Discovery of lucensimycins A and B from Streptomyces lucensis MA7349 using an antisense strategy

Inhibition of protein synthesis is one of the validated and highly successful targets for inhibition of bacterial growth; this mechanism is a target of a large number of clinical drugs. Ribosomal protein S4, a primary protein, is a potential target for the discovery of antibacterial agents. We describe, using an antisense-sensitized rpsD Streptomyces aureus strain, the discovery and activity of lucensimycins A and B. [structure: see text].     

Natural‐Product‐Inspired Aminoepoxybenzoquinones Kill Members of the Gram‐Negative Pathogen Salmonella by Attenuating Cellular Stress Response

Gram‐negative bacteria represent a challenging task for antibacterial drug discovery owing to their impermeable cell membrane and restricted uptake of small molecules. We herein describe the synthesis of natural‐product‐derived epoxycyclohexenones and explore their antibiotic activity against several pathogenic bacteria. A compound with activity against Salmonella Typhimurium was identified, and the target enzymes were unraveled by quantitative chemical proteomics. Importantly, two protein hits were linked to bacterial stress response, and corresponding assays revealed an elevated susceptibility to reactive oxygen species upon compound treatment. The consolidated inhibition of these targets provides a rationale for antibacterial activity and highlights epoxycyclohexenones as natural product scaffolds with suitable properties for killing Gram‐negative Salmonella.     

New Found Hope for Antibiotic Discovery: Lipid II Inhibitors

Research into antibacterial agents has recently gathered pace in light of the disturbing crisis of antimicrobial resistance. The development of modern tools offers the opportunity of reviving the fallen era of antibacterial discovery through uncovering novel lead compounds that target vital bacterial cell components, such as lipid II. This paper provides a summary of the role of lipid II as well as an overview and insight into the structural features of macrocyclic peptides that inhibit this bacterial cell wall component. The recent discovery of teixobactin, a new class of lipid II inhibitor has generated substantial research interests. As such, the significant progress that has been achieved towards its development as a promising antibacterial agent is discussed.     

Bacterial beta-ketoacyl-acyl carrier protein synthases as targets for antibacterial agents

As a result of increasing drug resistance in pathogenic bacteria, there is a critical need for novel broad-spectrum antibacterial agents. As fatty acid synthesis (FAS) in bacteria is an essential process for cell survival, the enzymes involved in the FAS pathway have emerged as promising targets for antimicrobial agents. Several lines of evidence have indicated that bacterial condensing enzymes are central to the initiation and elongation steps in bacterial fatty acid synthesis and play a pivotal role in the regulation of the entire fatty acid synthesis pathway. beta-ketoacyl-acyl carrier protein (ACP) synthases (KAS) from various bacterial species have been cloned, expressed and purified in large quantities for detailed enzymological, structural and screening studies. Availability of purified KAS from a variety of bacteria, along with a combination of techniques, including combinatorial chemistry, high-throughput screening, and rational drug design based on crystal structures, will undoubtedly aid in the discovery and development of much needed potent and broad-spectrum antibacterial agents. In this review we summarize the biochemical, biophysical and inhibition properties of beta-ketoacyl-ACP synthases from a variety of bacterial species.     

表面抗菌功能涂层的构建及在生物医用材料中的应用研究

近年来,生物医用材料在使用过程中产生的医源性感染问题层出不穷,对人们健康和生命造成严重威胁.表面抗菌涂层构建是解决该类医源性感染问题最有效的策略之一.目前,按照作用机制和功能不同将表面抗菌涂层分为接触式抗菌涂层、抗黏附抑菌涂层、抗黏附杀菌涂层以及智能抗菌涂层.表面抗菌涂层的构建不仅赋予了生物医用材料抗菌性能,有效解决了上述医源性感染问题,还可以提高材料的生物相容性,赋予其抗黏附、抗氧化、生物识别、传感等功能.本文旨在对目前表面抗菌涂层的种类、构建方法以及其在生物医用材料领域中的应用做一全面论述,为进一步开发高性能表面抗菌涂层并扩展其应用提供新思路.     

