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The modern lithium-ion battery (LIB) configuration was enabled by the “magic chemistry” between ethylene carbonate (EC) and graphitic carbon anode. Despite the constant changes of cathode chemistries with improved energy densities, EC-graphite combination remained static during the last three decades. While the interphase generated by EC protects the fragile graphitic structure, the intrinsic disadvantages of EC (high viscosity, high melting point, excessive interphase growth) lead to mediocre power density and poor performances of LIB at sub-zero temperatures, where lithium depositions form upon charging. Such performance compromises arise from the fundamental dilemma between requiring effective interphase protection and high impedance from excessive growth of interphase. In this work, we designed and synthesized a “double EC” molecule as electrolyte additive to resolve the above dilemma. Erythritol bis(carbonate) (EBC) possesses lower LUMO energy level than EC and hence tends to decompose prior to EC reduction, but its weak solvation toward Li+ restricts the extent of its reduction, thus minimizing the interphase thickness and the corresponding impedances. Electrolytes containing EBC enables both the charging and discharging of ampere-size LIB pouch cells at sub-zero temperatures from 0 to -20℃, demonstrating that the key approach to improve low temperature performances lies in how to tailor interphasial chemistry rather than the bulk electrolyte composition.

Developing structure-performance relationships with the underlying goal of optimizing known zeolite catalysts involves the manipulation of their physicochemical properties. Here, we systematically assessed the impact of mesoscopic gradients in acid site concentration, which has received little attention in the design of zeolite catalysts for hydrocarbon upgrading. A series of core-shell MEL-type zeolites were synthesized with catalytically active ZSM-11 cores and passivated silicalite-2 shells of varying thickness. Our findings revealed that ZSM-11@silicalite-2 particles with ultrathin shells (

Peptides have potential to be developed into immune checkpoint inhibitors, but the target interfaces are difficult to inhibit. Here, we explored an approach to mimic the binding surface of PD-1 to design inhibitors. Mimicking native PD-1 resulted in a mimetic with no activity. However, mimicking an affinity-optimized PD-1 resulted in the peptide mimetic MOPD-1 that displayed nanomolar affinity to PD-L1 and could inhibit PD-1:PD-L1 interactions in both protein- and cell-based assays. Mutagenesis and structural characterization using NMR spectroscopy and X-ray crystallography revealed that binding residues from the high affinity PD-1 are crucial for the bioactivity of MOPD-1. Furthermore, MOPD-1 was extremely stable in human serum and inhibited tumor growth in vivo, suggesting it has potential for use in cancer immunotherapy. The successful design of an inhibitor of PD-1:PD-L1 using the mimicry approach described herein illustrates the value of placing greater emphasis on optimizing the target interface before inhibitor design and is an approach that could have broader utility for the design of peptide inhibitors for other complex protein-protein interactions.

Proprotein convertase subtilisin/kexin type 9 (PCSK9) is a clinically validated target for treating cardiovascular disease (CVD) due to its involvement in cholesterol metabolism. Although approved monoclonal antibodies (alirocumab and evolocumab) that inhibit PCSK9 function are very effective in lowering cholesterol, their limitations, including high treatment costs, have so far prohibited widespread use. Accordingly, there is great interest in alternative drug modalities to antibodies. Like antibodies, peptides are valuable therapeutics due to their high target potency and specificity. Furthermore, being smaller than antibodies means they have access to more drug administration options, are less likely to induce adverse immunogenic responses, and are better suited to affordable production. This review surveys the current peptide-based landscape aimed towards PCSK9 inhibition, covering pre-clinical to patented drug candidates and comparing them to current cholesterol lowering therapeutics. Classes of peptides reported to be inhibitors include nature-inspired disulfide-rich peptides, combinatorially derived cyclic peptides, and peptidomimetics. Their functional activities have been validated in biophysical and cellular assays, and in some cases pre-clinical mouse models. Recent efforts report peptides with potent sub-nanomolar binding affinities to PCSK9, which highlights their potential to achieve antibody-like potency. Studies are beginning to address pharmacokinetic properties of PCSK9-targeting peptides in more detail. We conclude by highlighting opportunities to investigate their biological effects in pre-clinical models of cardiovascular disease. The anticipation concerning the PCSK9-targeting peptide landscape is accelerating and it seems likely that a peptide-based therapeutic for treating PCSK9-mediated hypercholesterolemia may be clinically available in the near future.

International audience; Two new mononuclear Fe(II) polymorphs, [(C2H5)4N]2[Fe(py3C-OEt)(NCS)3]2 (1) and [(C2H5)4N][Fe(py3C-OEt)(NCS)3] (2) (py3C-OEt = tris(pyridin-2-yl)ethoxymethane), have been synthesized and characterized by single-crystal X-ray diffraction, by magnetic and photomagnetic measurements, and by detailed variable-temperature infrared spectroscopy. The molecular structure, in both complexes, is composed of the same anionic [Fe(py3C-OEt)(NCS)3]− complex (two units for 1 and one unit for 2) generated by coordination to the Fe(II) metal center of one tridentate py3C-OEt tripodal ligand and three terminal κN-SCN coligands. Magnetic studies revealed that polymorph 2 displays a high-spin (HS) state over the entire studied temperature range (300–10 K), while complex 1 exhibits an abrupt and complete spin crossover (SCO) transition at ca. 132.3 K, the structural characterizations of which, performed at 295 and 100 K, show a strong modification, resulting from the thermal evolutions of the Fe–N bond lengths and of the distortion parameters (∑ and Θ) of the FeN6 coordination sphere, in agreement with the presence of HS and low-spin (LS) states at 295 and 100 K, respectively. This thermal transition has been also confirmed by the thermal evolution of the maximum absorbance for ν(NCS) vibrational bands recorded in the temperature range 200–10 K. In 1 the signature of a metastable photoinduced HS state has been observed using photomagnetic and photoinfrared spectroscopy, leading to a similar T(LIESST) relaxation temperature (LIESST = light-induced excited spin-state trapping) of 70 K.

