RESUMEN
Phosphatidylinositol (PtdIns) transfer proteins (PITPs) enhance the activities of PtdIns 4-OH kinases that generate signaling pools of PtdIns-4-phosphate. In that capacity, PITPs serve as key regulators of lipid signaling in eukaryotic cells. Although the PITP phospholipid exchange cycle is the engine that stimulates PtdIns 4-OH kinase activities, the underlying mechanism is not understood. Herein, we apply an integrative structural biology approach to investigate interactions of the yeast PITP Sec14 with small-molecule inhibitors (SMIs) of its phospholipid exchange cycle. Using a combination of X-ray crystallography, solution NMR spectroscopy, and atomistic MD simulations, we dissect how SMIs compete with native Sec14 phospholipid ligands and arrest phospholipid exchange. Moreover, as Sec14 PITPs represent new targets for the development of next-generation antifungal drugs, the structures of Sec14 bound to SMIs of diverse chemotypes reported in this study will provide critical information required for future structure-based design of next-generation lead compounds directed against Sec14 PITPs of virulent fungi.
Asunto(s)
Antifúngicos , Diseño de Fármacos , Proteínas de Transferencia de Fosfolípidos , Proteínas de Saccharomyces cerevisiae , Transporte Biológico/efectos de los fármacos , Fosfatidilinositoles/metabolismo , Proteínas de Transferencia de Fosfolípidos/antagonistas & inhibidores , Saccharomyces cerevisiae/metabolismo , Proteínas de Saccharomyces cerevisiae/antagonistas & inhibidores , Transducción de Señal , Antifúngicos/química , Antifúngicos/farmacologíaRESUMEN
Zero-valent iron (ZVI) is a promising material for the remediation of Cd-contaminated paddy soils. However, the effects of ZVI added during flooding or drainage processes on cadmium (Cd) retention remain unclear. Herein, Cd-contaminated paddy soil was incubated for 40 days of flooding and then for 15 days of drainage, and the underlying mechanisms of Cd immobilization coupled with Fe/S/N redox processes were investigated. The addition of ZVI to the flooding process was more conducive to Cd immobilization. Less potential available Cd was detected by adding ZVI before flooding, which may be due to the increase in paddy soil pH and newly formed secondary Fe minerals. Moreover, the reductive dissolution of Fe minerals promoted the release of soil colloids, thereby increasing significantly the surface sites and causing Cd immobilization. Additionally, the addition of ZVI before flooding played a vital role in Cd retention after soil drainage. In contrast, the addition of ZVI in the drainage phase was not conducive to Cd retention, which might be due to the rapid decrease in soil pH that inhibited Cd adsorption and further immobilization on soil surfaces. The findings of this study demonstrated that Cd availability in paddy soil was largely reduced by adding ZVI during the flooding period and provide a novel insight into the mechanisms of ZVI remediation in Cd-contaminated paddy soils.
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Oryza , Contaminantes del Suelo , Cadmio/análisis , Hierro , Suelo , Contaminantes del Suelo/análisis , MineralesRESUMEN
Chemical oxidation of As(III) by iron (Fe) oxyhydroxides has been proposed to occur under anoxic conditions and may play an important role in stabilization and detoxification of As in subsurface environments. However, this reaction remains controversial due to lack of direct evidence and poorly understood mechanisms. In this study, we show that As(III) oxidation can be facilitated by Fe oxyhydroxides (i.e., goethite) under anoxic conditions coupled with the reduction of structural Fe(III). An excellent electron balance between As(V) production and Fe(III) reduction is obtained. The formation of an active metastable Fe(III) phase at the defective surface of goethite due to atom exchange is responsible for the oxidation of As(III). Furthermore, the presence of defects (i.e., Fe vacancies) in goethite can noticeably enhance the electron transfer (ET) and atom exchange between the surface-bound Fe(II) and the structural Fe(III) resulting in a two time increase in As(III) oxidation. Atom exchange-induced regeneration of active goethite sites is likely to facilitate As(III) coordination and ET with structural Fe(III) based on electrochemical analysis and theoretical calculations showing that this reaction pathway is thermodynamically and kinetically favorable. Our findings highlight the synergetic effects of defects in the Fe crystal structure and Fe(II)-induced catalytic processes on anoxic As(III) oxidation, shedding a new light on As risk management in soils and subsurface environments.
