Experimental and Numerical Simulation Research on Controllable Shock Wave-Induced Shale Fracturing under Repeated Action.

Low permeability is a key geological factor constraining the development of shale gas, and reservoir modification to improve its permeability is a prerequisite. Controlled shock wave fracturing can induce the formation of complex fractures in reservoirs and is expected to become an important means of reservoir modification. However, the mechanism of controlled shock wave fracturing in shale and the geological engineering control factors are unclear. Therefore, this article reveals the mechanism and effect of shock wave modification through small-scale experiments and large-scale numerical simulations. Results show that as the impact number increases, a significant increase in large fractures and fracture connectivity within the shale samples is observed, while the correlation between the geometric parameters of the fractures and the number of impacts is weak. High-energy input in the model will cause a larger range of damage to the rock, accompanied by a smaller attenuation index, indicating that the speed of energy attenuation plays a decisive role in rock damage. The influence of crustal stress is greater than the speed of energy attenuation, and higher crustal stress will inhibit the formation of fractures. A moderate increase in the number of controllable shock waves is beneficial for the fracturing effect; however, further increasing the loading number of controllable shock waves will weaken the strengthening effect of the fracturing effect.

Tetrazine-Isonitrile Bioorthogonal Fluorogenic Reactions Enable Multiplex Labeling and Wash-Free Bioimaging of Live Cells.

Developing fluorogenic probes for simultaneous live cell labeling of multiple targets is crucial for understanding complex cellular events. The emerging [4+1] cycloaddition between tetrazine and isonitriles holds promise as a bioorthogonal tool, yet existing tetrazine probes lack reactivity and fluorogenicity. Here, we present the development of a series of tetrazine-functionalized bioorthogonal probes. By incorporating pyrazole adducts into the fluorophore scaffolds, the post-reacted probes displayed remarkable fluorescence turn-on ratios, up to 3184-fold. Moreover, these modifications are generalizable to various fluorophores, enabling a broad emission range from 473 to 659 nm. Quantum chemical calculations further elucidate the turn-on mechanisms. These probes enable the simultaneous labeling of multiple targets in live cells, without the need for a washing step. Consequently, our findings pave the way for advanced multiplex imaging and detection techniques for cellular studies.

Design of SC PEP with enhanced stability against pepsin digestion and increased activity by machine learning and structural parameters modeling.

Prolyl endopeptidases from Sphingomonas capsulata (SC PEP) has attracted much attention as promising oral therapy candidate for celiac sprue, however, its low stability in the gastric environment leads to unsatisfactory clinical results. Therefore, improving its stability against pepsin digestion at low pH is crucial for clinical applications, but challenging. In this study, machine learning and physical parameter model were combined to design SC PEP mutants. After iterations, 20 mutants had higher hydrolysis activity in stomach environment, which was up to 14.1-fold compared with wild-type SC PEP. Mutant M24 involving stable and active mutations and pegylated M24 (M24-PEG) had higher activity of hydrolyzing immunogen in bread than wild-type SC PEP in vitro and in vivo, and residual immunogens in simulated gastric environment were only 1/8 and 1/10 of that in the wild-type SC PEP group. The total residual immunogens in the gastrointestinal tract of mice in the M24 and M24-PEG groups were <20 ppm, reaching the standard of non-toxic food. Our results indicate that the combination of M24 (or M24-PEG) with EP-B2 may be a promising candidate for celiac disease, and the strategies developed in this study provide a paradigm for the design of SC PEP stability mutants.

A General and Modular Approach to BCP Alkylamines via Multicomponent Difunctionalization of [1.1.1]Propellane.

