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Phase separation underpins many biologically important cellular events such as RNA metabolism, signaling, and CO2 fixation. However, determining the composition of a phase-separated organelle is often challenging due to its sensitivity to environmental conditions, which limits the application of traditional proteomic techniques like organellar purification or affinity purification mass spectrometry to understand their composition. In Chlamydomonas reinhardtii, Rubisco is condensed into a crucial phase-separated organelle called the pyrenoid that improves photosynthetic performance by supplying Rubisco with elevated concentrations of CO2. Here, we developed a TurboID-based proximity labeling technique in which proximal proteins in Chlamydomonas chloroplasts are labeled by biotin radicals generated from the TurboID-tagged protein. By fusing 2 core pyrenoid components with the TurboID tag, we generated a high-confidence pyrenoid proxiome that contains most known pyrenoid proteins, in addition to new pyrenoid candidates. Fluorescence protein tagging of 7 previously uncharacterized TurboID-identified proteins showed that 6 localized to a range of subpyrenoid regions. The resulting proxiome also suggests new secondary functions for the pyrenoid in RNA-associated processes and redox-sensitive iron–sulfur cluster metabolism. This developed pipeline can be used to investigate a broad range of biological processes in Chlamydomonas, especially at a temporally resolved suborganellar resolution. A high-confidence proteome of the CO2-fixing algal pyrenoid is identified by proximity labeling. [ABSTRACT FROM AUTHOR]

Escherichia coli can transport and catabolize the common sialic acid, N-acetylneuraminic acid ( Neu5 Ac), as a sole source of carbon and nitrogen, which is an important mucus-derived carbon source in the mammalian gut. Herein we demonstrate that E. coli can also grow efficiently on the related sialic acids, N-glycolylneuraminic acid ( Neu5 Gc) and 3-keto-3-deoxy- d-glycero- d-galactonononic acid ( KDN), which are transported via the sialic acid transporter Nan T and catabolized using the sialic acid aldolase Nan A. Catabolism of Neu5 Gc uses the same pathway as Neu5 Ac, likely producing glycolate instead and acetate during its breakdown and catabolism of KDN requires Nan A activity, while other components of the Neu5 Ac catabolism pathway are non-essential. We also demonstrate that these two sialic acids can support growth of an E. coli ∆ nan T strain expressing sialic acid transporters from two bacterial pathogens, namely the tripartite ATP-independent periplasmic transporter Sia PQM from Haemophilus influenzae and the sodium solute symport transporter STM1128 from Salmonella enterica ssp. Typhimurium, suggesting that the ability to use Neu5 Gc and KDN in addition to Neu5 Ac is present in a number of human pathogens. [ABSTRACT FROM AUTHOR]

Host traits, such as migratory behavior, could facilitate the dispersal of disease-causing parasites, potentially leading to the transfer of infections both across geographic areas and between host species. There is, however, little quantitative information on whether variation in such host attributes does indeed affect the evolutionary outcome of host-parasite associations. Here, we employ Leucocytozoon blood parasites of birds, a group of parasites closely related to avian malaria, to study host-parasite coevolution in relation to host behavior using a phylogenetic comparative approach. We reconstruct the molecular phylogenies of both the hosts and parasites and use cophylogenetic tools to assess whether each host-parasite association contributes significantly to the overall congruence between the two phylogenies. We find evidence for a significant fit between host and parasite phylogenies in this system, but show that this is due only to associations between nonmigrant parasites and their hosts. We also show that migrant bird species harbor a greater genetic diversity of parasites compared with nonmigrant species. Taken together, these results suggest that the migratory habits of birds could influence their coevolutionary relationship with their parasites, and that consideration of host traits is important in predicting the outcome of coevolutionary interactions. [ABSTRACT FROM AUTHOR]

