Marcos Pires

Nature Medicine, Published online: 04 July 2019; doi:10.1038/s41591-019-0517-0

Machine learning can use patients’ demographic information and previous clinical history to help physicians select the antibiotics most likely to successfully treat urinary tract infections, despite growing levels of resistance.

The N-terminal residue influences protein stability through N-degron pathways. We used stability profiling of the human N-terminome to uncover multiple additional features of N-degron pathways. In addition to uncovering extended specificities of UBR E3 ligases, we characterized two related Cullin-RING E3 ligase complexes, Cul2^ZYG11B and Cul2^ZER1, that act redundantly to target N-terminal glycine. N-terminal glycine degrons are depleted at native N-termini but strongly enriched at caspase cleavage sites, suggesting roles for the substrate adaptors ZYG11B and ZER1 in protein degradation during apoptosis. Furthermore, ZYG11B and ZER1 were found to participate in the quality control of N-myristoylated proteins, in which N-terminal glycine degrons are conditionally exposed after a failure of N-myristoylation. Thus, an additional N-degron pathway specific for glycine regulates the stability of metazoan proteomes.

Selective by design: The ability to selectively target the membrane‐perturbing potential of peptides towards a distinct cellular population has eluded researchers. Herein, we report the de novo design of a new class of peptide that can either enter or lyse cells in an enzyme‐dependent and peptide‐concentration‐dependent manner.

Abstract

Selectively targeting the membrane‐perturbing potential of peptides towards a distinct cellular phenotype allows one to target distinct populations of cells. We report the de novo design of a new class of peptide whose ability to perturb cellular membranes is coupled to an enzyme‐mediated shift in the folding potential of the peptide into its bioactive conformation. Cells rich in negatively charged surface components that also highly express alkaline phosphatase, for example many cancers, are susceptible to the action of the peptide. The unfolded, inactive peptide is dephosphorylated, shifting its conformational bias towards cell‐surface‐induced folding to form a facially amphiphilic membrane‐active conformer. The fate of the peptide can be further tuned by peptide concentration to affect either lytic or cell‐penetrating properties, which are useful for selective drug delivery. This is a new design strategy to afford peptides that are selective in their membrane‐perturbing activity.

Nature Medicine, Published online: 01 July 2019; doi:10.1038/s41591-019-0516-1

Heat-inactivated Akkermansia muciniphila helps alleviate features of metabolic syndrome in overweight and obese subjects.

Nature Communications, Published online: 24 June 2019; doi:10.1038/s41467-019-10542-0

Standard DNA-based analyses of microbial communities cannot distinguish between active microbes and dead or dormant cells. Here, Couradeau et al. use BONCAT (bioorthogonal non-canonical amino acid tagging), flow cytometry, and 16S rRNA gene amplicon sequencing to identify active microbial cells in soils.

Tuberculosis, caused by the intracellular pathogen Mycobacterium tuberculosis, remains the world’s deadliest infectious disease. Sterilizing chemotherapy requires at least 6 months of multidrug therapy. Difficulty visualizing the subcellular localization of antibiotics in infected host cells means that it is unclear whether antibiotics penetrate all mycobacteria-containing compartments in the cell. Here, we combined correlated light, electron, and ion microscopy to image the distribution of bedaquiline in infected human macrophages at submicrometer resolution. Bedaquiline accumulated primarily in host cell lipid droplets, but heterogeneously in mycobacteria within a variety of intracellular compartments. Furthermore, lipid droplets did not sequester antibiotic but constituted a transferable reservoir that enhanced antibacterial efficacy. Thus, strong lipid binding facilitated drug trafficking by host organelles to an intracellular target during antimicrobial treatment.

Nature Medicine, Published online: 24 June 2019; doi:10.1038/s41591-019-0461-z

Bacteriotherapy using gut-derived bacteria from healthy human infants, but not infants with food allergies, suppresses food allergy symptoms in mice via induction of Foxp3 + Rorγt + regulatory T cells

Nature Medicine, Published online: 24 June 2019; doi:10.1038/s41591-019-0485-4

A closer look at the gut microbiome of elite marathon runners unveils a microbe-encoded enzymatic process that contributes to enhanced athletic performance.

Bacterial spores can remain dormant for years but possess the remarkable ability to germinate, within minutes, once nutrients become available. However, it still remains elusive how such instant awakening of cellular machineries is achieved. Utilizing Bacillus subtilis as a model, we show that YwlE arginine (Arg) phosphatase is crucial for...

Nanoparticles coated with bacterial outer membranes (denoted “OM‐NPs”) are reported as an anti‐adhesion nanomedicine that competes with source bacteria for binding sites and therefore inhibits bacterial adhesion to the host cells. Specifically, OM‐NPs prepared with Helicobacter pylori outer membrane were demonstrated to inhibit bacterial binding to the gastric epithelial cells and stomach tissues.