Over the past decade, bicyclo[1.1.1]pentane (BCP) motifs have come to the fore as valuable pharmaceutical bioisosteres of para-disubstituted benzenes. However, the limited approaches and requisite multistep syntheses of useful BCP building blocks are hampering early discovery research in medicinal chemistry. Herein we report the development of a modular strategy for the divergent preparation of functionalized BCP alkylamines. In this process, a general method to introduce fluoroalkyl groups to BCP scaffolds using readily available and easy-to-handle fluoroalkyl sulfinate salts was also developed. Moreover, this strategy can also be extended to S-centered radicals for incorporation of sulfones and thioethers into the BCP core. Overall, this multicomponent strategy enables rapid construction of BCP-type bioisosteres for applications in drug discovery.

Production of Recombinant Human Hybrid Ferritin with Heavy Chain and Light Chain in Escherichia coli and its Characterization.

BACKGROUND: Natural human ferritin generally contains 24 subunits with different ratios of heavy chain to light chain, and the ratio of both subunits varies depending on tissue distribution and pathological conditions. However, the production of recombinant hybrid ferritin with both subunits is more challenging. OBJECTIVE: This study aimed to prepare the recombinant hybrid ferritin for prokaryotic expression and characterize its structure and physicochemical properties. METHODS: A prokaryotic expression vector of pACYCDuet-1 harboring the two individual genes of human ferritin heavy chain and light chain (FTH/FTL-pACYCDuet-1) was constructed and transfected into Escherichia coli bacteria. Then the genes were co-induced by IPTG to express. RESULTS: The ferritin was purified by hydrophobic interaction chromatography combining size exclusion chromatography and verified by mass spectrometry and characterized by spectral and morphological analysis. CONCLUSION: FTH and FTL subunits were successfully co-assembled into a hybrid ferritin nanoparticle (rhFTH/L). The structure of rhFTH/L was demonstrated highly ordered and fairly compact. Besides, the hybrid rhFTH/L nanoparticle was shown more sensitive to thermal stress and reduced stability when compared with that of both individual rhFTH and rhFTL.

Photochemical Intermolecular [3σ + 2σ]-Cycloaddition for the Construction of Aminobicyclo[3.1.1]heptanes.

The development of synthetic strategies for the preparation of bioisosteric compounds is a demanding undertaking in medicinal chemistry. Numerous strategies have been developed for the synthesis of bicyclo[1.1.1]pentanes (BCPs), bridge-substituted BCPs, and bicyclo[2.1.1]hexanes. However, progress on the synthesis of bicyclo[3.1.1]heptanes, which serve as meta-substituted arene bioisosteres, has not been previously explored. Herein, we disclose the first photoinduced [3σ + 2σ] cycloaddition for the synthesis of trisubstituted bicyclo[3.1.1]heptanes using bicyclo[1.1.0]butanes and cyclopropylamines. This transformation not only uses mild and operationally simple conditions but also provides unique meta-substituted arene bioisosteres. The applicability of this method is showcased by simple derivatization reactions.

Milk-derived exosomes exhibit versatile effects for improved oral drug delivery.

As endogenous courier vesicles, exosomes play crucial roles in macromolecule transmission and intercellular communication. Therefore, exosomes have drawn increasing attention as biomimetic drug-delivery vehicles over the past few years. However, few studies have investigated the encapsulation of peptide/protein drugs into exosomes for oral administration. Additionally, the mechanisms underlying their biomimetic properties as oral delivery vehicles remain unknown. Herein, insulin-loaded milk-derived exosomes (EXO@INS) were fabricated and the in vivo hypoglycemic effect was investigated on type I diabetic rats. Surprisingly, EXO@INS (50 and 30 IU/kg) elicited a more superior and more sustained hypoglycemic effect compared with that obtained with subcutaneously injected insulin. Further mechanism studies indicated that the origin of excellent oral-performance of milk-derived exosomes combined active multi-targeting uptake, pH adaptation during gastrointestinal transit, nutrient assimilation related ERK1/2 and p38 MAPK signal pathway activation and intestinal mucus penetration. This study provides the first demonstration that multifunctional milk-derived exosomes offer solutions to many of the challenges arising from oral drug delivery and thus provide new insights into developing naturally-equipped nanovehicles for oral drug administration.