Extreme morphologies of many insular taxa suggest that islands have unusual properties that influence the tempo and mode of evolution. Yet whether insularity per se promotes rapid phenotypic evolution remains largely untested. We extend a phylogenetic comparative approach to test the influence of novel environments versus insularity on rates of body size and sexual size dimorphism diversification in Anolis. Rates of body size diversification among small-island and mainland species were similar to those of anole species on the Greater Antilles. However, the Greater Antilles taxa that colonized small islands and the mainland are ecologically nonrandom: rates of body size diversification among small-island and mainland species are high compared to their large-island sister taxa. Furthermore, rates of diversification in sexual size dimorphism on small islands are high compared to all large-island and mainland lineages. We suggest that elevated diversifying selection, particularly as a result of ecological release, may drive high rates of body size diversification in both small-island and mainland novel environments. In contrast, high abundance (prevalent among small-island lizard communities) mediating intraspecific resource competition and male–male competition may explain why sexual size dimorphism diversifies faster among small-island lineages than among their mainland and large-island relatives. [ABSTRACT FROM AUTHOR]

Background: Rhs genes are prominent features of bacterial genomes that have previously been implicated in genomic rearrangements in E. coli. By comparing rhs repertoires across the Enterobacteriaceae, this study provides a robust explanation of rhs diversification and evolution, and a mechanistic model of how rhs diversity is gained and lost. Results: Rhs genes are ubiquitous and comprise six structurally distinct lineages within the Enterobacteriaceae. There is considerable intergenomic variation in rhs repertoire; for instance, in Salmonella enterica, rhs are restricted to mobile elements, while in Escherichia coli one rhs lineage has diversified through transposition as older lineages have been deleted. Overall, comparative genomics reveals frequent, independent gene gains and losses, as well as occasional lateral gene transfer, in different genera. Furthermore, we demonstrate that Rhs 'core' domains and variable C-termini are evolutionarily decoupled, and propose that rhs diversity is driven by homologous recombination with circular intermediates. Existing C-termini are displaced by laterally acquired alternatives, creating long arrays of dissociated 'tips' that characterize the appearance of rhs loci. Conclusion: Rhs repertoires are highly dynamic among Enterobacterial genomes, due to repeated gene gains and losses. In contrast, the primary structures of Rhs genes are evolutionarily conserved, indicating that rhs sequence diversity is driven, not by rapid mutation, but by the relatively slow evolution of novel core/tip combinations. Hence, we predict that a large pool of dissociated rhs C-terminal tips exists episomally and these are potentially transmitted across taxonomic boundaries. [ABSTRACT FROM AUTHOR]

Tripartite ATP-independent periplasmic (TRAP) transporters are a family of extracytoplasmic solute receptor-dependent secondary transporters that are widespread in the prokaryotic world but which have not been extensively studied. Here, we present results of a genome-wide analysis of TRAP sequences and genome organization from application of TRAPDb, a relational database created for the collection, curation and analysis of TRAP sequences. This has revealed a specific enrichment in the number of TRAP transporters in several bacteria which is consistent with increased use of TRAP transporters in saline environments. Additionally, we report a number of new organizations of TRAP transporter genes and proteins which suggest the recruitment of TRAP transporter components for use in other biological contexts. Copyright © 2007 S. Karger AG, Basel [ABSTRACT FROM AUTHOR]

Campylobacter jejuniis a Gram-negative food-borne pathogen associated with gastroenteritis in humans as well as cases of the autoimmune disease Guillain–Barré syndrome.C. jejuniis asaccharolytic because it lacks an active glycolytic pathway for the use of sugars as a carbon source. This suggests an increased reliance on amino acids as nutrients and indeed the genome sequence of this organism indicates the presence of a number of amino acid uptake systems. Cj0982, also known as CjaA, is a putative extracytoplasmic solute receptor for one such uptake system as well as a major surface antigen and vaccine candidate. The crystal structure of Cj0982 reveals a two-domain protein with density in the enclosed cavity between the domains that clearly defines the presence of a bound cysteine ligand. Fluorescence titration experiments were used to demonstrate that Cj0982 binds cysteine tightly and specifically with aK d of∼10−7 M consistent with a role as a receptor for a high-affinity transporter. These data imply that Cj0982 is the binding protein component of an ABC-type cysteine transporter system and that cysteine uptake is important in the physiology ofC. jejuni. [ABSTRACT FROM AUTHOR]