Cyclic disulfide-rich peptides have attracted significant interest in drug development and biotechnology. Here, we describe a protocol for producing cyclic peptide precursors in Pichia pastoris that undergo in vitro enzymatic maturation into cyclic peptides using recombinant asparaginyl endopeptidases (AEPs). Peptide precursors are expressed with a C-terminal His tag and secreted into the media, enabling facile purification by immobilized metal affinity chromatography. After AEP-mediated cyclization, cyclic peptides are purified by reverse-phase high-performance liquid chromatography and characterized by mass spectrometry, peptide mass fingerprinting, NMR spectroscopy, and activity assays. We demonstrate the broad applicability of this protocol by generating cyclic peptides from three distinct classes that are either naturally occurring or synthetically backbone cyclized, and range in size from 14 amino acids with one disulfide bond, to 34 amino acids with a cystine knot comprising three disulfide bonds. The protocol requires 14 d to identify and optimize a high-expressing Pichia clone in small-scale cultures (24 well plates or 50 mL tubes), after which large-scale production in a bioreactor and peptide purification can be completed in 10 d. We use the cyclotide Momordica cochinchinensis trypsin inhibitor II as an example. We also include a protocol for recombinant AEP production in Escherichia coli as AEPs are emerging tools for orthogonal peptide and protein ligation. We focus on two AEPs that preferentially cyclize different peptide precursors, namely an engineered AEP with improved catalytic efficiency [C247A]OaAEP1b and the plant-derived MCoAEP2. Rudimentary proficiency and equipment in molecular biology, protein biochemistry and analytical chemistry are needed. The authors present a protocol for producing cyclic peptide precursors in Pichia pastoris that undergo in vitro enzymatic maturation into cyclic peptides using recombinant asparaginyl endopeptidases. Assays for characterizing purified cyclic peptides are also described.

Developing vapor phase infiltration (VPI) processes for area selective polymer nanopatterning requires substantial advancement in understanding precursor infiltration, precursor–polymer interaction and process parameters. In this work, infiltration receptive poly(2-vinylpyridine) (P2VP) and poly(4-vinylpyridine) (P4VP) brushes were exposed to a trimethylaluminum (TMA) VPI process and compared to a non-receptive polystyrene (PS) system. The interaction that takes place between TMA and P2VP/P4VP was analysed in detail and we report on notable advantages in the use of P4VP, arising from the difference in position of the pyridinic nitrogen. The VPI process was performed in a commercial atomic layer deposition reactor and the effects of the fundamental process parameters on the three polymer brushes were investigated to ensure optimal area selectivity. In situ X-ray photoelectron spectroscopy (XPS) measurements were supported by grazing angle Fourier transform infrared spectroscopy (GA-FTIR) and hard X-ray photoelectron spectroscopy (HAXPES). The report identifies several important factors when developing a VPI process to ensure area selectivity, while also demonstrating the use of novel pyridine containing polymers for VPI area selective purposes.

Peptides are regarded as promising next-generation therapeutics. However, an analysis of over 1000 bioactive peptide candidates suggests that many have underdeveloped affinities and could benefit from cyclization using a bridging linker sequence. Until now, the primary focus has been on the use of inert peptide linkers. Here, we show that affinity can be significantly improved by enriching the linker with functional amino acids. We engineered a peptide inhibitor of PCSK9, a target for clinical management of hypercholesterolemia, to demonstrate this concept. Cyclization linker optimization from library screening produced a cyclic peptide with ∼100-fold improved activity over the parent peptide and efficiently restored low-density lipoprotein (LDL) receptor levels and cleared extracellular LDL. The linker forms favorable interactions with PCSK9 as evidenced by thermodynamics, structure-activity relationship (SAR), NMR, and molecular dynamics (MD) studies. This PCSK9 inhibitor is one of many peptides that could benefit from bioactive cyclization, a strategy that is amenable to broad application in pharmaceutical design.

Purpose:Mutational activation of GNAQ or GNA11 (GNAQ/11), detected in >90% of uveal melanomas, leads to constitutive activation of oncogenic pathways, including MAPK and YAP. To date, chemo- or pathway-targeted therapies, either alone or in combination, have proven ineffective in the treatment of patients with metastatic uveal melanoma.Experimental Design:We tested the efficacy of chloroquine or hydroxychloroquine, in combination with MAPK pathway inhibition in GNAQ/11-mutated cells in vitro and in vivo and identified mechanisms of MEK1/2 inhibitor plus chloroquine-induced cytotoxicity.Results:Inhibition of GNAQ/11-mediated activation of MAPK signaling resulted in the induction of autophagy. Combined inhibition of Gα and autophagy or lysosome function resulted in enhanced cell death. Moreover, the combination of MEK1/2 inhibition, using trametinib, with the lysosome inhibitor, chloroquine, also increased cytotoxicity. Treatment of mice bearing GNAQ/11-driven melanomas with trametinib plus hydroxychloroquine resulted in inhibition of tumor growth and significantly prolonged survival. Interestingly, lysosomal- and autophagy-specific inhibition with bafilomycin A1 was not sufficient to promote cytotoxicity in combination with trametinib. However, the addition of YAP inhibition with trametinib plus bafilomycin A1 resulted in cell death at comparable levels to trametinib plus chloroquine (T/CQ) treatment. Furthermore, T/CQ-treated cells displayed decreased YAP nuclear localization and decreased YAP transcriptional activity. Expression of a constitutively active YAP5SA mutant conferred resistance to T/CQ-induced cell death.Conclusions:These results suggest that YAP, MEK1/2, and lysosome function are necessary and critical targets for the therapy of GNAQ/11-driven melanoma, and identify trametinib plus hydroxychloroquine as a potential treatment strategy for metastatic uveal melanoma.

An energy storage paradigm orthogonal to Li-ion battery chemistries can be conceptualized by employing anions as the primary charge carriers. F-ion conversion chemistries show promise but have limi...