Conversion of recombinant human ferritin light chain inclusion bodies into uniform nanoparticles in Escherichia coli for facile production.

Prokaryotic expression systems are widely used to produce many types of biologics because of their extreme conveniences and unmatchable cost. However, production of recombinant human ferritin light chain (rhFTL) protein is largely restrained because its expression in Escherichia coli tends to form inclusion bodies (IBs). In this study, a prokaryotic expression vector (FTL-pBV220) harboring the rhFTL gene was constructed using a pBV220 plasmid. The tag-free rhFTL was highly expressed and almost entirely converted to soluble form, and thus the rhFTL was successfully self-assembled into uniform nanoparticles in E. coli. To establish a simplified downstream process, a precipitation procedure based on the optimized incubation temperature, pH condition, and ionic strength was developed to remove impurities from the crude lysate supernatant. The rhFTL retained in the clarified supernatant was subsequently purified in a single step using Capto Butyl column resulting in a considerable recovery and high purity. The purified rhFTL was characterized and verified by mass spectrometry and spectral and morphological analyses. The results revealed that rhFTL exhibited highly ordered and fairly compact structures and the spherical structures were preserved.

Facile production of tag-free recombinant human interleukin-11 by transforming into soluble expression in Escherichia coli.

Human interleukin-11 (IL-11) is considered as a difficult-to-express protein in prokaryotic expression systems because of its low expression level and high tendency to form inclusion bodies. The current source of recombinant human IL-11 (rhIL-11) for therapeutic use is mainly obtained from a fusion protein synthesized by Escherichia coli, which requires an additional operation to cleave the fusion tag. Herein, we reported a strategy for the direct expression of tag-free rhIL-11 in E. coli. To explore the soluble expression of rhIL-11 without fusion tags in E. coli, we inserted the rhIL-11 gene into a pBV220 plasmid which is characterized by employing a temperature-sensitive pR/pL promoter to manipulate the transcription and translation of the gene of interest. As a result, the tag free rhIL-11 was efficiently expressed in the soluble form in E. coli. A two-step chromatography method, Capto Butyl-S combined with Capto Q, was developed to efficiently purify the tag-free rhIL-11 from the supernatant of the cell lysate. The resultant rhIL-11 showed a compact and highly ordered structure, as validated by circular dichroism and intrinsic fluorescence emission spectra. Additionally, the biological activity of the purified rhIL-11 was evaluated by TF-1 cell proliferation experiments and the results demonstrated that the E. coli expressed rhIL-11 is biologically active.

Novel thiophene-based donor-acceptor scaffolds as cathodes for rechargeable aqueous zinc-ion hybrid supercapacitors.

Well-defined π-conjugated thiophene donor-acceptor molecules play an important role in different fields ranging from synthetic chemistry to materials science. Their chemical structure provides specific electronic and physicochemical properties, which can be further tuned by the introduction of functional groups. Herein, we design and synthesize two novel thiophene-based π-conjugated donor-acceptor molecules TDA-1 and TDA-2 through Aldol and Knoevenagel condensations. In our proof-of-concept study we report for the first time on the use of small organic molecules employed in aqueous zinc-ion hybrid supercapacitors (Zn-HSCs),which exhibit capacitance as high as 206.7 and 235.2 F g-1 for TDA-1, and TDA-2, respectively.

One-Pot anti-Michael Regio- and Stereoselective Hydroamination of Activated N-Allenamides.

N-Allenamides, substituted by an ester at the γ-position, were obtained through addition of terminal ynamides with ethyl diazoacetate under copper catalysis for the first time. Regio- and stereoselective hydroamination of those activated N-allenamides provided exclusively E-configured captodative enamimes through a one-pot anti-Michael addition. Numerous ynamides as well as various secondary amines were adapted in this process.

Copper-Catalyzed Synthesis of Terminal vs. Fluorine-Substituted N-Allenamides via Addition of Diazo Compounds to Terminal Ynamides.