Asunto(s)
Compuestos de Hierro , Hierro , Hierro/química , Compuestos de Hierro/química , Minerales/química , Oxidación-Reducción , Compuestos Ferrosos/química , Compuestos Férricos/químicaRESUMEN
The dark production of reactive oxygen species (ROS) coupled to biogeochemical cycling of iron (Fe) plays a pivotal role in controlling arsenic transformation and detoxification. However, the effect of secondary atom incorporation into Fe(III) oxyhydroxides on this process is poorly understood. Here, we show that the presence of oxygen vacancy (OV) as a result of Cu incorporation in goethite substantially enhances the As(III) oxidation by Fe(II) under oxic conditions. Electrochemical and density functional theory (DFT) evidence reveals that the electron transfer (ET) rate constant is enhanced from 0.023 to 0.197 s-1, improving the electron efficiency of the surface-bound Fe(II) on OV defective surfaces. The cascade charge transfer from the surface-bound Fe(II) to O2 mediated by Fe(III) oxyhydroxides leads to the O-O bond of O2 stretching to 1.46-1.48 Å equivalent to that of superoxide (â¢O2-), and â¢O2- is the predominant ROS responsible for As(III) oxidation. Our findings highlight the significant role of atom incorporation in changing the ET process on Fe(III) oxyhydroxides for ROS production. Thus, such an effect must be considered when evaluating Fe mineral reactivity toward changing their surface chemistry, such as those noted here for Cu incorporation, which likely determines the fates of arsenic and other redox sensitive pollutants in the environments with oscillating redox conditions.
Asunto(s)
Arsénico , Compuestos Férricos , Compuestos Férricos/química , Oxígeno , Especies Reactivas de Oxígeno , Arsénico/química , Minerales/química , Oxidación-Reducción , Compuestos Ferrosos/química , Estrés OxidativoRESUMEN
Electron shuttles such cysteine play an important role in Fe cycle and its availability in soils, while the roles of pH and organic ligands in this process are poorly understood. Herein, the reductive dissolution process of goethite by cysteine were explored in the presence of organic ligands. Our results showed that cysteine exhibited a strong reactivity towards goethite - a typical iron minerals in paddy soils with a rate constant ranging from 0.01 to 0.1 hr-1. However, a large portion of Fe(II) appeared to be "structural species" retained on the surface. The decline of pH was favorable to generate more Fe(II) ions and enhancing tendency of Fe(II) release to solution. The decline of generation of Fe(II) by increasing pH was likely to be caused by a lower redox potential and the nature of cysteine pH-dependent adsorption towards goethite. Interestingly, the co-existence of oxalate and citrate ligands also enhanced the rate constant of Fe(II) release from 0.09 to 0.15 hr-1; nevertheless, they negligibly affected the overall generation of Fe(II) in opposition to the pH effect. Further spectroscopic evidence demonstrated that two molecules of cysteine could form disulfide bonds (S-S) to generate cystine through oxidative dehydration, and subsequently, inducing electron transfer from cysteine to the structural Fe(III) on goethite; meanwhile, those organic ligands act as Fe(II) "strippers". The findings of this work provide new insights into the understanding of the different roles of pH and organic ligands on the generation and release of Fe induced by electron shuttles in soils.
Asunto(s)
Cisteína , Compuestos de Hierro , Compuestos Férricos , Concentración de Iones de Hidrógeno , Ligandos , Minerales , Oxidación-Reducción , SolubilidadRESUMEN
The membrane-bound serine protease matriptase belongs to a rare subset of serine proteases that display significant activity in the zymogen form. Matriptase is critically involved in epithelial differentiation and homeostasis, and insufficient regulation of its proteolytic activity directly causes onset and development of malignant cancer. There is strong evidence that the zymogen activity of matriptase is sufficient for its biological function(s). Activated matriptase is inhibited by the two Kunitz-type inhibitor domain-containing hepatocyte growth factor activator inhibitors 1 (HAI-1) and HAI-2, however, it remains unknown whether the activity of the matriptase zymogen is regulated. Using both purified proteins and a cell-based assay, we show that the catalytic activity of the matriptase zymogen towards a peptide-based substrate as well as the natural protein substrates, pro-HGF and pro-prostasin, can be inhibited by HAI-1 and HAI-2. Inhibition of zymogen matriptase by HAI-1 and HAI-2 appears similar to inhibition of activated matriptase and occurs at comparable inhibitor concentrations. This indicates that HAI-1 and HAI-2 interact with the active sites of zymogen and activated matriptase in a similar manner. Our results suggest that HAI-1 and HAI-2 regulate matriptase zymogen activity and thus may act as regulators of matriptase trans(auto)-activation. Due to the main localisation of HAI-2 in the ER and HAI-1 in the secretory pathway and on the cell surface, this regulation likely occurs both in the secretory pathway and on the plasma membrane. Regulation of an active zymogen form of a protease is a novel finding.