Membrane Sensor Histidine Kinases: Insights from Structural,Ligand and Inhibitor Studies of Full-Length Proteins and Signalling Domains for Antibiotic Discovery

There is an urgent need to find new antibacterial agents to combat bacterial infections, including agents that inhibit novel, hitherto unexploited targets in bacterial cells. Amongst novel targets are two-component signal transduction systems (TCSs) which are the main mechanism by which bacteria sense and respond to environmental changes. TCSs typically comprise a membrane-embedded sensory protein (the sensor histidine kinase, SHK) and a partner response regulator protein. Amongst promising targets within SHKs are those involved in environmental signal detection (useful for targeting specific SHKs) and the common themes of signal transmission across the membrane and propagation to catalytic domains (for targeting multiple SHKs). However, the nature of environmental signals for the vast majority of SHKs is still lacking, and there is a paucity of structural information based on full-length membrane-bound SHKs with and without ligand. Reasons for this lack of knowledge lie in the technical challenges associated with investigations of these relatively hydrophobic membrane proteins and the inherent flexibility of these multidomain proteins that reduces the chances of successful crystallisation for structural determination by X-ray crystallography. However, in recent years there has been an explosion of information published on (a) methodology for producing active forms of full-length detergent-, liposome- and nanodisc-solubilised membrane SHKs and their use in structural studies and identification of signalling ligands and inhibitors; and (b) mechanisms of signal sensing and transduction across the membrane obtained using sensory and transmembrane domains in isolation, which reveal some commonalities as well as unique features. Here we review the most recent advances in these areas and highlight those of potential use in future strategies for antibiotic discovery. This Review is part of a Special Issue entitled “Interactions of Bacterial Molecules with Their Ligands and Other Chemical Agents” edited by Mary K. Phillips-Jones.     

A revised pathway proposed for Staphylococcus aureus wall teichoic acid biosynthesis based on in vitro reconstitution of the intracellular steps

Resistance to every family of clinically used antibiotics has emerged, and there is a pressing need to explore unique antibacterial targets. Wall teichoic acids (WTAs) are anionic polymers that coat the cell walls of many Gram-positive bacteria. Because WTAs play an essential role in Staphylococcus aureus colonization and infection, the enzymes involved in WTA biosynthesis are proposed to be targets for antibiotic development. To facilitate the discovery of WTA inhibitors, we have reconstituted the intracellular steps of S. aureus WTA biosynthesis. We show that two intracellular steps in the biosynthetic pathway are different from what was proposed. The work reported here lays the foundation for the discovery and characterization of inhibitors of WTA biosynthetic enzymes to assess their potential for treating bacterial infections.     

Targeting bacterial virulence: inhibitors of type III secretion in Yersinia

Agents that target bacterial virulence without detrimental effect on bacterial growth are useful chemical probes for studies of virulence and potential candidates for drug development. Several gram-negative pathogens employ type III secretion to evade the innate immune response of the host. Screening of a chemical library with a luciferase reporter gene assay in viable Yersinia pseudotuberculosis furnished several compounds that inhibit the reporter gene signal expressed from the yopE promoter and effector protein secretion at concentrations with no or modest effect on bacterial growth. The selectivity patterns observed for inhibition of various reporter gene strains indicate that the compounds target the type III secretion machinery at different levels. Identification of this set of inhibitors illustrates the approach of utilizing cell-based assays to identify compounds that affect complex bacterial virulence systems.     