Ruthenium-catalysed azide-alkyne cycloaddition (RuAAC) provides access to 1,5-disubstituted 1,2,3-triazole motifs in peptide engineering applications. However, investigation of this motif as a disulfide mimetic in cyclic peptides has been limited, and the structural consequences remain to be studied. We report synthetic strategies to install various triazole linkages into cyclic peptides through backbone cyclisation and RuAAC cross-linking reactions. These linkages were evaluated in four serine protease inhibitors based on sunflower trypsin inhibitor-1. NMR and X-ray crystallography revealed exceptional consensus of bridging distance and backbone conformations (RMSD

Disulfide bonds typically introduce conformational constraints into peptides and proteins, conferring improved biopharmaceutical properties and greater therapeutic potential. In our opinion, disulfide-rich microdomains from proteins are potentially a rich and under-explored source of drug leads. A survey of the UniProt protein database shows that these domains are widely distributed throughout the plant and animal kingdoms, with the EGF-like domain being the most abundant of these domains. EGF-like domains exhibit large diversity in their disulfide bond topologies and calcium binding modes, which we classify in detail here. We found that many EGF-like domains are associated with disease phenotypes, and the interactions they mediate are potential therapeutic targets. Indeed, EGF-based therapeutic leads have been identified, and we further propose that these domains can be optimized to expand their therapeutic potential using chemical design strategies. This Review highlights the potential of disulfide-rich microdomains as future peptide therapeutics.

Macrocyclic, disulfide-rich peptides have found widespread applications in drug design and development. Current peptide production strategies rely heavily on solid phase peptide synthesis (SPPS) requiring large amounts of hazardous/toxic reagents and solvents which have negative environmental impacts. A possible solution is to develop a sustainable hybrid production platform incorporating recombinant production of cyclic peptide precursors in yeast followed by enzymatic maturation of these precursors into cyclic peptides using asparaginyl endopeptidases in vitro. Harnessing the efficient secretory pathway of Pichia pastoris, peptide precursors, cloned downstream of the α-mating factor secretion signal, were purified from culture supernatant mitigating the need for complex purification. To demonstrate the broad utility of the platform, three distinct classes of cyclic peptides were produced; two were structurally validated by NMR and shown to be functionally equivalent to their synthetically produced versions. Furthermore, using this platform we report the first recombinant production of any α-conotoxin in its native “globular” conformation. Using scale-up production in bioreactors, cyclic peptide yields of 85–97 mg L−1 of culture were achieved, far exceeding the highest yields so far achieved for cyclic disulfide-rich peptides in any recombinant process. This platform can potentially unlock production and facilitate applications of cyclic disulfide-rich peptides previously inaccessible through large-scale chemical synthesis and reduce their environmental burden.

The insecticidal effects of ω-hexatoxin-Hv1a, κ-hexatoxin-Hv1c and ω/κ-hexatoxin-Hv1h are currently attributed to action at calcium and potassium channels. By characterizing the binding of these toxins to neuronal membranes, we show that they have more potent effects as positive allosteric modulators (PAMs) of insect nicotinic acetylcholine receptors (nAChRs), consistent with their neuroexcitatory toxicology. Alanine scanning analysis of ω-hexatoxin-Hv1a reveals a structure-activity relationship for binding that mirrors that for insecticidal activity. Spinosyn A does not compete with ω-hexatoxin-Hv-1a for binding, and we show that these two PAMs have distinct pharmacology of binding indicating that they act at different receptor populations. These toxins represent valuable tools for the characterization of insect nAChRs and for the development of more selective agrochemicals.

Loops at protein-protein interfaces are a rich source of peptide leads that have high specificity and low toxicity. Although such peptides typically need to be constrained to overcome thermodynamic and metabolic limitations, design guidelines to obtain a successfully constrained peptides, and thus facilitate the transition from loop to drug, are relatively poorly formulated. In this work, we surveyed the structures of interface loops and found the position of the terminal residues to be a key determinant of conformation. We used this knowledge to improve the process of molecular grafting, a valuable approach for constraining and stabilising peptides by fusing them to a suitable scaffold. We show that an informed choice of where a loop is "anchored" to a scaffold improves its form and function. This knowledge can help guide the choice of loop and its matching scaffold, and thus increase the success rate for designing stable and potent peptide drug leads.

Cyclotides have attracted great interest as drug design scaffolds because of their unique cyclic cystine knotted topology. They are classified into three subfamilies, among which the bracelet subfamily represents the majority and comprises the most bioactive cyclotides, but are the most poorly utilized in drug design applications. A long-standing challenge has been the very low in vitro folding yields of bracelets, hampering efforts to characterize their structures and activities. Herein, we report substantial increases in bracelet folding yields enabled by a single point mutation of residue Ile-11 to Leu or Gly. We applied this discovery to synthesize mirror image enantiomers and used quasi-racemic crystallography to elucidate the first crystal structures of bracelet cyclotides. This study provides a facile strategy to produce bracelet cyclotides, leading to a general method to easily access their atomic resolution structures and providing a basis for development of biotechnological applications.

Despite the effectiveness of direct-acting antiviral agents in treating hepatitis C virus (HCV), cases of treatment failure have been associated with the emergence of resistance-associated substitutions. To better guide clinical decision-making, we developed and validated a near-whole-genome HCV genotype-independent next-generation sequencing strategy. HCV genotype 1–6 samples from direct-acting antiviral agent treatment-naïve and -treated HCV-infected individuals were included. Viral RNA was extracted using a NucliSens easyMAG and amplified using nested reverse transcription-polymerase chain reaction. Libraries were prepared using Nextera XT and sequenced on the Illumina MiSeq sequencing platform. Data were processed by an in-house pipeline (MiCall). Nucleotide consensus sequences were aligned to reference strain sequences for resistance-associated substitution identification and compared to NS3, NS5a, and NS5b sequence data obtained from a validated in-house assay optimized for HCV genotype 1. Sequencing success rates (defined as achieving &gt, 100-fold read coverage) approaching 90% were observed for most genotypes in samples with a viral load &gt, 5 log10 IU/mL. This genotype-independent sequencing method resulted in &gt, 99.8% nucleotide concordance with the genotype 1-optimized method, and 100% agreement in genotype assignment with paired line probe assay-based genotypes. The assay demonstrated high intra-run repeatability and inter-run reproducibility at detecting substitutions above 2% prevalence. This study highlights the performance of a freely available laboratory and bioinformatic approach for reliable HCV genotyping and resistance-associated substitution detection regardless of genotype.