Asunto(s)
Proteínas Inhibidoras de Proteinasas Secretoras/metabolismo , Serina Endopeptidasas/metabolismo , Membrana Celular/metabolismo , Células HEK293 , Humanos , Glicoproteínas de Membrana/metabolismo , Vías SecretorasRESUMEN
Matriptase is a member of the type-II transmembrane serine protease (TTSP) family and plays a crucial role in the development and maintenance of epithelial tissues. As all chymotrypsin-like serine proteases, matriptase is synthesized as a zymogen (proform), requiring a cleavage event for full activity. Recent studies suggest that the zymogen of matriptase possesses enough catalytic activity to not only facilitate autoactivation, but also carry out its in vivo functions, which include activating several proteolytic and signaling cascades. Inhibition of zymogen matriptase may therefore be a highly effective approach for limiting matriptase activity. To this end, here we sought to characterize the catalytic activity of human zymogen matriptase and to develop mAb inhibitors against this enzyme form. Using a mutated variant of matriptase in which the serine protease domain is locked in the zymogen conformation, we confirmed that the zymogen form of human matriptase has catalytic activity. Moreover, the crystal structure of the catalytic domain of zymogen matriptase was solved to 2.5 Å resolution to characterize specific antibody-based matriptase inhibitors and to further structure-based studies. Finally, we describe the first antibody-based competitive inhibitors that target both the zymogen and activated forms of matriptase. We propose that these antibodies provide a more efficient way to regulate matriptase activity by targeting the protease both before and after its activation and may be of value for both research and preclinical applications.
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Anticuerpos Monoclonales/química , Precursores Enzimáticos/química , Inhibidores de Proteasas/química , Proteolisis , Serina Endopeptidasas/química , Cristalografía por Rayos X , Precursores Enzimáticos/antagonistas & inhibidores , Células HEK293 , Humanos , Dominios ProteicosRESUMEN
Hepatocyte growth factor activator inhibitor-1 (HAI-1) is a type I transmembrane protein and inhibitor of several serine proteases, including hepatocyte growth factor activator and matriptase. The protein is essential for development as knock-out mice die in utero due to placental defects caused by misregulated extracellular proteolysis. HAI-1 contains two Kunitz-type inhibitor domains (Kunitz), which are generally thought of as a functionally self-contained protease inhibitor unit. This is not the case for HAI-1, where our results reveal how interdomain interactions have evolved to stimulate the inhibitory activity of an integrated Kunitz. Here we present an x-ray crystal structure of an HAI-1 fragment covering the internal domain and Kunitz-1. The structure reveals not only that the previously uncharacterized internal domain is a member of the polycystic kidney disease domain family but also how the two domains engage in interdomain interactions. Supported by solution small angle x-ray scattering and a combination of site-directed mutagenesis and functional assays, we show that interdomain interactions not only stabilize the fold of the internal domain but also stimulate the inhibitory activity of Kunitz-1. By completing our structural characterization of the previously unknown N-terminal region of HAI-1, we provide new insight into the interplay between tertiary structure and the inhibitory activity of a multidomain protease inhibitor. We propose a previously unseen mechanism by which the association of an auxiliary domain stimulates the inhibitory activity of a Kunitz-type inhibitor (i.e. the first structure of an intramolecular interaction between a Kunitz and another domain).