Pseudaminic acid is a non-mammalian sugar found in the surface glycoconjugates of many bacteria, including several human pathogens, and is a virulence factor thought to facilitate immune evasion. The final step in the biosynthesis of the nucleotide activated form of the sugar, CMP-Pse5Ac7Ac is performed by a CMP-Pse5Ac7Ac synthetase (PseF). Here we present the biochemical and structural characterization of PseF from Aeromonas caviae (AcPseF), with AcPseF displaying metal-dependent activity over a broad pH and temperature range. Upon binding to CMP-Pse5Ac7Ac, AcPseF undergoes dynamic movements akin to other CMP-ulosonic acid synthetases. The enzyme clearly discriminates Pse5Ac7Ac from other ulosonic acids, through active site interactions with side-chain functional groups and by positioning the molecule in a hydrophobic pocket. Finally, we show that AcPseF binds the CMP-Pse5Ac7Ac side chain in the lowest energy conformation, a trend that we observed in the structures of other enzymes of this class. [Display omitted] • Crystal structures of AcPseF in the Apo and product-bound states were determined • AcPseF displays metal-dependent activity over a broad pH and temperature range • Pse5Ac7Ac is positioned within a hydrophobic pocket in the AcPseF active site • AcPseF binds the CMP-Pse5Ac7Ac side chain in the lowest energy tg conformation Keenan, Cowan et al. determined crystal structures of the CMP-Pse5Ac7Ac synthetase, AcPseF, in the ligand-free and product-bound forms. They provide molecular-level insights into how a Pse5Ac7Ac-processing enzyme might recognize and discriminate Pse5Ac7Ac from other ulosonic acids, which may inform the development of novel chemical tools or even antimicrobial inhibitors. [ABSTRACT FROM AUTHOR]

Tripartite ATP-independent periplasmic (TRAP) transporters are widespread in prokaryotes and are responsible for the transport of a variety of different ligands, primarily organic acids. TRAP transporters can be divided into two subclasses; DctP-type and TAXI type, which share the same overall architecture and substrate-binding protein requirement. DctP-type transporters are very well studied and have been shown to transport a range of compounds including dicarboxylates, keto acids, and sugar acids. However, TAXI-type transporters are relatively poorly understood. To address this gap in our understanding, we have structurally and biochemically characterized VC0430 from Vibrio cholerae. We show it is a monomeric, high affinity glutamate-binding protein, which we thus rename VcGluP. VcGluP is stereoselective, binding the L-isomer preferentially, and can also bind L-glutamine and L-pyroglutamate with lower affinity. Structural characterization of ligand-bound VcGluP revealed details of its binding site and biophysical characterization of binding site mutants revealed the substrate binding determinants, which differ substantially from those of DctP-type TRAPs. Finally, we have analyzed the interaction between VcGluP and its cognate membrane component, VcGluQM (formerly VC0429) in silico, revealing an architecture hitherto unseen. To our knowledge, this is the first transporter in V. cholerae to be identified as specific to glutamate, which plays a key role in the osmoadaptation of V. cholerae, making this transporter a potential therapeutic target. [ABSTRACT FROM AUTHOR]

The production of human body odor is the result of the action of commensal skin bacteria, including Staphylococcus hominis, acting to biotransform odorless apocrine gland secretions into volatile chemicals like thioalcohols such as 3-methyl-3-sulphanylhexan-1-ol (3M3SH). As the secreted odor precursor Cys-Gly-3M3SH contains a dipeptide, yet the final enzyme in the biotransformation pathway only functions on Cys-3M3SH, we sought to identify the remaining step in this human-adapted biochemical pathway using a novel coupled enzyme assay. Purification of this activity from S. hominis extracts led to the identification of the M20A-family PepV peptidase (ShPepV) as the primary Cys-Gly-3M3SH dipeptidase. To establish whether this was a primary substrate for PepV, the recombinant protein was purified and demonstrated broad activity against diverse dipeptides. The binding site for Cys-Gly-3M3SH was predicted using modeling, which suggested mutations that might accommodate this ligand more favorably. Indeed, a D437A resulted in an almost sixfold increase in the kcat/Km, whereas other introduced mutations reduced or abolished function. Together, these data identify an enzyme capable of catalyzing the missing step in an ancient human-specific biochemical transformation and suggest that the production of 3M3SH uses neither a dedicated transporter nor a peptidase for its breakdown, with only the final cleavage step, catalyzed by PatB cysteine-S-conjugate β-lyase, being a unique enzyme. [ABSTRACT FROM AUTHOR]