Angiotensinogen fine-tunes the tightly controlled activity of the renin-angiotensin system by modulating the release of angiotensin peptides that control blood pressure. One mechanism by which this modulation is achieved is via angiotensinogen's Cys18–Cys138 disulfide bond that acts as a redox switch. Molecular dynamics simulations of each redox state of angiotensinogen reveal subtle dynamic differences between the reduced and oxidised forms, particularly at the N-terminus. Surface plasmon resonance data demonstrate that the two redox forms of angiotensinogen display different binding kinetics to an immobilised anti-angiotensinogen monoclonal antibody. Mass spectrometry mapped the epitope for the antibody to the N-terminal region of angiotensinogen. We therefore provide evidence that the different redox forms of angiotensinogen can be detected by an antibody-based detection method.

Despite mounting evidence that materials imperfections are a major obstacle to practical applications of superconducting qubits, connections between microscopic material properties and qubit coherence are poorly understood. Here, we combine measurements of transmon qubit relaxation times (T1) with spectroscopy and microscopy of the polycrystalline niobium films used in qubit fabrication. By comparing films deposited using three different techniques, we reveal correlations between T1 and intrinsic film properties such as grain size, enhanced oxygen diffusion along grain boundaries, and the concentration of suboxides near the surface. Qubit and resonator measurements show signatures of two-level system defects, which we propose to be hosted in the grain boundaries and surface oxides. We also show that the residual resistance ratio of the polycrystalline niobium films can be used as a figure of merit for qubit lifetime. This comprehensive approach to understanding qubit decoherence charts a pathway for materials-driven improvements of superconducting qubit performance. Understanding the connection between qubit coherence and microscopic materials properties is vital for improving device performance. Here, the relaxation times of superconducting transmon qubits are found to be directly correlated with Nb film properties such as grain size and surface oxide composition.

Inhibiting the Nrf2:Keap1 interaction to trigger cytoprotective gene expression is a promising treatment strategy for oxidative stress-related diseases. A short linear motif from Nrf2 has the potential to directly inhibit this protein-protein interaction, but poor stability and limited cellular uptake impede its therapeutic development. To address these limitations, we utilized an integrated molecular grafting strategy to re-engineer the Nrf2 motif. We combined the motif with an engineered non-native disulfide bond and a cell-penetrating peptide onto a single multifunctionalizable and ultrastable molecular scaffold, namely, the cyclotide MCoTI-II, resulting in the grafted peptide MCNr-2c. The engineered disulfide bond enhanced the conformational rigidity of the motif, resulting in a nanomolar affinity of MCNr-2c for Keap1. The cell-penetrating peptide led to an improved cellular uptake and increased ability to enhance the intracellular expression of two well-described Nrf2-target genes NQO1 and TALDO1. Furthermore, the stability of the scaffold was inherited by the grafted peptide, which became resistant to proteolysis in serum. Overall, we have provided proof-of-concept for a strategy that enables the encapsulation of multiple desired and complementary activities into a single molecular entity to design a Keap1-targeted inhibitor. We propose that this integrated approach could have broad utility for the design of peptide drug leads that require multiple functions and/or biopharmaceutical properties to elicit a therapeutic activity.

The seasonal variability of the carbonate system in the eastern Mediterranean Sea (EMed) was investigated based on discrete total alkalinity (AT), total dissolved inorganic carbon (CT), and pH measurements collected during three cruises around Crete between June 2018 and March 2019. This study presents a detailed description of this new carbonate chemistry dataset in the eastern Mediterranean Sea. We show that the North Western Levantine Basin (NWLB) is unique in terms of range of AT variation vs. CT variation in the upper water column over an annual cycle. The reasons for this singularity of the NWLB can be explained by the interplay between strong evaporation and the concomitant consumption of CT by autotrophic processes. The high range of AT variations, combined to temperature changes, has a strong impact on the variability of the seawater pCO2 (pCO2SW). Based on Argo float data, an entire annual cycle for pCO2SW in the NWLB has been reconstructed in order to estimate the temporal sequence of the potential “source” and “sink” of atmospheric CO2. By combining this dataset with previous observations in the NWLB, this study shows a significant ocean acidification and a decrease in the oceanic surface pHT25 of −0.0024 ± 0.0004 pHT25 units.a–1. The changes in the carbonate system are driven by the increase of atmospheric CO2 but also by unexplained temporal changes in the surface AT content. If we consider that the EMed will, in the future, encounter longer, more intense and warmer summer seasons, this study proposes some perspectives on the carbonate system functioning of the “future” EMed.

In the eastern Mediterranean Sea, satellites have observed events of spring surface‐chlorophyll increase in the Rhodes Gyre region recurring intermittently. Few in situ biogeochemical data, however, exist to confirm their consistency, elucidate their seasonal characteristics, or discriminate among the possible drivers. During the year 2018, an array of BGC‐Argo floats was deployed in the region, collecting the first‐ever annual time series of in situ profiles of biogeochemical parameters in this area. Their observations demonstrated that nitrates, driven by mixed‐layer dynamics, were available at surface from December 2018 onwards and could have sustained phytoplankton growth. Phytoplankton accumulation at the surface was observed by satellite only in March 2019 when the mixed‐layer depth shoaled. These findings confirm that blooms occurring before the start of seasonal stratification are not easily recorded by satellite observations and reaffirm the need to consolidate the BGC‐Argo network to establish time series of the evolution of biogeochemical processes. Plain Language Summary The Levantine Sea, the easternmost area of Mediterranean Sea, is considered one of the poorest oceans on the Earth in terms of abundance of phytoplankton, the microscopic organisms that fuel the marine food web. However, historical data and satellite maps of chlorophyll (the pigment that reveals phytoplankton presence in the water) show episodic increases in the concentration of this pigment in the area near the island of Rhodes. To elucidate the characteristics of these events, a set of six robotic instruments (i.e. the BGC‐Argo floats) was deployed in the Levantine Sea in 2018. A BGC‐Argo float is an autonomous, free‐floating instrument that makes oceanic observations over the first 2,000 meters of the water column on a regular basis. This article presents an analysis of the data collected by these six robots. They provided the very first annual time series of biogeochemical observations in area, including during winter, when ship and satellite data are hard to collect. Our results reveal the increase in phytoplankton occurring before the start of seasonal stratification, increase that is not easily recorded by satellite observations and reaffirm the need to consolidate the BGC‐Argo network to establish time series of the evolution of biogeochemical processes.