Asunto(s)
Glicoproteínas de Membrana/metabolismo , Enfermedades Renales Poliquísticas/metabolismo , Secuencia de Aminoácidos , Animales , Cristalografía por Rayos X , Glicoproteínas de Membrana/química , Ratones , Ratones Noqueados , Modelos Moleculares , Datos de Secuencia Molecular , Unión Proteica , Conformación Proteica , Proteínas Inhibidoras de Proteinasas Secretoras , Proteínas Recombinantes/química , Proteínas Recombinantes/metabolismo , Dispersión del Ángulo Pequeño , Homología de Secuencia de Aminoácido , Resonancia por Plasmón de SuperficieRESUMEN
Fibrinolysis is a process responsible for the dissolution of formed thrombi to re-establish blood flow after thrombus formation. Plasminogen activator inhibitor-1 (PAI-1) inhibits urokinase-type and tissue-type plasminogen activator (uPA and tPA) and is the major negative regulator of fibrinolysis. Inhibition of PAI-1 activity prevents thrombosis and accelerates fibrinolysis. However, a specific antagonist of PAI-1 is currently unavailable for therapeutic use. We screened a panel of uPA variants with mutations at and near the active site to maximize their binding to PAI-1 and identified a potent PAI-1 antagonist, PAItrap. PAItrap is the serine protease domain of urokinase containing active-site mutation (S195A) and four additional mutations (G37bR-R217L-C122A-N145Q). PAItrap inhibits human recombinant PAI-1 with high potency (Kd = 0.15 nM) and high specificity. In vitro using human plasma, PAItrap showed significant thrombolytic activity by inhibiting endogenous PAI-1. In addition, PAItrap inhibits both human and murine PAI-1, allowing the evaluation in murine models. In vivo, using a laser-induced thrombosis mouse model in which thrombus formation and fibrinolysis are monitored by intravital microscopy, PAItrap reduced fibrin generation and inhibited platelet accumulation following vascular injury. Therefore, this work demonstrates the feasibility to generate PAI-1 inhibitors using inactivated urokinase.
Asunto(s)
Fragmentos de Péptidos/farmacología , Inhibidor 1 de Activador Plasminogénico/metabolismo , Activador de Plasminógeno de Tipo Uroquinasa/metabolismo , Secuencia de Aminoácidos , Animales , Dominio Catalítico , Cristalografía por Rayos X , Modelos Animales de Enfermedad , Fibrinólisis , Humanos , Concentración 50 Inhibidora , Cinética , Ratones Endogámicos C57BL , Modelos Moleculares , Proteínas Mutantes/química , Fragmentos de Péptidos/química , Unión Proteica , Trombosis/patología , Activador de Plasminógeno de Tipo Uroquinasa/química , Activador de Plasminógeno de Tipo Uroquinasa/farmacologíaRESUMEN
A decade ago, motif at N-terminus with eight-cysteines (MANEC) was defined as a new protein domain family. This domain is found exclusively at the N-terminus of >400 multi-domain type-1 transmembrane proteins from animals. Despite the large number of MANEC-containing proteins, only one has been characterized at the protein level: hepatocyte growth factor activator inhibitor-1 (HAI-1). HAI-1 is an essential protein, as knockout mice die in utero due to placental defects. HAI-1 is an inhibitor of matriptase, hepsin and hepatocyte growth factor (HGF) activator, all serine proteases with important roles in epithelial development, cell growth and homoeostasis. Dysregulation of these proteases has been causatively implicated in pathological conditions such as skin diseases and cancer. Detailed functional understanding of HAI-1 and other MANEC-containing proteins is hampered by the lack of structural information on MANEC. Although many MANEC sequences exist, sequence-based database searches fail to predict structural homology. In the present paper, we present the NMR solution structure of the MANEC domain from HAI-1, the first three-dimensional (3D) structure from the MANEC domain family. Unexpectedly, MANEC is a new subclass of the PAN/apple domain family, with its own unifying features, such as two additional disulfide bonds, two extended loop regions and additional α-helical elements. As shown for other PAN/apple domain-containing proteins, we propose a similar active role of the MANEC domain in intramolecular and intermolecular interactions. The structure provides a tool for the further elucidation of HAI-1 function as well as a reference for the study of other MANEC-containing proteins.