A copper-mediated coupling reaction between ynamides and diazo-compounds to produce N-allenamides is reported for the first time. This method enables facile and rapid access to terminal N-allenamides by using commercially available TMS-diazomethane with wide functional group compatibility on the nitrogen. Furthermore, the ubiquity of molecules containing a fluorine moiety in medicine, in agricultural, and material science requires the continuous search of new building blocks, including this unique surrogate. The CuI/diazo protocol was successfully applied to the synthesis of fluorine-substituted N-allenamides. DFT calculations provided insights in the mechanism involved.

Design SMAP29-LysPA26 as a Highly Efficient Artilysin against Pseudomonas aeruginosa with Bactericidal and Antibiofilm Activity.

Antimicrobial resistance (AMR) is a major issue to global health. The multidrug-resistant (MDR) Gram-negative infections, particularly infected by carbapenem-resistant pathogens, urgently need efficient antibiotics and novel therapy. However, the scientific challenges of aiming for innovative approaches against Gram-negative bacteria have hindered the research and development of antibiotic drugs. Phage-derived endolysins are bacteriolytic and specific for a bacterial species or genus, providing a promising antibiotic strategy. However, the outer membrane of Gram-negative bacteria could prevent the peptidoglycan layer from the hydrolysis of endolysins. Antimicrobial peptides usually destabilize the outer membrane and could enhance the antibiotic activity of endolysins. In this study, we designed new artilysins with antimicrobial-peptide SMAP29 fusion at the N-terminal of LysPA26 (named as AL-3AA, AL-9AA, and AL-15AA), and evaluated them. The results showed artilysin AL-3AA to be highly bactericidal; even 0.05 mg/mL AL-3AA could reduce 5.81 log units P. aeruginosa without EDTA in 60 min. It killed P. aeruginosa rapidly and dose-dependently through cell lysis. AL-3AA inhibited P. aeruginosa PAO1 biofilm formation and significantly decreased mature P. aeruginosa biofilms. It also had potential broad-spectrum activity against susceptible Gram-negative bacteria in the hospital, including K. pneumoniae and E. coli. The antibacterial mechanism investigation has provided valuable information about the antibacterial action of AL-3AA, which can lyse and disintegrate the bacterial quickly. These results suggested AL-3AA could be a new and promising antimicrobial agent for the combat of P. aeruginosa. IMPORTANCE Antimicrobial resistance (AMR) is a major issue to global health, particularly the multidrug-resistant (MDR) Gram-negative infections, which pose great challenges. Even new antibiotics research is ongoing, antibiotics used to treat Gram-negative bacteria in the clinical are limited in a small set of molecular scaffolds, and biomolecular categories of antibiotics are urgently needed. In this study, we designed new proteins by combining antimicrobial peptides and endolysins for synergistic bactericidal effects. One of designed proteins, named AL-3AA, showed highly bactericidal, and killed P. aeruginosa rapidly and dose-dependently through cell lysis. It also killed Klebsiella pneumoniae and Escherichia coli, showing potential broad-spectrum activity against susceptible Gram-negative bacteria in the hospital. All results suggest AL-3AA could be a new and promising antimicrobial agent for the combat of P. aeruginosa.

Quantum frequency conversion from ultraviolet to visible band through waveguides in a period-poled MgO:LiTaO3 crystal.

In research on hybrid quantum networks, visible or near-infrared frequency conversion has been realized. However, technical limitations mean that there have been few studies involving the ultraviolet band, and unfortunately the wavelengths of the rare-earth or alkaline-earth metal atoms or ions that are used widely in research on quantum information are often in the UV band. Therefore, frequency conversion of the ultraviolet band is very important. In this paper, we demonstrate a quantum frequency conversion between ultraviolet and visible wavelengths by fabricating waveguides in a period-poled MgO:LiTaO3 crystal with a laser writing system, which will be used to connect the wavelength of the dipole transition of 171Yb+ at 369.5 nm and the absorption wavelength of Eu3+ at 580 nm in a solid-state quantum memory system. An external conversion efficiency of 0.85% and a signal-to-noise ratio of greater than 500 are realized with a pumping power of 3.28 W at 1018 nm. Furthermore, we complete frequency conversion of the classical polarization state by means of a symmetric optical setup based on the fabricated waveguide, and the process fidelity of the conversion is (96.13 ± 0.021)%. This converter paves the way for constructing a hybrid quantum network and realizing a quantum router in the ultraviolet band in the future.

