Natural biomolecules have been used extensively as chiral scaffolds that bind/surround metal complexes to achieve stereoselectivity in catalytic reactions. ATP is ubiquitously found in nature as an ...energy-storing molecule and can complex diverse metal cations. However, in biotic reactions ATP-metal complexes are thought to function mostly as co-substrates undergoing phosphoanhydride bond cleavage reactions rather than participating in catalytic mechanisms. Here, we report that a specific Cu(II)-ATP complex (Cu
·ATP) efficiently catalyses Diels-Alder reactions with high reactivity and enantioselectivity. We investigate the substrates and stereoselectivity of the reaction, characterise the catalyst by a range of physicochemical experiments and propose the reaction mechanism based on density functional theory (DFT) calculations. It is found that three key residues (N7, β-phosphate and γ-phosphate) in ATP are important for the efficient catalytic activity and stereocontrol via complexation of the Cu(II) ion. In addition to the potential technological uses, these findings could have general implications for the chemical selection of complex mixtures in prebiotic scenarios.
The diverse structures of DNA serve as potent chiral scaffolds for DNA-based asymmetric catalysis, yet in most cases tens to hundreds of nucleotides in DNA hybrid catalysts hinder the deep insight ...into their structure-activity relationship. Owing to the structural simplicity and design flexibility of nucleotides, nucleotide-based catalysts have been emerging as a promising way to obtain fine structural information and understand the catalytic mechanisms. Herein, we found that a cyclic dinucleotide of cyclic di-AMP (c-di-AMP) and 1,10-phenanthroline copper(II) nitrate (Cu(phen)(NO
)
) are assembled to a c-di-AMP-based catalyst (c-di-AMP/Cu(phen)(NO
)
), which could fast achieve enantioselective fluorination in water with 90-99% yields and up to 90% enantiomeric excess (ee). The host-guest interaction between c-di-AMP and Cu(phen)(NO
)
has been proposed mainly in a supramolecular interaction mode as evidenced by spectroscopic techniques of ultraviolet-visible, fluorescence, circular dichroism, and nuclear magnetic resonance. Cu(phen)(NO
)
tightly binds to c-di-AMP with a binding constant of 1.7 ± 0.3 × 10
M
, and the assembly of c-di-AMP/Cu(phen)(NO
)
shows a modest rate enhancement to carbon-fluorine bond formations as supported by kinetic studies.
The diverse structures of nucleic acids as scaffolds have brought the significant advancement for DNA‐based enantioselective catalysis, yet RNA‐based enantioselective catalysis is lacking ...investigation. Herein, we report a small, natural RNA of cyclic di‐AMP (c‐di‐AMP) and Cu2+ ions assemble into an artificial metalloribozyme (c‐di‐AMP·Cu2+), that could effectively catalyze the enantioselective Diels–Alder reactions with up to 80 % ee. The enantioselective catalytic performance of c‐di‐AMP·Cu2+ has been studied by thorough investigations of different metal cofactors, c‐di‐AMP/Cu2+ molar ratios, additives, buffers and c‐di‐AMP analogues. In addition, the assembly of c‐di‐AMP·Cu2+ gives rise to 300‐fold and 5‐fold rate acceleration compared to the uncatalyzed reaction and Cu2+ ions, respectively. This work provides a simple and efficient strategy to construct the RNA‐based catalysts that would expand the current nucleic acids‐based catalysis and might hint the possible catalytic RNA in primordial chemistry.