Asunto(s)
Modelos Moleculares , Proteínas Inhibidoras de Proteinasas Secretoras/química , Secuencias de Aminoácidos , Secuencia de Aminoácidos , Sustitución de Aminoácidos , Humanos , Datos de Secuencia Molecular , Proteínas Mutantes/química , Proteínas Mutantes/clasificación , Proteínas Mutantes/metabolismo , Resonancia Magnética Nuclear Biomolecular , Conformación Proteica , Pliegue de Proteína , Dominios y Motivos de Interacción de Proteínas , Isoformas de Proteínas/química , Señales de Clasificación de Proteína , Estabilidad Proteica , Estructura Terciaria de Proteína , Proteínas Inhibidoras de Proteinasas Secretoras/clasificación , Proteínas Inhibidoras de Proteinasas Secretoras/genética , Proteínas Inhibidoras de Proteinasas Secretoras/metabolismo , Proteínas Recombinantes/química , Proteínas Recombinantes/clasificación , Proteínas Recombinantes/metabolismo , Dispersión del Ángulo Pequeño , Solubilidad , Difracción de Rayos XRESUMEN
The natural product embelin was found to have PAI-1 inhibitory activity with the IC50 value of 4.94µM. Based on the structure of embelin, a series of analogues were designed, synthesized, and evaluated for their ability to inhibit PAI-1. The SAR study on these compounds disclosed that the inhibitory potency largely depended on the hydroxyl groups at C2 and C5, and the length of the alkyl chains at C3 and C6. Compound 11 displayed the best PAI-1 inhibitory potency with the IC50 value of 0.18µM.
Asunto(s)
Benzoquinonas/química , Benzoquinonas/farmacología , Inhibidor 1 de Activador Plasminogénico/química , Inhibidor 1 de Activador Plasminogénico/metabolismo , Benzoquinonas/síntesis química , Sitios de Unión , Diseño de Fármacos , Modelos Moleculares , Relación Estructura-ActividadRESUMEN
G-quadruplexes (G4s) formed by guanine-rich nucleic acids induce genome instability through impeding DNA replication fork progression. G4s are stable DNA structures, the unfolding of which require the functions of DNA helicases. Pif1 helicase binds preferentially to G4 DNA and plays multiple roles in maintaining genome stability, but the mechanism by which Pif1 unfolds G4s is poorly understood. Here we report the co-crystal structure of Saccharomyces cerevisiae Pif1 (ScPif1) bound to a G4 DNA with a 5' single-stranded DNA (ssDNA) segment. Unlike the Thermus oshimai Pif1-G4 structure, in which the 1B and 2B domains confer G4 recognition, ScPif1 recognizes G4 mainly through the wedge region in the 1A domain that contacts the 5' most G-tetrad directly. A conserved Arg residue in the wedge is required for Okazaki fragment processing but not for mitochondrial function or for suppression of gross chromosomal rearrangements. Multiple substitutions at this position have similar effects on resolution of DNA duplexes and G4s, suggesting that ScPif1 may use the same wedge to unwind G4 and dsDNA. Our results reveal the mechanism governing dsDNA unwinding and G4 unfolding by ScPif1 helicase that can potentially be generalized to other eukaryotic Pif1 helicases and beyond.