具有双活性序列的新型抗菌肽的设计及性质

设计合成了具有2个活性序列的线性和环状多肽及具有单个活性序列的短链多肽, 研究了它们的杀菌活性、 细胞毒性及溶血性. 结果表明, 线性肽和环状肽的杀菌活性高于短链肽. 利用计算模拟的方法计算了多肽与细菌细胞膜中一种重要的成分磷脂酰甘油(DMPG)的结合能. 结果表明, 多肽-DMPG的结合能与多肽的杀菌活性具有较高的相关性, 线性和环状多肽与DMPG的结合能大于短链肽. 线性和环状多肽均含有2个活性序列, 可提供多个荷正电氨基酸与荷负电的磷脂结合, 结合能较大, 杀菌活性较强. 采用模拟生物膜对其中几条多肽的作用机理进行了初步研究. 结果表明, 该类多肽有可能使正常哺乳动物细胞的细胞膜产生孔洞; 而对于细菌细胞膜, 多肽并未在膜上产生明显孔洞, 而是引起了细菌细胞膜的聚集.     

Isotopically Labeled Desthiobiotin Azide (isoDTB) Tags Enable Global Profiling of the Bacterial Cysteinome

Rapid development of bacterial resistance has led to an urgent need to find new druggable targets for antibiotics. In this context, residue-specific chemoproteomic approaches enable proteome-wide identification of binding sites for covalent inhibitors. Described here are easily synthesized isotopically labeled desthiobiotin azide (isoDTB) tags that shortened the chemoproteomic workflow and allowed an increased coverage of cysteines in bacterial systems. They were used to quantify 59 % of all cysteines in essential proteins in Staphylococcus aureus and enabled the discovery of 88 cysteines that showed high reactivity, which correlates with functional importance. Furthermore, 268 cysteines that are engaged by covalent ligands were identified. Inhibition of HMG-CoA synthase was verified and will allow addressing the bacterial mevalonate pathway through a new target. Overall, a broad map of the bacterial cysteinome was obtained, which will facilitate the development of antibiotics with novel modes-of-action.     

Small organometallic compounds as antibacterial agents

The emergence of bacterial resistance to commercial antibiotics is an issue of global importance. During the last two decades, the number of antibacterial agents that have been discovered and introduced into the market has steadily declined and failed to meet the challenges posed by rapidly increasing resistance of the pathogens against common antibacterial drugs. The development of new classes of compounds to control the virulence of the pathogens is therefore urgently required. This perspective describes the historical development in brief and recent advances on the preparation of small organometallic compounds as new classes of antibacterial agents with potential for clinical development.     

Isotopically Labeled Desthiobiotin Azide (isoDTB) Tags Enable Global Profiling of the Bacterial Cysteinome

Rapid development of bacterial resistance has led to an urgent need to find new druggable targets for antibiotics. In this context, residue‐specific chemoproteomic approaches enable proteome‐wide identification of binding sites for covalent inhibitors. Described here are easily synthesized isotopically labeled desthiobiotin azide (isoDTB) tags that shortened the chemoproteomic workflow and allowed an increased coverage of cysteines in bacterial systems. They were used to quantify 59 % of all cysteines in essential proteins in Staphylococcus aureus and enabled the discovery of 88 cysteines that showed high reactivity, which correlates with functional importance. Furthermore, 268 cysteines that are engaged by covalent ligands were identified. Inhibition of HMG‐CoA synthase was verified and will allow addressing the bacterial mevalonate pathway through a new target. Overall, a broad map of the bacterial cysteinome was obtained, which will facilitate the development of antibiotics with novel modes‐of‐action.     

Applications of Activity‐Based Protein Profiling (ABPP) and Bioimaging in Drug Discovery

Activity‐based protein profiling (ABPP) and bioimaging have been developed in recent years as powerful technologies in drug discovery. Specifically, both approaches can be applied in critical steps of drug development, such as therapy target discovery, high‐throughput drug screening and target identification of bioactive molecules. We have been focused on the development of various strategies that enable simultaneous activity‐based protein profiling and bioimaging studies, thus facilitating an understanding of drug actions and potential toxicities. In this Minireview, we summarize these novel strategies and applications, with the aim of promoting these technologies in drug discovery.     