The tripartite ATP-independent periplasmic (TRAP) transporters enable Vibrio cholerae and Haemophilus influenzae to acquire sialic acid, aiding their colonization of human hosts. This process depends on SiaP, a substrate-binding protein (SBP) that captures and delivers sialic acid to the transporter. We identified 11 nanobodies that bind specifically to the SiaP proteins from H. influenzae (HiSiaP) and V. cholerae (VcSiaP). Two nanobodies inhibited sialic acid binding. Detailed structural and biophysical studies of one nanobody-SBP complex revealed an allosteric inhibition mechanism, preventing ligand binding and releasing pre-bound sialic acid. A hydrophobic surface pocket of the SBP is crucial for the allosteric mechanism and for the conformational rearrangement that occurs upon binding of sialic acid to the SBP. Our findings provide new clues regarding the mechanism of TRAP transporters, as well as potential starting points for novel drug design approaches to starve these human pathogens of important host-derived molecules. Biophysical and structural characterization of the TRAP transporter SBP SiaP in complex with a nanobody provides insights into an allosteric mechanism of inhibition. [ABSTRACT FROM AUTHOR]

An amendment to this paper has been published and can be accessed via a link at the top of the paper [ABSTRACT FROM AUTHOR]

Sexual selection is proposed to be a powerful driver of phenotypic evolution in animal systems. At macroevolutionary scales, sexual selection can theoretically drive both the rate and direction of phenotypic evolution, but this hypothesis remains contentious. Here, we find that differences in the rate and direction of plumage colour evolution are predicted by a proxy for sexual selection intensity (plumage dichromatism) in a large radiation of suboscine passerine birds (Tyrannida). We show that rates of plumage evolution are correlated between the sexes, but that sexual selection has a strong positive effect on male, but not female, interspecific divergence rates. Furthermore, we demonstrate that rapid male plumage divergence is biased towards carotenoid-based (red/yellow) colours widely assumed to represent honest sexual signals. Our results highlight the central role of sexual selection in driving avian colour divergence, and reveal the existence of convergent evolutionary responses of animal signalling traits under sexual selection. What factors explain variation in the pace and trajectory of evolutionary divergence between lineages? Here, the authors show that a proxy measure for sexual selection intensity predicts both the rate and direction of plumage colour evolution in a diverse radiation of New World passerine birds. [ABSTRACT FROM AUTHOR]

Progress towards the total synthesis of the macrolide natural product anthracimycin is described. This new approach utilises an intermolecular Diels-Alder strategy followed by epimeirsation to form the key trans-decalin framework. The route culminates in the stereoselective synthesis of an advanced tricyclic lactone intermediate, containing five contiguous sterogenic centres with the correct relative and absolute stereochemistry required for the anthracimycin core motif. [ABSTRACT FROM AUTHOR]

A review is presented of the article "A comprehensive phylogeny of birds (Aves) using targeted next-generation DNA sequencing" by Richard O. Prum, Jacob S. Berv, Alex Dornburg, and others concerning genome sequencing of 198 species of birds which allows determination of the early evolutionary relationships of modern avians.