Abstract

Anti‐adhesion therapies interfere with the bacterial adhesion to the host and thus avoid direct disruption of bacterial cycles for killing, which may alleviate resistance development. Herein, an anti‐adhesion nanomedicine platform is made by wrapping synthetic polymeric cores with bacterial outer membranes. The resulting bacterium‐mimicking nanoparticles (denoted “OM‐NPs”) compete with source bacteria for binding to the host. The “top‐down” fabrication of OM‐NPs avoids the identification of the adhesins and bypasses the design of agonists targeting these adhesins. In this study, OM‐NPs are made with the membrane of Helicobacter pylori and shown to bind with gastric epithelial cells (AGS cells). Treatment of AGS cells with OM‐NPs reduces H. pylori adhesion and such anti‐adhesion efficacy is dependent on OM‐NP concentration and its dosing sequence.

Commensal-derived antigens may contribute to autoimmunity by coincidentally mimicking immune-targeted self-structures. Ruff et al. report that human autoreactive lymphocytes and autoantibodies cross-react with homologous regions expressed by the human gut commensal Roseburia intestinalis. Pathogenic APS-generated autoantibodies target bacterial DNA methyltransferase. Gavage of susceptible mice with R. intestinalis triggers autoimmune pathologies.

Nature Microbiology, Published online: 17 June 2019; doi:10.1038/s41564-019-0480-z

Heteroresistance to multiple antibiotics is prevalent across carbapenem-resistant Enterobacteriaceae clinical isolates, but drug combinations that exploit multiple heteroresistance can be used to effectively treat multidrug-resistant infections.

Nature Chemical Biology, Published online: 17 June 2019; doi:10.1038/s41589-019-0279-5

A chemical proteomics strategy identifies DCAF16 as a potential ubiquitin ligase recruiter for cysteine-directed electrophilic PROTACs to promote the degradation of nuclear proteins.

Nature Communications, Published online: 13 June 2019; doi:10.1038/s41467-019-10556-8

The mechanism by which environmental pH controls the virulence of the pathogen Streptococcus pyogenes is unclear. Here, Do et al. show that changes in pH affect the activity of the virulence regulator RopB via its interaction with a quorum-sensing peptide signal.

Antibiotics testing: The natural product elegaphenone kills Gram‐positive Enterococcus faecalis and inhibits the virulence of Gram‐negative Pseudomonas aeruginosa. Its intrinsic benzophenone structure was utilized to determine its cellular targets by chemical proteomics.

Abstract

Natural products represent a rich source of antibiotics that address versatile cellular targets. The deconvolution of their targets via chemical proteomics is often challenged by the introduction of large photocrosslinkers. Here we applied elegaphenone, a largely uncharacterized natural product antibiotic bearing a native benzophenone core scaffold, for affinity‐based protein profiling (AfBPP) in Gram‐positive and Gram‐negative bacteria. This study utilizes the alkynylated natural product scaffold as a probe to uncover intriguing biological interactions with the transcriptional regulator AlgP. Furthermore, proteome profiling of a Pseudomonas aeruginosa AlgP transposon mutant provided unique insights into the mode of action. Elegaphenone enhanced the elimination of intracellular P. aeruginosa in macrophages exposed to sub‐inhibitory concentrations of the fluoroquinolone antibiotic norfloxacin.

A blessing in disguise: Conjugating a diglycine (GG) to an antibiotic prodrug (chloramphenicol succinate) increases the rate of prodrug activation by increasing the rate of ester‐bond hydrolysis catalyzed by bacterial esterases. Dipeptide conjugation to antibiotic prodrugs can increase antibiotic efficacy and reduce adverse drug effects.

Abstract

Antimicrobial drug resistance demands novel approaches for improving the efficacy of antibiotics, especially against Gram‐negative bacteria. Herein, we report that conjugating a diglycine (GG) to an antibiotic prodrug drastically accelerates intrabacterial ester‐bond hydrolysis required for activating the antibiotic. Specifically, the attachment of GG to chloramphenicol succinate (CLsu) generates CLsuGG, which exhibits about an order of magnitude higher inhibitory efficacy than CLsu against Escherichia coli. Further studies reveal that CLsuGG undergoes rapid hydrolysis, catalyzed by intrabacterial esterases (e.g., BioH and YjfP), to generate chloramphenicol (CL) in E. coli. Importantly, the conjugate exhibits lower cytotoxicity to bone marrow stromal cells than CL. Structural analogues of CLsuGG indicate that the conjugation of GG to an antibiotic prodrug is an effective strategy for accelerating enzymatic prodrug hydrolysis and enhancing the antibacterial efficacy of antibiotics.