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ADN Helicasas , G-Cuádruplex , Proteínas de Saccharomyces cerevisiae , Saccharomyces cerevisiae , ADN Helicasas/metabolismo , ADN Helicasas/química , ADN Helicasas/genética , Proteínas de Saccharomyces cerevisiae/metabolismo , Proteínas de Saccharomyces cerevisiae/química , Proteínas de Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo , ADN/metabolismo , ADN/química , ADN/genética , ADN de Cadena Simple/metabolismo , ADN de Cadena Simple/química , Cristalografía por Rayos X , Modelos Moleculares , Unión Proteica , Replicación del ADN , Inestabilidad GenómicaRESUMEN
Arsenic (As) from mine wastewater is a significant source for acidic paddy soil pollution, and its mobility can be influenced by alternating redox conditions. However, mechanistic and quantitative insights into the biogeochemical cycles of exogenous As in paddy soil are still lacking. Herein, the variations of As species in paddy soil spiking with As(III) or As(V) were investigated in the process of 40 d of flooding followed 20 d of drainage. During flooding process, available As was immobilized in paddy soil spiking As(III) and the immobilized As was activated in paddy soil spiking As(V) owing to deprotonation. The contributions of Fe oxyhydroxides and humic substances (HS) to As immobilization in paddy soil spiking As(III) were 80.16% and 18.64%, respectively. Whereas the contributions of Fe oxyhydroxides and HS to As activation in paddy soil spiking As(V) were 47.9% and 52.1%, respectively. After entering drainage, available As was mainly immobilized by Fe oxyhydroxides and HS and adsorbed As(III) was oxidized. The contribution of Fe oxyhydroxides to As fixation in paddy soil spiking As(III) and As(V) was 88.82% and 90.26%, respectively, and of HS to As fixation in paddy soil spiking As(III) and As(V) was 11.12% and 8.95%, respectively. Based on the model fitting results, the activation of Fe oxyhydroxides and HS bound As followed with available As(V) reduction were key processes during flooding. This may be because the dispersion of soil particles and release of soil colloids activated the adsorbed As. Immobilization of available As(III) by amorphous Fe oxyhydroxides followed with adsorbed As(III) oxidation were key processes during drainage. This may be ascribe to the occurrence of coprecipitation and As(III) oxidation mediated by reactive oxygen species from Fe(II) oxidation. The results are beneficial for a deeper understanding of As species transformation at the interface of paddy soil-water as well as an estimation pathway for the impacts of key biogeochemical cycles on exogenous As species under a redox-alternating condition.
Asunto(s)
Arsénico , Oryza , Contaminantes del Suelo , Sustancias Húmicas , Arsénico/química , Suelo/química , Hierro/química , Contaminantes del Suelo/análisis , Oxidación-Reducción , Oryza/metabolismoRESUMEN
Telomeres are specialized nucleoprotein structures at the ends of linear chromosomes. The progressive shortening of steady-state telomere length in normal human somatic cells is a promising biomarker for age-associated diseases. However, there remain substantial challenges in quantifying telomere length due to the lack of high-throughput method with nucleotide resolution for individual telomere. Here, we describe a workflow to capture telomeres using newly designed telobaits in human culture cell lines as well as clinical patient samples and measure their length accurately at nucleotide resolution using single-molecule real-time (SMRT) sequencing. Our results also reveal the extreme heterogeneity of telomeric variant sequences (TVSs) that are dispersed throughout the telomere repeat region. The presence of TVSs disrupts the continuity of the canonical (5'-TTAGGG-3')n telomere repeats, which affects the binding of shelterin complexes at the chromosomal ends and telomere protection. These findings may have profound implications in human aging and diseases.
Asunto(s)
Complejo Shelterina , Telómero , Humanos , Telómero/genética , EnvejecimientoRESUMEN
Proprotein convertase subtilisin/kexin type 9 (PCSK9) is a secretory serine protease synthesized primarily by the liver. It mainly promotes the degradation of low-density lipoprotein receptor (LDL-R) by binding LDL-R, reducing low-density lipoprotein cholesterol (LDL-C) clearance. In addition to regulating LDL-R, PCSK9 inhibitors can also bind Toll-like receptors (TLRs), scavenger receptor B (SR-B/CD36), low-density lipoprotein receptor-related protein 1 (LRP1), apolipoprotein E receptor-2 (ApoER2) and very-low-density lipoprotein receptor (VLDL-R) reducing the lipoprotein concentration and slowing thrombosis. In addition to cardiovascular diseases, PCSK9 is also used in pancreatic cancer, sepsis, and Parkinson's disease. Currently marketed PCSK9 inhibitors include alirocumab, evolocumab, and inclisiran, as well as small molecules, nucleic acid drugs, and vaccines under development. This review systematically summarized the application, preclinical studies, safety, mechanism of action, and latest research progress of PCSK9 inhibitors, aiming to provide ideas for the drug research and development and the clinical application of PCSK9 in cardiovascular diseases and expand its application in other diseases.