Reaction of Fe(II) with the tris-(pyridin-2-yl)ethoxymethane (py3C-OEt) tripodal ligand in the presence of the pseudohalide ancillary NCSe− (E = S, Se, BH3) ligand leads to the mononuclear complex [Fe(py3C-OEt)2][Fe(py3C-OEt)(NCSe)3]2·2CH3CN (3), which has been characterised as an isomorph of the two previously reported complexes, Fe(py3C-OEt)2][Fe(py3C-OEt)(NCE)3]2·2CH3CN, with E = S (1), BH3 (2). X-ray powder diffraction of the three complexes (1–3), associated with the previously reported single crystal structures of 1–2, revealed a monomeric isomorph structure for 3, formed by the spin crossover (SCO) anionic [Fe(py3C-OEt)(NCSe)3]− complex, associated with the low spin (LS) [Fe(py3C-OEt)2]2+ cationic complex and two solvent acetonitrile molecules. In the [Fe(py3C-OEt)2]2+ complex, the metal ion environment involves two py3C-OEt tridentate ligands, while the [Fe(py3C-OEt)(NCSe)3]− anion displays a hexacoordinated environment involving three N-donor atoms of one py3C-OEt ligand and three nitrogen atoms arising from the three (NCSe)− coligands. The magnetic studies for 3 performed in the temperature range 300-5-400 K, indicated the presence of a two-step SCO transition centred around 170 and 298 K, while when the sample was heated at 400 K until its complete desolvation, the magnetic behaviour of the high temperature transition (T1/2 = 298 K) shifted to a lower temperature until the two-step behaviour merged with a gradual one-step transition at ca. 216 K.

For more than two decades the SOHO/SWAN instrument has been monitoring the full-sky hydrogen backscattered Lyman-α emission, and the derived three-dimensional solar wind proton flux. We present a comparison of the time series of the latitude-integrated hydrogen ionization rates (β) derived from the inversion of the SWAN full-sky maps with the integrated coronal electron density derived from the inversion of SOHO/LASCO-C2 white light images. The analysis shows a variable time lag of the SWAN β of a few Carrington rotations, correlated with the solar cycle phase (larger delay during solar maxima compared to minima). This is a direct consequence of the variation of the size of the hydrogen ionization cavity and the time it takes for hydrogen atoms to propagate in the inner heliosphere. This effect should be taken into account in studies of the interstellar neutral populations in interplanetary space.

Legionella pneumophila cell surface hydrophobicity and charge are important determinants of their mobility and persistence in engineered water systems (EWS). These surface properties may differ depending on the growth phase of L. pneumophila resulting in variable adhesion and persistence within EWS. We describe the growth-dependent variations in L. pneumophila cell surface hydrophobicity and surface charge using the microbial adhesion to hydrocarbon assay and microelectrophoresis, respectively, and their role in cell adhesion to stainless steel using a quartz crystal microbalance with dissipation (QCM-D) monitoring instrument. We observed a steady increase in L. pneumophila hydrophobicity during their lifecycle in culture media. Cell surfaces of stationary phase L. pneumophila were significantly more hydrophobic than their lag and midexponential counterparts. No significant changes in L. pneumophila cell surface charge were noted. Morphology of L. pneumophila remained relatively constant throughout their lifecycle. In the QCM-D study, lag and exponential phase L. pneumophila weakly adhered to stainless steel surfaces resulting in viscoelastic layers. In contrast, stationary phase bacteria were tightly and irreversibly bound to the surfaces, forming rigid layers. Our results suggest that the stationary phase of L. pneumophila would highly favour their adhesion to plumbing surfaces and persistence in EWS.

Proprotein convertase subtilisin/kexin type 9 (PCSK9) is a clinically validated target for treating hypercholesterolemia. Peptide-based PCSK9 inhibitors have attracted pharmaceutical interest, but the effect of multivalency on bioactivity is poorly understood. Here we designed bivalent and tetravalent dendrimers, decorated with the PCSK9 inhibitory peptides Pep2-8[RRG] or P9-38, to study relationships between peptide binding affinity, peptide valency, and PCSK9 inhibition. Increased valency resulted in improved PCSK9 inhibition for both peptides, with activity improvements of up to 100-fold achieved for the P9-38-decorated dendrimers compared to monomeric P9-38 in in vitro competition binding assays. Furthermore, the P9-38-decorated dendrimers showed improved potency at restoring functional low-density lipoprotein (LDL) receptor levels and internalizing LDL in the presence of PCSK9, demonstrating significant cell-based activity at picomolar concentrations. This study demonstrates the potential of increasing valency as a strategy for increasing the efficacy of peptide-based PCSK9 therapeutics.

Doped ferroelectric HfO2 is highly promising for integration into complementary metal-oxide semiconductor (CMOS) technology for devices such as ferroelectric nonvolatile memory and low-power field-effect transistors (FETs). We report the direct measurement of the energy barriers between various metal electrodes (Pt, Au, Ta, TaN, Ti/Pt, Ni, Al) and hafnium zirconium oxide (Hf0.58Zr0.42O2, HZO) using internal photoemission (IPE) spectroscopy. Results are compared with valence band offsets determined using the three-sample X-ray photoelectron spectroscopy (XPS) as well as the two-sample hard X-ray photoelectron spectroscopy (HAXPES) techniques. Both XPS and IPE indicate roughly the same dependence of the HZO barrier on metal work function with a slope of 0.8 ± 0.5. XPS and HAXPES-derived barrier heights are on average about 1.1 eV smaller than barrier heights determined by IPE, suggesting the presence of negative charge in the HZO.