A small cyclic RNA of c‐di‐AMP and Cu2+ ions could assemble into an artificial metalloribozyme, that is able to catalyze the enantioselective Diels–Alder reactions with up to 80 % ee. The presence of c‐di‐AMP in the metalloribozyme not only achieves the chirality transfer but also gives rise to a significant rate enhancement.
is a foodborne pathogen that can colonize the small intestine of the host and cause diarrhea. The alternative sigma factor RpoN plays a vital role in regulating motility, carbon utilization and ...affects host colonization in
RIMD2210633. In this study, transcriptome and phenotypic analysis further expanded our understanding of the RpoN regulon in
. A deletion mutant of
(Δ
) was subjected to RNA-seq for systemic identification of the RpoN-controlled genes. Compared with the wild-type (WT), 399 genes were differentially expressed in the Δ
strain. Moreover, 264 genes were down-regulated in the Δ
strain, including those associated with nitrogen utilization (
), glutamine synthetase (
), formate dehydrogenase (
and
-
), quorum sensing (
and
), polar flagellar systems, and type VI secretion system 2 (T6SS2). Quantitative real-time reverse transcription PCR (qRT-PCR) and electrophoretic mobility shift assay (EMSA) further confirmed that RpoN could directly bind to the promoters of these genes associated with polar flagellar systems (
and
), lateral flagellar systems (
and
), T6SS2 (
and
) and glutamine synthetase (
), and then positively regulate the expression of these systems. A RpoN-binding motif was identified in
using the MEME suite and verified by the EMSA. Besides, the deletion of
caused a significant decrease in hemolytic activity, adhesion, and cytotoxicity. Our results provide new cues to better understand the regulatory networks of RpoN protein to motility, T6SS2, and metabolism in
.
is a moderately halophilic foodborne pathogen that is mainly distributed in marine and freshwater environments. The transition of
between aquatic ecosystems and hosts is essential for infection. Both ...freshwater and host environments have low salinity. In this study, we sought to further investigate the effects of low salinity (0.5% NaCl) on the fitness and virulence of
. We found that
could survive in Luria-Bertani (LB) and M9 mediums with different NaCl concentrations, except for the M9 medium containing 9% NaCl. Our results further showed that
cultured in M9 medium with 0.5% NaCl had a higher cell density than that cultured at other NaCl concentrations when it entered the stationary phase. Therefore, we compared the transcriptomes of
wild type (WT) cultured in an M9 medium with 0.5% and 3% NaCl at the stationary phase using RNA-seq. A total of 658 genes were significantly differentially expressed in the M9 medium with 0.5% NaCl, including regulators, osmotic adaptive responses (compatible solute synthesis systems, transporters, and outer membrane proteins), and virulence factors (T3SS1 and T6SS1). Furthermore, a low salinity concentration in the M9 medium induced the expression of T3SS1 to mediate the cytotoxicity of
to HeLa cells. Similarly, low salinity could also induce the secretion of the T3SS2 translocon protein VPA1361. These factors may result in the high pathogenicity of
in low-salinity environments. Taken together, these results suggest that low salinity (0.5% NaCl) could affect gene expression to mediate fitness and virulence, which may contribute to the transition of
between aquatic ecosystems and the host.
Vibrio parahaemolyticus is an increasingly important foodborne pathogen that cause acute gastroenteritis in humans. However, the prevalence and transmission of this pathogen in freshwater food ...remains unclear. This study aimed to determine the molecular characteristics and genetic relatedness of V. parahaemolyticus isolates obtained from freshwater food, seafood, environmental, and clinical samples. A total of 138 (46.6%) isolates were detected from 296 food and environmental samples, and 68 clinical isolates from patients. Notably, V. parahaemolyticus was more prevalent in freshwater food (56.7%, 85/150) than in seafood (38.8%, 49/137). Virulence phenotype analyses revealed that the high motility of isolates from freshwater food (40.0%) and clinical isolates (42.0%) was higher than that of isolates from seafood (12.2%), whereas the biofilm-forming capacity of freshwater food isolates (9.4%) was lower than that of seafood (22.4%) and clinical isolates (15.9%). Virulence genes analysis showed that 46.4% of the clinical isolates contained the tdh gene encoding thermostable direct hemolysin (TDH) and only two freshwater food isolates contained the trh gene encoding TDH-related hemolysin (TRH). Multilocus sequence typing (MLST) analysis divided the 206 isolates into 105 sequence types (STs), including 56 (53.3%) novel STs. ST2583, ST469, and ST453 have been isolated from freshwater food and clinical samples. Whole-genome sequence (WGS) analyses revealed that the 206 isolates were divided into five clusters. Cluster II contained isolates from freshwater food and clinical samples, whereas the other clusters contained isolates from seafood, freshwater food, and clinical samples. In addition, we observed that ST2516 had the same virulence pattern, with a close phylogenetic relationship to ST3. The increased prevalence and adaption of V. parahaemolyticus in freshwater food is a potential cause of clinical cases closely related to the consumption of V. parahaemolyticus contaminated freshwater food.