Large-tuning-range frequency stabilization of an ultraviolet laser by an open-loop piezoelectric ceramic controlled Fabry-Pérot cavity.

We demonstrate a laser frequency stabilization method with large tuning range to stabilize a UV laser by installing piezoelectric ceramic actuators into a Fabry-Pérot cavity with an ultra-low expansion spacer. To suppress piezoelectric drift, a two-layer symmetrical structure is adopted for the piezoelectric actuator, and a 14.7 GHz tuning range is achieved. The short-term drift of the piezoelectric ceramics caused by temperature and creep is eliminated, and the long-term drift is 0.268 MHz/h when the Fabry-Pérot cavity is sealed in a chamber without a vacuum environment. The long-term frequency drift is mainly caused by stress release and is eliminated by compensating the cavity voltage with an open loop. Without the need for an external reference or a vacuum environment, the laser frequency stabilization system is greatly simplified, and it can be extended to wavelengths ranging from ultraviolet to infrared. Owing to its simplicity, stability, and large tuning range, it is applicable in cold atom and trapped ion experiments.

Design BH3 domain fusion protein as targeting pro-apoptotic self-assembling nanoparticles.

Cancer is a serious global health issue, and apoptosis is a logical and practical cancer therapeutic strategy. Apoptosis responses to internal and external signals. Both BH3 domain in the pro-apoptotic proteins and truncated BH3 domain can stimulate cell apoptosis. However, the faults of peptides in systemic administration restrict the applications of truncated BH3 domain. Ferritin, as an attractive nanoparticle with the capacity of self-assemble to unique hollow spherical structure, could display truncated BH3 domain an N-terminal. Thus, in this study, we designed a pro-apoptosis self-assembling protein nanoparticle by BH3 domain fusion at N-terminal of ferritin. We evaluated the size, cytotoxicity and pro-apoptosis effect of these nanoparticles. The results showed that RGD-BH3-HFn, BH3-HFn and HFn had uniformly spherical structure with sizes at 26.08 ± 0.11 nm, 22.07 ± 0.67 nm, and 16.81 ± 0.88 nm, respectively; RGD-BH3-HFn has stronger cytotoxicity against tumor cells than BH3-HFn and HFn. The total apoptosis ratios (including necrosis) of C6 cells induced by RGD-BH3-HFn, BH3-HFn, and HFn proteins were 15.24%, 10.13% and 2.14%, respectively; those of bEnd.3 cells were 15.47%, 7.33% and 1.70%, respectively; while the total apoptosis rate (including necrosis) of MCF-7 cells were 3.24%, 4.9% and - 1.68%, respectively. The results suggested self-assembling RGD-BH3-HFn could target to C6 cells and bEnd.3 cells, and enhance tumor cells apoptosis, its apoptosis effect against C6 cells was 7.11-fold that of HFn, and apoptosis effect against bEnd.3 cells was 9.08-fold that of HFn. These results indicated BH3 domain can be designed as targeting pro-apoptotic nanoparticles.

Direct Synthesis of CF2H-Substituted 2-Amidofurans via Copper-Catalyzed Addition of Difluorinated Diazoacetone to Ynamides.

The significance of molecules containing difluoromethyl groups is driven by their potential applications in pharmaceutical and agrochemical science. Methods for the incorporation of lightly fluorinated groups such as CF2H have been less well developed. Here we report the use of difluorinated diazoacetone as a practical reagent for the direct synthesis of CF2H-substituted 2-amidofurans through addition to ynamides. These newly designed difluorinated amidofurans were elaborated to create new nitrogen-containing frameworks that would be challenging to obtain otherwise.