Background: Worldwide 3.4 billion tonnes of municipal solid waste (MSW) will be produced annually by 2050, however, current approaches to MSW management predominantly involve unsustainable practices like landfilling and incineration. The organic fraction of MSW (OMSW) typically comprises ~ 50% lignocellulose-rich material but is underexplored as a biomanufacturing feedstock due to its highly inconsistent and heterogeneous composition. This study sought to overcome the limitations associated with studying MSW-derived feedstocks by using OMSW produced from a realistic and reproducible MSW mixture on a commercial autoclave system. The resulting OMSW fibre was enzymatically hydrolysed and used to screen diverse microorganisms of biotechnological interest to identify robust species capable of fermenting this complex feedstock. Results: The autoclave pre-treated OMSW fibre contained a polysaccharide fraction comprising 38% cellulose and 4% hemicellulose. Enzymatic hydrolysate of OMSW fibre was high in d-glucose (5.5% w/v) and d-xylose (1.8%w/v) but deficient in nitrogen and phosphate. Although relatively low levels of levulinic acid (30 mM) and vanillin (2 mM) were detected and furfural and 5-hydroxymethylfurfural were absent, the hydrolysate contained an abundance of potentially toxic metals (0.6% w/v). Hydrolysate supplemented with 1% yeast extract to alleviate nutrient limitation was used in a substrate-oriented shake-flask screen with eight biotechnologically useful microorganisms (Clostridium saccharoperbutylacetonicum, Escherichia coli, Geobacillus thermoglucosidasius, Pseudomonas putida, Rhodococcus opacus, Saccharomyces cerevisiae, Schizosaccharomyces pombe and Zymomonas mobilis). Each species' growth and productivity were characterised and three species were identified that robustly and efficiently fermented OMSW fibre hydrolysate without significant substrate inhibition: Z. mobilis, S. cerevisiae and R. opacus, respectively produced product to 69%, 70% and 72% of the maximum theoretical fermentation yield and could theoretically produce 136 kg and 139 kg of ethanol and 91 kg of triacylglycerol (TAG) per tonne of OMSW. Conclusions: Developing an integrated biorefinery around MSW has the potential to significantly alleviate the environmental burden of current waste management practices. Substrate-oriented screening of a representative and reproducible OMSW-derived fibre identified microorganisms intrinsically suited to growth on OMSW hydrolysates. These species are promising candidates for developing an MSW biorefining platform and provide a foundation for future studies aiming to valorise this underexplored feedstock. [ABSTRACT FROM AUTHOR]

Characterising RNA–protein interaction dynamics is fundamental to understand how bacteria respond to their environment. In this study, we have analysed the dynamics of 91% of the Escherichia coli expressed proteome and the RNA-interaction properties of 271 RNA-binding proteins (RBPs) at different growth phases. We find that 68% of RBPs differentially bind RNA across growth phases and characterise 17 previously unannotated proteins as bacterial RBPs including YfiF, a ncRNA-binding protein. While these new RBPs are mostly present in Proteobacteria, two of them are orthologs of human mitochondrial proteins associated with rare metabolic disorders. Moreover, we reveal novel RBP functions for proteins such as the chaperone HtpG, a new stationary phase tRNA-binding protein. For the first time, the dynamics of the bacterial RBPome have been interrogated, showcasing how this approach can reveal the function of uncharacterised proteins and identify critical RNA–protein interactions for cell growth which could inform new antimicrobial therapies. Synopsis: A dynamic analysis of RNA-protein interaction rewiring across growth phases detects extensive reorganisation of the RBPome and reveals the RNA binding properties for 17 unannotated E. coli proteins and their differential impact on cell growth and evolutionary conservation. Mass spectrometry characterisation of the dynamics of 91% of the expressed E. coli proteome highlights extensive variation during cell growth. Determining RNA-protein interaction dynamics reveals that 68% of the RBPome differentially binds RNA according to the bacterial growth stage. iCLIP analysis of YfiF and HtpG unveils novel roles for these two proteins as ncRNA binders. A dynamic analysis of RNA-protein interaction rewiring across growth phases detects extensive reorganisation of the RBPome and reveals the RNA binding properties for 17 unannotated E. coli proteins and their differential impact on cell growth and evolutionary conservation. [ABSTRACT FROM AUTHOR]

Cell surface sugar 5,7‐diacetyl pseudaminic acid (Pse5Ac7Ac) is a bacterial analogue of the ubiquitous sialic acid, Neu5Ac, and contributes to the virulence of a number of multidrug resistant bacteria, including ESKAPE pathogens Pseudomonas aeruginosa, and Acinetobacter baumannii. Despite its discovery in the surface glycans of bacteria over thirty years ago, to date no glycosyltransferase enzymes (GTs) dedicated to the synthesis of a pseudaminic acid glycosidic linkage have been unequivocally characterised in vitro. Herein we demonstrate that A. baumannii KpsS1 is a dedicated pseudaminyltransferase enzyme (PseT) which constructs a Pse5Ac7Ac‐α(2,6)‐Glcp linkage, and proceeds with retention of anomeric configuration. We utilise this PseT activity in tandem with the biosynthetic enzymes required for CMP‐Pse5Ac7Ac assembly, in a two‐pot, seven enzyme synthesis of an α‐linked Pse5Ac7Ac glycoside. Due to its unique activity and protein sequence, we also assign KpsS1 as the prototypical member of a previously unreported GT family (GT118). [ABSTRACT FROM AUTHOR]