Asunto(s)
Enfermedades Cardiovasculares , Ácidos Nucleicos , Enfermedades Cardiovasculares/tratamiento farmacológico , Enfermedades Cardiovasculares/prevención & control , LDL-Colesterol/metabolismo , Humanos , Lipoproteínas VLDL , Proteína 1 Relacionada con Receptor de Lipoproteína de Baja Densidad , Proproteína Convertasa 9/metabolismo , SubtilisinasRESUMEN
Extensive studies have been devoting to investigating the catalytic efficiency of zero-valent iron (Fe0)-based bimetals with persulfate (PS), while little is known in the stoichiometric efficiency, underlying mechanisms and reaction center of zero-valent bimetallic catalysts in activating PS. Herein, nanoscale zero-valent Fe/Cu catalysts in decomposing 2,4-dichlorophenol (DCP) have been investigated. The results show that the increase of Cu ratio from 0 to 0.75 significantly enhances the DCP degradation with a rate constant of 0.025 min-1 for Fe0 to 0.097 min-1 for Fe/Cu(0.75) at pH â¼3.3, indicating Cu is likely the predominate reaction centers over Fe. The PS decomposition is reduced with the increase of Cu ratios, suggesting the stoichiometric efficiency of Fe/Cu in activating PS is notably enhanced from 0.024 for Fe0 to 0.11 for Fe/Cu(0.75). Analyses indicate Cu atoms are likely the predominant reaction site for DCP decomposition, and Fe atoms synergistically enhance the activity of Cu as indicated by DFT calculations. Both SO4â¦- and â¦OH radicals are responsible for reactions, and the contribution of SO4â¦- is decreased at higher pH conditions. The findings of this work provide insight into the stoichiometric efficiency and the reaction center of Fe/Cu catalysts to activate PS for pollutant removals.
RESUMEN
Invasive fungal infections are accompanied by high mortality rates that range up to 90%. At present, only three different compound classes are available for use in the clinic, and these often suffer from low bioavailability, toxicity, and drug resistance. These issues emphasize an urgent need for novel antifungal agents. Herein, we report the identification of chemically versatile benzamide and picolinamide scaffolds with antifungal properties. Chemogenomic profiling and biochemical assays with purified protein identified Sec14p, the major phosphatidylinositol/phosphatidylcholine transfer protein in Saccharomyces cerevisiae, as the sole essential target for these compounds. A functional variomics screen identified resistance-conferring residues that localized to the lipid-binding pocket of Sec14p. Determination of the X-ray co-crystal structure of a Sec14p-compound complex confirmed binding in this cavity and rationalized both the resistance-conferring residues and the observed structure-activity relationships. Taken together, these findings open new avenues for rational compound optimization and development of novel antifungal agents.
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Antifúngicos/metabolismo , Benzamidas/química , Ácidos Picolínicos/química , Amidas/química , Amidas/metabolismo , Amidas/farmacología , Secuencia de Aminoácidos , Antifúngicos/química , Antifúngicos/farmacología , Aspergillus/efectos de los fármacos , Benzamidas/metabolismo , Benzamidas/farmacología , Sitios de Unión , Candida albicans/efectos de los fármacos , Cristalografía por Rayos X , Farmacorresistencia Fúngica/efectos de los fármacos , Pruebas de Sensibilidad Microbiana , Simulación de Dinámica Molecular , Mutagénesis Sitio-Dirigida , Proteínas de Transferencia de Fosfolípidos/química , Proteínas de Transferencia de Fosfolípidos/genética , Proteínas de Transferencia de Fosfolípidos/metabolismo , Ácidos Picolínicos/metabolismo , Ácidos Picolínicos/farmacología , Estructura Terciaria de Proteína , Saccharomyces cerevisiae/efectos de los fármacos , Saccharomyces cerevisiae/metabolismo , Proteínas de Saccharomyces cerevisiae/química , Proteínas de Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/metabolismo , Alineación de Secuencia , Relación Estructura-ActividadRESUMEN
Plasminogen activator inhibitor-1 (PAI-1), a serpin, is the physiological inhibitor of tissue-type and urokinase-type plasminogen activators and thus also an inhibitor of fibrinolysis and tissue remodeling. It is a potential therapeutic target in many pathological conditions, including thrombosis and cancer. Several types of PAI-1 antagonist have been developed, but the structural basis for their action has remained largely unknown. Here we report X-ray crystal structure analysis of PAI-1 in complex with a small-molecule antagonist, embelin. We propose a mechanism for embelin-induced rapid conversion of PAI-1 into a substrate for its target proteases and the subsequent slow conversion of PAI-1 into an irreversibly inactivated form. Our work provides structural clues to an understanding of PAI-1 inactivation by small-molecule antagonists and an important step toward the design of drugs targeting PAI-1.