Alpha-asarone Improves Cognitive Function of APP/PS1 Mice and Reducing Aß42, P-tau and Neuroinflammation, and Promoting Neuron Survival in the Hippocampus.

Alzheimer's disease (AD) is a progressive neurodegenerative disease most often characterized by memory impairment and cognitive decline. Alpha-asarone has been reported to have the potential to treat AD. Our previous studies have found that alpha-asarone improves aged rats' cognitive function by alleviating neuronal excitotoxicity via type A gamma-aminobutyric acid (GABA) receptors. GABA level's change, neuroinflammation, and dysfunctional autophagy are found to be associated with AD. However, the effect of alpha-asarone on cognitive function of APP/PS1 transgenic mice and its underlying mechanism in terms of aggregation of amyloid-ß42 (Aß42) and phosphorylated tau (p-tau), glutamic acid decarboxylase (GAD) level, neuroinflammation, and autophagy are unclear. Accordingly, we attempted to explore whether alpha-asarone improves AD mice's cognitive function and alleviates pathological symptoms by regulating GAD level, inhibiting neuroinflammation, or restore autophagy. We found that alpha-asarone enhanced spatial learning memory and decreased Aß42 and p-tau levels without influencing the GAD level in APP/PS1 transgenic mice. Also, it decreased the GFAP expression and reduced pro-inflammatory cytokines levels, thus alleviating neuroinflammation. Furthermore, alpha-asarone decreased the excess number of autophagosomes and promoted hippocampal neurons' survival. In conclusion, the results confirmed the therapeutic effect of alpha-asarone on AD-related astrogliosis, dysfunctional autophagy, and neuronal damage, which indicates its great potential to treat AD.

Super-resolved Imaging of a Single Cold Atom on a Nanosecond Timescale.

In cold atomic systems, fast and high-resolution microscopy of individual atoms is crucial, since it can provide direct information on the dynamics and correlations of the system. Here, we demonstrate nanosecond-scale two-dimensional stroboscopic pictures of a single trapped ion beyond the optical diffraction limit, by combining the main idea of ground-state depletion microscopy with quantum-state transition control in cold atoms. We achieve a spatial resolution up to 175 nm using a NA=0.1 objective in the experiment, which represents a more than tenfold improvement compared with direct fluorescence imaging. To show the potential of this method, we apply it to observe the secular motion of the trapped ion; we demonstrate a temporal resolution up to 50 ns with a displacement detection sensitivity of 10 nm. Our method provides a powerful tool for probing particle positions, momenta, and correlations, as well as their dynamics in cold atomic systems.

Innate promiscuity of the CYP706 family of P450 enzymes provides a suitable context for the evolution of dinitroaniline resistance in weed.

Increased metabolism is one of the main causes for evolution of herbicide resistance in weeds, a major challenge for sustainable food production. The molecular drivers of this evolution are poorly understood. We tested here the hypothesis that a suitable context for the emergence of herbicide resistance could be provided by plant enzymes with high innate promiscuity with regard to their natural substrates. A selection of yeast-expressed plant cytochrome P450 enzymes with well documented narrow to broad promiscuity when metabolizing natural substrates was tested for herbicide metabolism competence. The positive candidate was assayed for capacity to confer herbicide tolerance in Arabidopsis thaliana. Our data demonstrate that Arabidopsis thaliana CYP706A3, with the most promiscuous activity on monoterpenes and sesquiterpenes for flower defence, can also oxidize plant microtubule assembly inhibitors, dinitroanilines. Ectopic overexpression of CYP706A3 confers dinitroaniline resistance. We show, in addition, that the capacity to metabolize dinitroanilines is shared by other members of the CYP706 family from plants as diverse as eucalyptus and cedar. Supported by three-dimensional (3D) modelling of CYP706A3, the properties of enzyme active site and substrate access channel are discussed together with the shared physicochemical properties of the natural and exogenous substrates to explain herbicide metabolism.