This corrects the article DOI: 10.1038/nature21074 [ABSTRACT FROM AUTHOR]

The formation of complex multidomain proteins has occurred many times during evolution. For enzymes, this often confers the addition of new functions or new routes to regulate function. Herein we review how this same process has impacted on the function of membrane transporters and channels, proteins that, due to their integral membrane location, are potentially more constrained in the fusions they can accommodate. Using examples primarily from bacterial systems, we illustrate diverse instances of functional fusions and find evidence for promiscuous fusion partners that have fused to many different classes of membrane protein. We consider the evidence that topology and stoichiometry issues might limit the range of fusions that are selected in nature and attempt to find examples where a functional benefit of direct fusion over split proteins has been demonstrated. Finally, we consider whether the reverse process of gene fission has been important in membrane transporter evolution. [ABSTRACT FROM AUTHOR]

Widely documented, megaevolutionary jumps in phenotypic diversity continue to perplex researchers because it remains unclear whether these marked changes can emerge from microevolutionary processes. Here, we tackle this question using new approaches for modeling multivariate traits to evaluate the magnitude and distribution of elaboration and innovation in the evolution of bird beaks. We find that elaboration, evolution along the major axis of phenotypic change, is common at both macro- and megaevolutionary scales, whereas innovation, evolution away from the major axis of phenotypic change, is more prominent at megaevolutionary scales. The major axis of phenotypic change among species beak shapes at megaevolutionary scales is an emergent property of innovation across clades. Our analyses suggest that the reorientation of phenotypes via innovation is a ubiquitous route for divergence that can arise through gradual change alone, opening up further avenues for evolution to explore. [ABSTRACT FROM AUTHOR]

What drives the evolution of plumage color in birds? Bird color is likely to be under both natural and sexual selection where natural selection may favor evolution toward crypsis or camouflage whereas sexual selection may favor evolution toward conspicuousness. The responses to selection are predicted to relate to species' ecology, behavior, and life history. Key hypotheses have focused on habitat and light environment, breeding strategy, territoriality, and hunting behavior. We tested these potential causes of color variation in the Coraciiformes, a colorful clade of non-passerine birds, using phylogenetic comparative methods and data on chromatic and achromatic properties of plumage coloration measured from museum specimens. We found that correlates of color evolution in Coraciiformes vary across body regions and depend on the focal color property (chromatic or achromatic properties of plumage coloration). While the light environment showed widespread effects on coloration in multiple body regions for both color properties, selection pressures related to behavioral characteristics had more spatially localized effects (e.g. territoriality on achromatic properties of wing feathers and hunting strategy on chromatic properties of belly feathers). Our results reveal both general patterns that may hold across other bird clades and more nuanced effects of selection that are likely to be mediated through the visual ecology of the signaler and receiver and the behavioral characteristics of Coraciiform species. [ABSTRACT FROM AUTHOR]

A key challenge in mobilising growing numbers of digitised biological specimens for scientific research is finding high-throughput methods to extract phenotypic measurements on these datasets. In this paper, we test a pose estimation approach based on Deep Learning capable of accurately placing point labels to identify key locations on specimen images. We then apply the approach to two distinct challenges that each requires identification of key features in a 2D image: (i) identifying body region-specific plumage colouration on avian specimens and (ii) measuring morphometric shape variation in Littorina snail shells. For the avian dataset, 95% of images are correctly labelled and colour measurements derived from these predicted points are highly correlated with human-based measurements. For the Littorina dataset, more than 95% of landmarks were accurately placed relative to expert-labelled landmarks and predicted landmarks reliably captured shape variation between two distinct shell ecotypes ('crab' vs 'wave'). Overall, our study shows that pose estimation based on Deep Learning can generate high-quality and high-throughput point-based measurements for digitised image-based biodiversity datasets and could mark a step change in the mobilisation of such data. We also provide general guidelines for using pose estimation methods on large-scale biological datasets. Author summary: As the digitisation of natural history collections continues apace, a wealth of information is waiting to be mobilised from these vast digital datasets that can help address many evolutionary and ecological questions. Deep Learning has achieved success on many real-world tasks such as face recognition and image classification. Here, we use deep learning to measure phenotypic traits of specimens by placing points on photos of birds and periwinkles. We show that the measurements produced by Deep Learning are generally accurate and very similar to manual measurements taken by experts. As Deep Learning methods vastly reduce the time required to produce these measurements, our results demonstrate the great potential of Deep Learning for future biodiversity studies. [ABSTRACT FROM AUTHOR]