An ordered 3D printed chromatography stationary phase was used to purify M13 bacteriophage (M13) directly from crude cell culture. This new approach, which offers the same advantages as expanded bed adsorption (EBA) with regard to tolerating solids-laden feed streams but without the corresponding issues associated with fluidized bed stability that affect the latter, can be described as “printed monolith adsorption (PMA)”. PMA columns (5, 10 and 15 cm length by 1 cm diameter) were made via a wax templating method from cross-linked cellulose hydrogel and functionalized with a quaternary amine ligand. The recovery of M13 was found to be strongly linked to load flow rate, with the highest recovery 89.7% ± 6% for 1.4 × 1011 pfu/mL of resin occurring at 76 cm/h with a 10 cm column length. A recovery of 87.7% ± 5% for 1.49 × 1011 pfu/mL of media was achieved with a 15 cm column length under conditions comparable to a reported EBA process. The PMA process was completed three times faster than EBA because PMA flow rates can readily be adjusted during operation, with high flow rates and low back pressure, which is unique to the ordered monolithic media geometry used. Equilibration, wash, and cleaning steps were carried out at high flow rates (611 cm/h), minimizing process time and were limited only by the volumetric flow rate capacity of the pumps used, rather than column back pressure (

The European Parliament recently approved a new law banning single-use plastic items for 2021 such as plastic plates, cutlery, straws, cotton swabs, and balloon sticks. Transition to a bioeconomy involves the substitution of these banned products with biodegradable materials. Several materials such as polylactic acid (PLA), polybutylene adipate terephthalate (PBAT), poly(butylene succinate) (PBS), polyhydroxybutyrate-valerate (PHBV), Bioplast, and Mater-Bi could be good candidates to substitute cotton swabs, but their biodegradability needs to be tested under marine conditions. In this study, we described the microbial life growing on these materials, and we evaluated their biodegradability in seawater, compared with controls made of non-biodegradable polypropylene (PP) or biodegradable cellulose. During the first 40 days in seawater, we detected clear changes in bacterial diversity (Illumina sequencing of 16S rRNA gene) and heterotrophic activity (incorporation of 3H-leucine) that coincided with the classic succession of initial colonization, growth, and maturation phases of a biofilm. Biodegradability of the cotton swab sticks was then tested during another 94 days under strict diet conditions with the different plastics as sole carbon source. The drastic decrease of the bacterial activity on PP, PLA, and PBS suggested no bacterial attack of these materials, whereas the bacterial activity in PBAT, Bioplast, Mater-Bi, and PHBV presented similar responses to the cellulose positive control. Interestingly, the different bacterial diversity trends observed for biodegradable vs. non-biodegradable plastics allowed to describe potential new candidates involved in the degradation of these materials under marine conditions. This better understanding of the bacterial diversity and activity dynamics during the colonization and biodegradation processes contributes to an expanding baseline to understand plastic biodegradation in marine conditions and provide a foundation for further decisions on the replacement of the banned single-used plastics.

9 pages, 7 figures, 3 tables, Coastal marine ecosystems are strongly influenced by different occasional events, such as intense winds, mixing, rain and river discharges. These events can directly or indirectly cause changes in dissolved organic matter (DOM) quality through a cascade of different biotic and abiotic processes. Changes in DOM quality are often associated with changes in DOM optical properties. Thus, examining the dynamics of chromophoric DOM (CDOM) can provide valuable information about biological and physical processes that have occurred in the ecosystem. Episodic meteorological events, particularly in temperate areas, appear very abruptly and induce very rapid responses; therefore, high time-resolved measurements are needed to capture them. We used a weekly sampling scheme to characterize DOM and nutrient dynamics in the NW Mediterranean coastal station ‘SOLA’. From February 2013 to April 2014, we measured several physical and chemical variables including temperature, salinity, inorganic nutrients, chlorophyll a dissolved organic carbon (DOC), CDOM and fluorescent DOM (FDOM). During this period, two extremely high fresh water intrusions greatly influenced the dynamics of some DOM fractions, in particular the FDOM. Inorganic nutrients and chlorophyll a showed seasonal patterns: A winter period characterized by a high nutrient concentration in surface waters favored the phytoplankton spring bloom; then, summer stratification extended until autumn. This stratification led to nutrient depletion and, consequently, lower chlorophyll a values in the photic zone. The CDOM and FDOM optical active fractions did not follow temporal trends similar to total DOC. This was likely because the potential sources and sinks of these DOM pools are microbial activity and light exposure, and these were acting simultaneously but in opposite directions. Interestingly, DOC exhibited the highest concentrations in summer, coinciding with nutrient and chlorophyll a minima. To explain this mismatch, we propose a sequence of abiotic and biotic phenomena that drive DOC temporal dynamics, This study was supported by the MERMeX/MISTRALS project and is a contribution to international SOLAS program and the projects ANIMA (CTM2015-65720-R) and DOREMI (CTM2012-34294) from the Spanish Ministerio de Economia y Competitividad. E.D. Sánchez-Pérez would like to thank the Consejo Nacional de Ciencia y Tecnologia (CONACyT) for their financial support through a PhD fellowship, With the funding support of the ‘Severo Ochoa Centre of Excellence’ accreditation (CEX2019-000928-S), of the Spanish Research Agency (AEI)

Summary NTRK1 gene fusions are actionable drivers of numerous human malignancies. Here, we show that expression of the TPR-NTRK1 fusion kinase in immortalized mouse pancreatic ductal epithelial (IMPE) (pancreas) or mouse lung epithelial (MLE-12) cells is sufficient to promote rapidly growing tumors in mice. Both tumor models are exquisitely sensitive to targeted inhibition with entrectinib, a tropomyosin-related kinase A (TRKA) inhibitor. Initial regression of NTRK1-driven tumors is driven by induced expression of BIM, such that BIM silencing leads to a diminished response to entrectinib in vivo. However, the emergence of drug-resistant disease limits the long-term durability of responses. Based on the reactivation of RAF>MEK>ERK signaling observed in entrectinib-treated tumors, we show that the combination of entrectinib plus the MEK1/2 inhibitor cobimetinib dramatically forestalls the onset of drug resistance in vivo. Collectively, these data provide a mechanistic rationale for rapid clinical deployment of combined inhibition of TRKA plus MEK1/2 in NTRK1-driven cancers.