Pilicides-small molecules targeting bacterial virulence

In a time of emerging bacterial resistance there is a vital need for new targets and strategies in antibacterial therapy. Using uropathogenic Escherichia coli as a model pathogen we have developed a class of compounds, pilicides, which inhibit the formation of virulence-associated organelles termed pili. The pilicides interfere with a highly conserved bacterial assembly and secretion system called the chaperone-usher pathway, which is abundant in a vast number of Gram-negative pathogens and serves to assemble multi-protein surface fibers (pili/fimbriae). This class of compounds provides a platform to gain insight into important biological processes such as the molecular mechanisms of the chaperone-usher pathway and the sophisticated function of pili. Pili are primarily involved in bacterial adhesion, invasion and persistence to host defenses. On this basis, pilicides can aid the development of new antibacterial agents.     

Photo-leucine incorporation reveals the target of a cyclodepsipeptide inhibitor of cotranslational translocation

Photoaffinity labeling is a powerful tool to identify protein targets of biologically active small molecules, yet is often limited by the size, chemical properties, and availability of photoreactive groups. We report an improved synthesis of photo-leucine, a diazirine-based photoreactive analogue of leucine, and demonstrate its incorporation into a cyclodepsipeptide inhibitor of cotranslational translocation. Photoaffinity labeling in a crude membrane fraction, followed by "click chemistry" with a rhodamine-azide reporter, enabled the identification of Sec61alpha, the structural core of the Sec61 translocation channel, as the inhibitor's target.     

Computer-aided design and discovery of protein–protein interaction inhibitors as agents for anti-HIV therapy

Protein–protein interactions (PPI) are involved in most of the essential processes that occur in organisms. In recent years, PPI have become the object of increasing attention in drug discovery, particularly for anti-HIV drugs. Although the use of combinations of existing drugs, termed highly active antiretroviral therapy (HAART), has revolutionized the treatment of HIV/AIDS, problems with these agents, such as the rapid emergence of drug-resistant HIV-1 mutants and serious adverse effects, have highlighted the need for further discovery of new drugs and new targets. Numerous investigations have shown that PPI play a key role in the virus’s life cycle and that blocking or modulating them has a significant therapeutic potential. Here we summarize the recent progress in computer-aided design of PPI inhibitors, mainly focusing on the selection of the drug targets (HIV enzymes and virus entry machinery) and the utilization of peptides and small molecules to prevent a variety of protein–protein interactions (viral–viral or viral–host) that play a vital role in the progression of HIV infection.     

Construction of Antibacterial N‐Halamine Polymer Nanomaterials Capable of Bacterial Membrane Disruption for Efficient Anti‐Infective Wound Therapy

The increasing occurrence of bacterial infection at the wound sites is a serious global problem, demanding the rapid development of new antibacterial materials for wound dressing to avoid the abuse of antibiotics and thereby antibiotic resistance. In this work, the authors first report on antibacterial N‐halamine polymer nanomaterials based on a strategic copolymerization of 3‐allyl‐5,5‐dimethylhydantoin (ADMH) and methyl methacrylate (MMA), which exhibits in vitro and in vivo antimicrobial efficacy against pathogenic bacteria including Staphylococcus aureus and Escherichia coli. Particularly, when a biological evaluation is run for wound therapy, the N‐halamine polymer nanomaterials exhibit a powerful antibacterial efficiency and wound healing ability after a series of histological examination of mouse wound. After the evaluation of biological and chemical surroundings, the proposed four‐stage mechanism suggests that, with unique antibacterial N? Cl bonds, the N‐halamine polymer nanomaterials can disrupt the bacterial membrane, as a result causing intracellular content leaked out and thereby cell death. Based on the synergistic action of antibacterial and wound therapy, the N‐halamine polymer nanomaterials are expected to be promising as wound dressing materials in medical healing and biomaterials.     