Due to rising resistance, new antibacterial strategies are needed, including methods for targeted antibiotic release. As targeting vectors, chelating molecules called siderophores that are released by bacteria to acquire iron have been investigated for conjugation to antibacterials, leading to the clinically approved drug cefiderocol. The use of small‐molecule catalysts for prodrug activation within cells has shown promise in recent years, and here we investigate siderophore‐linked ruthenium catalysts for the activation of antibacterial prodrugs within cells. Moxifloxacin‐based prodrugs were synthesised, and their catalyst‐mediated activation was demonstrated under anaerobic, biologically relevant conditions. In the absence of catalyst, decreased antibacterial activities were observed compared to moxifloxacin versus Escherichia coli K12 (BW25113). A series of siderophore‐linked ruthenium catalysts were investigated for prodrug activation, all of which displayed a combinative antibacterial effect with the prodrug, whereas a representative example displayed little toxicity against mammalian cell lines. By employing complementary bacterial growth assays, conjugates containing siderophore units based on catechol and azotochelin were found to be most promising for intracellular prodrug activation. [ABSTRACT FROM AUTHOR]

Acylation of diverse carbohydrates occurs across all domains of life and can be catalysed by proteins with a membrane bound acyltransferase-3 (AT3) domain (PF01757). In bacteria, these proteins are essential in processes including symbiosis, resistance to viruses and antimicrobials, and biosynthesis of antibiotics, yet their structure and mechanism are largely unknown. In this study, evolutionary co-variance analysis was used to build a computational model of the structure of a bacterial O-antigen modifying acetyltransferase, OafB. The resulting structure exhibited a novel fold for the AT3 domain, which molecular dynamics simulations demonstrated is stable in the membrane. The AT3 domain contains 10 transmembrane helices arranged to form a large cytoplasmic cavity lined by residues known to be essential for function. Further molecular dynamics simulations support a model where the acyl-coA donor spans the membrane through accessing a pore created by movement of an important loop capping the inner cavity, enabling OafB to present the acetyl group close to the likely catalytic resides on the extracytoplasmic surface. Limited but important interactions with the fused SGNH domain in OafB are identified, and modelling suggests this domain is mobile and can both accept acyl- groups from the AT3 and then reach beyond the membrane to reach acceptor substrates. Together this new general model of AT3 function provides a framework for the development of inhibitors that could abrogate critical functions of bacterial pathogens. [ABSTRACT FROM AUTHOR]

Biodiversity is facing a global extinction crisis that will reduce ecological trait diversity, evolutionary history, and ultimately ecosystem functioning and services. 1–4 A key challenge is understanding how species losses will impact morphological and phylogenetic diversity at global scales. 5,6 Here, we test whether the loss of species threatened with extinction according to the International Union for Conservation of Nature (IUCN) leads to morphological and phylogenetic homogenization 7,8 across both the whole avian class and within each biome and ecoregion globally. We use a comprehensive set of continuous morphological traits extracted from museum collections of 8,455 bird species, including geometric morphometric beak shape data, 9 and sequentially remove species from those at most to least threat of extinction. We find evidence of morphological, but not phylogenetic, homogenization across the avian class, with species becoming more alike in terms of their morphology. We find that most biome and ecoregions are expected to lose morphological diversity at a greater rate than predicted by species loss alone, with the most imperiled regions found in East Asia and the Himalayan uplands and foothills. Only a small proportion of assemblages are threatened with phylogenetic homogenization, in particular parts of Indochina. Species extinctions will lead to a major loss of avian ecological strategies, but not a comparable loss of phylogenetic diversity. As the decline of species with unique traits and their replacement with more widespread generalist species continues, the protection of assemblages at most risk of morphological and phylogenetic homogenization should be a key conservation priority. • Predicted loss of birds will drive exceptional declines in morphological diversity • Species extinctions lead to a major loss of ecological strategies and functions • Most biomes and ecoregions will experience morphological homogenization • Phylogenetic diversity tends to decline as expected as species go extinct The global extinction crisis will lead to widespread losses of morphological diversity. Hughes et al. show that predicted species extinctions drive far greater declines of ecological strategies than predicted, with important ramifications for humans as ecosystem services are lost. In contrast, phylogenetic diversity declines as expected. [ABSTRACT FROM AUTHOR]