International audience; Particulate organic matter (POM) in coastal systems is a mixture of different organic matter (OM) sources originating from land and sea. Among sources, bacterial biomass plays a large role in OM processing and carbon recycling in the ocean and is often neglected as a source in common approaches. The present study proposes to use elemental and isotopic ratio of carbon and nitrogen (C:N, δ13C, δ15N) and fatty acids to investigate the relationship between bacteria and surface water POM composition of three systems with different characteristics (two marine and one estuarine) over an annual cycle. Overall, our results highlight a positive relationship between bacterial contribution and continental degraded or undergoing degradation POM for all the studied systems and an inverse relationship with pelagic primary producers. At multisystem scale, high bacterial contribution is linked to high proportion of refractory terrestrial material characterizing estuarine stations whereas in marine systems, the occurrence of bacteria is mainly linked to river POM. Over the annual cycle, bacterial markers are more abundant during the winter period characterized by larger river and/or benthic POM contribution. This seasonal pattern is mainly driven by changes in river flows and resuspension. This study highlights the importance of bacterial compartment as a component of coastal and estuarine POM. Even though these results remains semi-quantitative, similar studies in other types of systems can help to understand microbial role in OM dynamic and to better estimate bacterial source in carbon budgets and food web studies.

Disulfide-rich peptides are a class of molecules for which NMR spectroscopy has been the primary tool for structural characterization. Here, we explore whether the process can be achieved by using structural information encoded in chemical shifts. We examine (i) a representative set of five cyclic disulfide-rich peptides that have high-resolution NMR and X-ray structures and (ii) a larger set of 100 disulfide-rich peptides from the PDB. Accuracy of the calculated structures was dependent on the methods used for searching through conformational space and for identifying native conformations. Although Hα chemical shifts could be predicted reasonably well using SHIFTX, agreement between predicted and experimental chemical shifts was sufficient for identifying native conformations for only some peptides in the representative set. Combining chemical shift data with the secondary structure information and potential energy calculations improved the ability to identify native conformations. Additional use of sparse distance restraints or homology information to restrict the search space also improved the resolution of the calculated structures. This study demonstrates that abbreviated methods have potential for elucidation of peptide structures to high resolution and further optimization of these methods, e.g., improvement in chemical shift prediction accuracy, will likely help transition these methods into the mainstream of disulfide-rich peptide structural biology.

This review highlights the predominant role that NMR has had in determining the structures of cyclotides, a fascinating class of macrocyclic peptides found in plants. Cyclotides contain a cystine knot, a compact structural motif that is constrained by three disulfide bonds and able to resist chemical and biological degradation. Their resistance to proteolytic degradation has made cyclotides appealing as drug leads. Herein, we examine the developments that led to the identification and conclusive determination of the disulfide connectivity of cyclotides and describe in detail the structural features of exemplar cyclotides. We also review the role that X-ray crystallography has played in resolving cyclotide structures and describe how racemic crystallography opened up the possibility of obtaining previously inaccessible X-ray structures of cyclotides.

Rapid clearance by renal filtration is a major impediment to the translation of small bioactive biologics into drugs. To extend serum t(1/2), a commonly used approach is to attach drug leads to the G-related albumin-binding domain (ABD) to bind albumin and evade clearance. Despite the success of this approach in extending half-lives of a wide range of biologics, it is unclear whether the existing constructs are optimized for binding and size; any improvements along these lines could lead to improved drugs. Characterization of the biophysics of binding of an ABD to albumin in solution could shed light on this question. Here, we examine the binding of an ABD to human serum albumin using isothermal titration calorimetry and assess the structural integrity of the ABD using CD, NMR, and molecular dynamics. A structure-activity analysis of truncations of the ABD suggests that downsized variants could replace the full-length domain. Reducing size could have the benefit of reducing potential immunogenicity problems. We further showed that one of these variants could be used to design a bifunctional molecule with affinity for albumin and a serum protein involved in cholesterol metabolism, PCSK9, demonstrating the potential utility of these fragments in the design of cholesterol-lowering drugs. Future work could extend these in vitro binding studies to other ABD variants to develop therapeutics. Our study presents new understanding of the solution structural and binding properties of ABDs, which has implications for the design of next-generation long-lasting therapeutics.

Northwest Atlantic leatherback sea turtles (Dermochelys coriacea) are endangered and low hatch success limits potential for population recovery. We examined essential and nonessential metal concentrations in 43 eggs from nests on St. Kitts to determine if there was a relationship with hatch success. Whole homogenized embryos and undeveloped eggs contained detectable concentrations of arsenic, barium, copper, iron, selenium, vanadium, and zinc, but not beryllium, cadmium, chromium, cobalt, lead, mercury, molybdenum, and thallium. Of detected metals, only vanadium concentrations negatively correlated with hatch success (P = 0.01). Manganese and vanadium were associated with pneumonia occurring in the nest, and arsenic with renal mineralization. This study adds to the knowledge regarding baseline values for environmental contaminants in sea turtles, supporting the trend that leatherback eggs have relatively low concentrations of toxic metals, lacking a strong relationship with hatch success, and normally contain the essential elements copper, iron, selenium, and zinc.

Retinitis pigmentosa (RP) and associated inherited retinal diseases (IRDs) are caused by rod photoreceptor degeneration, necessitating therapeutics promoting rod photoreceptor survival. To address this, we tested compounds for neuroprotective effects in zebrafish and mouse RP models, reasoning drugs effective across species may translate better clinically. We first performed a large-scale phenotypic drug screen using a larval zebrafish model of inducible RP. 2,934 compounds, mostly human-approved drugs, were tested across six concentrations. Statistically, 113 compounds achieved “hit” status. Secondary tests of 42 high-priority hits confirmed eleven lead compounds. Nine leads were then evaluated in mouse RP models, with six exhibiting neuroprotective effects. An analysis of potential mechanisms of action suggested complementary activities. Paired lead compound assays in zebrafish showed additive neuroprotective effects for the majority. These results highlight the value of cross-species phenotypic drug discovery and suggest combinatorial drug therapies may provide enhanced therapeutic benefits for patients with RP and IRDs.

