to restricted populations of bovine mammary epithelial cells (MECs). It was expressed at a moderate level in late pregnancy, at a low level through lactation, was induced early in milk stasis, and expressed at high levels in most MECs during mid to late involution and inflammation/mastitis. The mature M-SAA3 peptide was expressed in Escherichia coli, antibodies made, and shown to have antibacterial activity against E. coli, Streptococcus uberis and Pseudomonas aeruginosa. These results suggest that the mammary SAA3 may have a role in protection of the mammary gland during remodelling and infection and possibly in the neonate gastrointestinal tract.     

Synthetic analogs of anoplin show improved antimicrobial activities

We present the antimicrobial and hemolytic activities of the decapeptide anoplin and 19 analogs thereof tested against methicillin‐resistant Staphylococcus aureus ATCC 33591 (MRSA), Escherichia coli (ATCC 25922), Pseudomonas aeruginosa (ATCC 27853), vancomycin‐resistant Enterococcus faecium (ATCC 700221) (VRE), and Candida albicans (ATCC 200955). The anoplin analogs contain substitutions in amino acid positions 2, 3, 5, 6, 8, 9, and 10. We use these peptides to study the effect of altering the charge and hydrophobicity of anoplin on activity against red blood cells and microorganisms. We find that increasing the charge and/or hydrophobicity improves antimicrobial activity and increases hemolytic activity. For each strain tested, we identify at least six anoplin analogs with an improved therapeutic index compared with anoplin, the only exception being Enterococcus faecium, against which only few compounds are more specific than anoplin. Both 2Nal⁶ and Cha⁶ show improved therapeutic index against all strains tested. Copyright © 2013 European Peptide Society and John Wiley & Sons, Ltd.     

Self‐assembling organo‐peptide bolaphiles with KLK tripeptide head groups display selective antibacterial activity

In keeping with recent efforts to generate compounds for antibiotic and microbicide development, we focused on the creation of non‐natural organo‐peptide hybrids of antimicrobial peptide amides (KLK(L)_nKLK‐NH₂) derived from sapecin B and a self‐assembling oligoglycine organo‐peptide bolaphile containing an ω‐amino fatty acid residue. The hybrid organo‐peptide bolaphiles with two cationic KLK tripeptide motifs linked with an ω‐amino acid residue (penta‐, octa‐ or undecamethylene chain) maintained the self‐assembling properties of the root oligoglycine bolaphile. Electron microscopy clearly revealed complex supramolecular architectures for both sapecin B‐derived peptides and the hybrid analogues. FT‐IR spectroscopy indicated that the supramolecular structures were composed primarily of β‐sheets. CD revealed that the hybrid bolaphiles did not share the same secondary structures as the sapecin B peptides in solution. However, although secondary structures of antimicrobial peptides are central in the activity, the organo‐peptide bolaphiles also retained the potent antimicrobial activity of the leader sapecin B‐derived peptide against both Gram‐positive and Gram‐negative bacteria. In general, the hybrids were more selective than the sapecin B peptides, as they displayed little or no appreciable haemolytic activity. The results obtained herald a new approach for the design of purpose‐built hybrid organo‐peptide bolaphiles. Copyright © 2013 European Peptide Society and John Wiley & Sons, Ltd.     

Antioxidant and antimicrobial activities of rosemary extracts linked to their polyphenol composition

Rosmarinus officinalis extracts were investigated by a combination of bioassays and biochemical analysis to identify bioactive compounds. The 2,2-diphenyl-2-picrylhydracyl hydrate (DPPH) radical scavenging method, Folin–Ciocaulteau method and HPLC chromatography were used to study the distribution and levels of antioxidants (AOXs). Antimicrobial activity analysis was carried out using the disk diffusion and broth dilution techniques. A good correlation between the AOX activities and total phenol content in the extracts was found. Although all rosemary extracts showed a high radical scavenging activity, a different efficacy as antimicrobial agent was observed. Methanol extract containing 30% of carnosic acid, 16% of carnosol and 5% of rosmarinic acid was the most effective antimicrobial against Gram positive bacteria (minimal inhibition concentration, MIC, between 2 and 15 μg/ml), Gram negative bacteria (MIC between 2 and 60 μg/ml) and yeast (MIC of 4 μg/ml). By contrast, water extract containing only 15% of rosmarinic acid showed a narrow activity. MIC value of the methanol and water extracts is in a good correlation with the values obtained with pure carnosic acid and rosmarinic acid, respectively. Therefore, our results suggested that the antimicrobial rosemary extracts efficacy was associated with their specific phenolic composition. Carnosic acid and rosmarinic acid may be the main bioactive antimicrobial compounds present in rosemary extracts. From a practical point of view, rosemary extract may be a good candidate for functional foods as well as for pharmaceutical plant-based products.