A novel Trojan Horse conjugate consisting of an SO 2 -releasing 2,4-dinitrobenzenesulfonamide group attached to the monocatecholate siderophore aminochelin was synthesized to examine whether a bidentate catecholate siderophore unit could help potentiate the antimicrobial activity of SO 2 -releasing prodrugs. The conjugate obtained displays rapid SO 2 release on reaction with glutathione, and proved more active against Staphylococcus aureus than a comparable SO 2 -releasing prodrug lacking the siderophore unit, although activity required micromolar concentrations. The conjugate was inactive against wild-type Escherichia coli , but activity was observed against an entA mutant strain that is unable to produce its major siderophores. Hence, the poor activity of the conjugate in wild-type E. coli may be due to the production of native siderophores that can compete with the conjugate for iron binding and uptake. Trojan Horse approach: the attachment of an iron-binding didentate catecholate siderophore unit potentiates the antimicrobial activity of an SO 2 -releasing prodrug in S. aureus. [Display omitted] • Synthesis of siderophore-based 2,4-dinitrobenzenesulfonamide conjugate and 3 analogues. • Rapid SO 2 release from the siderophore conjugate upon reaction with glutathione. • Siderophore-enhanced antibacterial activity against Staphylococcus aureus. • The conjugate was inactive against wild-type Escherichia coli. • Activity was observed against E. coli mutant unable to produce own siderophores. [ABSTRACT FROM AUTHOR]

Robust systematic approaches for the metabolic engineering of cell factories remain elusive. The available models for predicting phenotypical responses and mechanisms are incomplete, particularly within the context of compound toxicity that can be a significant impediment to achieving high yields of a target product. This study describes a Multi-Omic Based Production Strain Improvement (MOB psi) strategy that is distinguished by integrated time-resolved systems analyses of fed-batch fermentations. As a case study, MOB psi was applied to improve the performance of an Escherichia coli cell factory producing the commodity chemical styrene. Styrene can be bio-manufactured from phenylalanine via an engineered pathway comprised of the enzymes phenylalanine ammonia lyase and ferulic acid decarboxylase. The toxicity, hydrophobicity, and volatility of styrene combine to make bio-production challenging. Previous attempts to create styrene tolerant E. coli strains by targeted genetic interventions have met with modest success. Application of MOB psi identified new potential targets for improving performance, resulting in two host strains (E. coli NST74Δ aaeA and NST74Δ aaeA cpxP o) with increased styrene production. The best performing re-engineered chassis, NST74Δ aaeA cpxP o , produced ∼3 × more styrene and exhibited increased viability in fed-batch fermentations. Thus, this case study demonstrates the utility of MOB psi as a systematic tool for improving the bio-manufacturing of toxic chemicals. • Product toxicity impairs the performance of cell factories and limits yields. • Systematic approaches to improve strains producing toxic chemicals are few. • MOB psi integrates time-resolved multi-omic and analytic data from production runs. • The power of MOB psi was tested with an E. coli cell factory producing styrene. • New genetic interventions were implemented that enhanced styrene production. [ABSTRACT FROM AUTHOR]

The article discusses how sialic acid is utilized by bacterial pathogens. Sialic acid is important in various cellular processes such as intercellular adhesion and cell signaling. Pathogenic bacteria use sialic acid in two ways. First, they can coat themselves in sialic acid to provide resistance to components of the host's innate immune response. Second, bacteria can use sialic acid as a nutrient. Bacteria acquire sialic acid either by scavenging from the environment or by de novo biosynthesis. Bacteria can also modify sialic acid by O-acetylation.