In recent years, 3D printing has enabled the fabrication of complex designs, with low-cost customization and an ever-increasing range of materials. Yet, these abilities have also created an enormous challenge in optimizing a large number of process parameters, especially in the 3D printing of swellable, non-toxic, biocompatible and biodegradable materials, so-called bio-ink materials. In this work, a cellulose gel, made out of aqueous solutions of cellulose, sodium hydroxide and urea, was used to demonstrate the formation of a shear thinning bio-ink material necessary for an extrusion-based 3D printing. After analysing the shear thinning behaviour of the cellulose gel by rheometry a Design of Experiments (DoE) was applied to optimize the 3D bioprinter settings for printing the cellulose gel. The optimum print settings were then used to print a human ear shape, without a need for support material. The results clearly indicate that the found settings allow the printing of more complex parts with high-fidelity. This confirms the capability of the applied method to 3D print a newly developed bio-ink material.

Purpose To compare corneal endothelial damage associated with 2 techniques for preloaded Descemet membrane endothelial keratoplasty (DMEK): a tri-folded graft stored in a plastic cartridge designed for DMEK and a scrolled graft stored in a modified Jones Tube, at the time of preparation and after shipping. Methods DMEK grafts were prepared at the Rocky Mountain Lions Eye Bank. The grafts were either tri-folded and loaded in a plastic cartridge or scrolled and loaded into a modified Jones Tube. In each group, the grafts were then either immediately removed from the cartridges or shipped for 48 hours. The grafts were then stained with Calcein AM and imaged using a fluorescent microscope. Endothelial cell loss (ECL) was determined using trainable segmentation in Fiji by 2 graders. At each time point, rates of ECL loss were compared across the 2 groups. To explore the role of donor characteristics, a multivariable regression model was produced to account for method (tri-folding vs. scroll), donor age, donor gender, death-to-preservation time, death-to-preparation time, and shipping. Results A total of 40 grafts were prepared, processed, imaged, and analyzed. No significant difference in cell loss was seen between groups at either time point alone. In the multivariate model, no significant increase in cell loss was associated with either tri-folding (3.7% less ECL; P = 0.051) or shipping (4.3% less ECL; P = 0.049). Conclusions All techniques used resulted in clinically acceptable levels of ECL. Tri-folded tissue in a plastic cartridge did not result in ECL inferior to a scroll when prepared either immediately or preloaded for 48 hours.

Cenozoic sedimentary rocks in the southern Texas Gulf Coastal Plains contain abundant continental carbonates that are useful for reconstructing terrestrial paleoclimate and paleoenvironment in a region near sea-level. Our field observations and thin section characterizations of the Oligocene and Miocene continental carbonates in south Texas identified three types of pedogenic carbonates, including rhizoliths, carbonate nodules, and platy horizons, and two types of groundwater carbonates, including carbonate-cemented beds and carbonate concretions, with distinctive macromorphologic and micromorphologic features. Based on preservations of authigenic microfabrics and variations of carbon and oxygen isotopic compositions, we suggest these carbonates experienced minimal diagenesis, and their stable isotopic compositions reflect paleoclimate and paleoenvironment in south Texas. Our Oligocene and Miocene carbonate clumped isotope temperatures (T(Δ47)) are 23–28 °C, slightly less than or comparable to the range of modern mean annual and mean warm season air temperature (21–27 °C) in the study area. These T(Δ47) values do not show any dependency on carbonate-type, or trends through time suggesting that groundwater carbonates were formed at shallow depths. These data could indicate that air temperature in south Texas was relatively stable since the early Oligocene. The reconstructed paleo-surface water δ18O values are similar to modern surface water which could indicate that meteoric water δ18O values also remained stable since the early Oligocene. Mean pedogenic carbonate δ13C values increased ~ − 4.6‰ during the late Miocene, most likely reflecting an expansion of C4 grassland in south Texas. This study provides the first mid- and late Cenozoic continental records of paleoclimate and paleoecology in a low-latitude, near sea-level region.

Several cyclic peptides have been reported to have unexpectedly high membrane permeability. Of these, cyclosporin A is perhaps the most well-known example, particularly in light of its relatively high molecular weight. Observations that cyclosporin A changes conformation depending on its solvent environment led to the hypothesis that conformational dynamics is a prerequisite for its permeability; however, this hypothesis has been difficult to validate experimentally. Here, we use molecular dynamics simulations to explicitly determine the conformational behavior of cyclosporin A and other related cyclic peptides as they spontaneously transition between different environments, including through a lipid bilayer. These simulations are referenced against simulations in explicit water, chloroform, and cyclohexane and further validated against NMR experiments, measuring conformational exchange, nuclear spin relaxation, and three-dimensional structures in membrane-mimicking environments, such as in dodecylphosphocholine micelles, to build a comprehensive understanding of the role of dynamics. We find that conformational flexibility is a key determinant of the membrane permeability of cyclosporin A and similar membrane-permeable cyclic peptides, as conformationally constrained variants have limited movement into, then through, and finally out of the membrane in silico. We envisage that a better understanding of dynamics might thus provide new opportunities to modulate peptide function and enhance their delivery.

As the quest toward novel materials proceeds, improved characterization technologies are needed. In particular, the atomic thickness in graphene and other 2D materials renders some conventional technologies obsolete. Characterization technologies at wafer level are needed with enough sensitivity to detect strain in order to inform fabrication. In this work, NEXAFS spectroscopy was combined with simulations to predict lattice parameters of graphene grown on copper and further transferred to a variety of substrates. The strains associated with the predicted lattice parameters are in agreement with experimental findings. The approach presented here holds promise to effectively measure strain in graphene and other 2D systems at wafer levels to inform manufacturing environments.

Summary Fluoride-ion batteries are an attractive energy storage concept analogous to lithium-ion batteries but feature an inverted paradigm where anions are the principal charge carriers. Insertion hosts that can reversibly insert fluoride ions at room temperature are exceedingly sparse. Here, we report that topochemical insertion of fluoride ions in FeSb2O4 involves Fe2+/Fe3+ redox but is mediated by multi-center synergies between iron and antimony centers. Separation of the redox center from the p-block coordination site alleviates structural strain by enabling compensatory contraction and expansion of FeO6 and SbO3 polyhedra, respectively. p-block electron lone pairs play a critical role in weakening anion-lattice interactions, enabling reversible fluoride-ion diffusion across microns. The results illuminate the key principle that interactions traceable to stereoactive lone pairs can be used to mediate anion-lattice interactions and suggest that anion insertion hosts can be designed by pairing redox-active transition metals with p-block cations bearing stereochemically active electron lone pairs.