Dap on target: By introducing the non‐proteinogenic amino acid 2,3‐diaminopropionic acid (Dap), a pH‐responsive anticancer peptide has been developed. The Dap peptide exerts over tenfold increased toxicity at pH 6.0 specific to cancer tissues compared with that at pH 7.4. This strategy will lead to a new mechanism of cancer tissue targeting to enhance cancer selectivity.

Abstract

Endowment of pH responsivity to anticancer peptides is a promising approach to achieve better selectivity to cancer tissues. In this research, a template peptide was designed based on magainin 2, an antimicrobial peptide with anticancer activity, and a series of peptides were designed by replacing different numbers of lysine with the unnatural amino acid, 2,3diaminopropionic acid (Dap), which has a positive charge at weakly acidic pH in cancer tissues, but is neutral at physiological pH 7.4. These Dap‐containing peptides are expected to interact more strongly with tumor cells than with normal cells because 1) weakly acidic conditions form in tumors, and 2) the membrane of tumor cells is more anionic than that of normal cells. Although all examined peptides showed potent cytotoxicities to multidrug‐resistant cancer cells at a weakly acidic pH (ED₅₀≈5 μm), the toxicity decreased with an increase in the number of Dap at pH 7.4 (8 Dap residues resulted in ED₅₀≈60 μm). Furthermore, the introduction of Dap reduced cytotoxicity against normal cells. Thus, Dap led to significantly improved cancer targeting due to a pH‐dependent charge shift. Fluorescence imaging and model membrane experiments supported this charge‐shift model.

Nature Biotechnology, Published online: 27 May 2019; doi:10.1038/s41587-019-0135-x

A general approach to mask antibodies using coiled-coil domains improves anti-tumor therapeutic efficacy.

Membranes matter: Peptides that target lipid II are promising templates for antibiotic development. Unfortunately, almost all data on their binding mechanisms have been obtained in artificial membrane mimics, which might not necessarily reflect the physiologically relevant situation. This might be solved by modern structural biology approaches that provide direct access to the pharmacologically relevant states in bacterial cell membranes.

Abstract

The alarming rise of antimicrobial resistance (AMR) imposes severe burdens on healthcare systems and the economy worldwide, urgently calling for the development of new antibiotics. Antimicrobial peptides could be ideal templates for next‐generation antibiotics, due to their low propensity to cause resistance. An especially promising branch of antimicrobial peptides target lipid II, the precursor of the bacterial peptidoglycan network. To develop these peptides into clinically applicable compounds, detailed information on their pharmacologically relevant modes of action is of critical importance. Here we review the binding modes of a selection of peptides that target lipid II and highlight shortcomings in our molecular understanding that, at least partly, relate to the widespread use of artificial membrane mimics for structural studies of membrane‐active antibiotics. In particular, with the example of the antimicrobial peptide nisin, we showcase how the native cellular membrane environment can be critical for understanding of the physiologically relevant binding mode.

Wu et al. show that in Mycobacterium smegmatis, the putative cell wall amidase Ami3 can accumulate to toxicity under the stabilizing influence of Pmt mannosylation. To control Ami3 levels, an essential complex between the periplasmic serine protease HtrA and the lipoprotein LppZ regulates Ami3 levels, maintaining cellular integrity.

It is widely believed that preference in recognition of structural features of bacterial peptidoglycan (PG) drives specificity in the Drosophila antibacterial response. Vaz et al. challenge this dogma by showing that accessibility to PG plays a central role in bacterial sensing and that structural discrimination is much less stringent than previously thought.

Nature Communications, Published online: 22 May 2019; doi:10.1038/s41467-019-10354-2

Turning tumour promoting macrophages into an anti-tumour phenotype is an attractive therapeutic strategy. Here, the authors develop a polysaccharide-based structure that mimicks pathogen-associated molecular patterns and, by activating the toll-like receptors on macrophage surface, promotes a safe anti-tumour immune response in mouse models.

Straighten up! During domain swapping, proteins mutually interconvert structural elements to form a di‐/oligomer. To achieve domain swapping by design, insertion of a rigid polyproline rod has been investigated. Crystal structure and small‐angle X‐ray scattering analyses confirm the extended orientation of the two subunits.

Abstract

During domain swapping, proteins mutually interconvert structural elements to form a di‐/oligomer. Engineering this process by design is important for creating a higher order protein assembly with minimal modification. Herein, a simple design strategy is shown for domain‐swapping formation by loop deletion and insertion of a polyproline rod. Crystal structures revealed the formation of the domain‐swapped dimers and polyproline portion formed a polyproline II (PPII) structure. Small‐angle X‐ray scattering demonstrated that an extended orientation of domain‐swapped dimer was retained in solution. It is found that a multiple of three of inserting proline residue is favored for domain swapping because of the helical nature of PPII. The rigid nature of the polyproline rod enables precise control of the interdomain distance and orientation.