•The high prevalence of Vibrio parahaemolyticus in the freshwater food.•A specific clonal populations of V. parahaemolyticus strains isolated from freshwater food.•The potential transmission of V. parahaemolyticus from freshwater food to humans.
Based on commercially available WC (particle sizes of 0.9 μm and 2.4 μm) and Co, a dual-scale structure WC-6Co cemented carbide was prepared via conventional roller ball milling and following vacuum ...sintering. Effect of ball milling process on the microstructure, densities and mechanical properties were investigated together with the toughening mechanism of the dual-scale structure. The results showed that the dual-scale structured cemented carbide has excellent comprehensive mechanical properties, and its transverse rupture strength, hardness and fracture toughness are 3077 MPa, 1570 HV30 and 10.6 MPa m1/2, respectively. The drilling spherical teeth was prepared by the optimized technology and subjected to impact wear test, the failure mechanism of cemented carbide spherical teeth in drilling operations is studied. The CM-2 spherical teeth had a more reasonable distribution structure of coarse and fine of WC, and exhibited more excellent impact wear resistance in the impact wear test of granite.
•The dual-scale structure WC-6Co cemented carbide was prepared by conventional roller ball milling and vacuum sintering.•The transverse rupture strength, hardness and fracture toughness of the dual-scale structure cemented carbide reached 3077 MPa, 1570 HV30 and 10.6 MPa•m1/2, respectively.•The results show that the CM-2 spherical teeth with higher ball milling intensity exhibits more excellent impact wear resistance in the impact wear test of granite.
Ti6Al4V-x wt%TiB2 titanium matrix composites (x = 0, 0.01, 1, 3.4, 4.8) were prepared by selective laser melting (SLM), followed by solution-aging treatment. The microstructure and mechanical ...properties of the SLM-formed samples and the heat-treated ones were systematically investigated. In the SLM-formed samples, in-situ TiB whiskers clustered around the original position of the TiB2 particles, which reacted with the matrix through dynamic dissolution. The phase transformation of α' → α + β and the coarsening of TiB whiskers occurred during solution-aging treatment. The microhardness of the samples was enhanced with the increase of TiB2, while the heat treatment was harmful to microhardness. The fine grain strengthening and the dispersion strengthening caused by TiB whiskers enhanced the quasi-static compressive strength of the samples. The samples were modified successfully by solution-aging, and the compressive strength and strain of the Ti6Al4V-0.01wt.%TiB2 composite reached 1957 MPa and 23.7% respectively. The samples had higher ductility but lower strength under dynamic load, and the heat treatment had little effect on the dynamic properties.
•The transformation model during SLM was established by studying the microstructure of TMCs with different TiB2 contents.•Solution-aging treatment was used to modify the TMCs, and the microstructure evolution was analyzed.•The comprehensive mechanical properties of the SLM-formed and solution-aging treated TMCs were studies.
Recombinant adenoviruses have broad applications for gene delivery and expression. Furthermore, the adenovirus packaging system facilitates the expression of RNA-guided CRISPR/Cas9 nuclease ...complexes. In this study, we developed a novel system, named AdBlue, for the construction of recombinant adenoviruses using an enzymatic assembly strategy. This system could significantly reduce the time and labor required to generate adenoviral vectors. When applied to CRISPR/Cas9 design, it simplifies the preparation of recombinant adenoviruses carrying nuclease complexes and can induce high levels of site-specific mutagenesis. Our system has outstanding advantages for adenovirus preparation and could be a useful molecular engineering tool for gene delivery and editing.
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