Suppressing Alpha-Hemolysin as Potential Target to Screen of Flavonoids to Combat Bacterial Coinfection

The rapid emergence of bacterial coinfection caused by cytosolic bacteria has become a huge threat to public health worldwide. Past efforts have been devoted to discover the broad-spectrum antibiotics, while the emergence of antibiotic resistance encourages the development of antibacterial agents. In essence, bacterial virulence is a factor in antibiotic tolerance. However, the discovery and development of new antibacterial drugs and special antitoxin drugs is much more difficult in the antibiotic resistance era. Herein, we hypothesize that antitoxin hemolytic activity can serve as a screening principle to select antibacterial drugs to combat coinfection from natural products. Being the most abundant natural drug of plant origins, flavonoids were selected to assess the ability of antibacterial coinfections in this paper. Firstly, we note that four flavonoids, namely, baicalin, catechin, kaempferol, and quercetin, have previously exhibited antibacterial abilities. Then, we found that baicalin, kaempferol, and quercetin have better inhibitions of hemolytic activity of Hla than catechin. In addition, kaempferol and quercetin, have therapeutic effectivity for the coinfections of Staphylococcus aureus and Pseudomonas aeruginosa in vitro and in vivo. Finally, our results indicated that kaempferol and quercetin therapied the bacterial coinfection by inhibiting S. aureus α-hemolysin (Hla) and reduced the host inflammatory response. These results suggest that antitoxins may play a promising role as a potential target for screening flavonoids to combat bacterial coinfection.     

Synthesis,Biological Evaluation and Docking Studies of Ring-Opened Analogues of Ipomoeassin F

The plant-derived macrocyclic resin glycoside ipomoeassin F (Ipom-F) binds to Sec61α and significantly disrupts multiple aspects of Sec61-mediated protein biogenesis at the endoplasmic reticulum, ultimately leading to cell death. However, extensive assessment of Ipom-F as a molecular tool and a therapeutic lead is hampered by its limited production scale, largely caused by intramolecular assembly of the macrocyclic ring. Here, using in vitro and/or in cellula biological assays to explore the first series of ring-opened analogues for the ipomoeassins, and indeed all resin glycosides, we provide clear evidence that macrocyclic integrity is not required for the cytotoxic inhibition of Sec61-dependent protein translocation by Ipom-F. Furthermore, our modeling suggests that open-chain analogues of Ipom-F can interact with multiple sites on the Sec61α subunit, most likely located at a previously identified binding site for mycolactone and/or the so-called lateral gate. Subsequent in silico-aided design led to the discovery of the stereochemically simplified analogue 3 as a potent, alternative lead compound that could be synthesized much more efficiently than Ipom-F and will accelerate future ipomoeassin research in chemical biology and drug discovery. Our work may also inspire further exploration of ring-opened analogues of other resin glycosides.     

Synthesis of magnetite nanoparticles for bio- and nanotechnology: genetic engineering and biomimetics of bacterial magnetosomes

Magnetotactic bacteria (MTB) have the ability to navigate along the Earth's magnetic field. This so-called magnetotaxis is a result of the presence of magnetosomes, organelles which comprise nanometer-sized intracellular crystals of magnetite (Fe(3)O(4)) enveloped by a membrane. Because of their unique characteristics, magnetosomes have a high potential for nano- and biotechnological applications, which require a specifically designed particle surface. The functionalization of magnetosomes is possible either by chemical modification of purified particles or by genetic engineering of magnetosome membrane proteins. The second approach is potentially superior to chemical approaches as a large variety of biological functions such as protein tags, fluorophores, and enzymes may be directly incorporated in a site-specific manner during magnetosome biomineralization. An alternative to the bacterial production of magnetosomes are biomimetic approaches, which aim to mimic the bacterial biomineralization pathway in vitro. In MTB a number of magnetosome proteins with putative functions in the biomineralization of the nanoparticles have been identified by genetic and biochemical approaches. The initial results obtained by several groups indicate that some of these proteins have an impact on nanomagnetite properties in vitro. In this article the key features of magnetosomes are discussed, an overview of their potential applications are given, and different strategies are proposed for the functionalization of magnetosome particles and for the biomimetism of their biomineralization pathway.