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Abstract The G‐protein‐coupled human cannabinoid receptor 1 (CB1) is a promising therapeutic target for pain management, inflammation, obesity, and substance abuse disorders. The structures of CB1‐Gi complexes in synthetic agonist‐bound forms have been resolved to date. However, the commercial drug recognition and Gq coupling mechanisms of CB1 remain elusive. Herein, the cryo‐electron microscopy (cryo‐EM) structure of CB1‐Gq complex, in fenofibrate‐bound form, at near‐atomic resolution, is reported. The structure elucidates the delicate mechanisms of the precise fenofibrate recognition and Gq protein coupling by CB1 and will facilitate future drug discovery and design.

This paper focuses on the northwest region, which is related to China’s overall ecological security and ethnic stability. This paper selects the neighboring regions of Dingxi City, Gannan Tibetan Autonomous Prefecture and Linxia Hui Autonomous Prefecture as the starting point, deeply and systematically analyzes the impact of different lifestyles on the environment. Using environmental economics, ecological economics, environmental sociology and other related theories, ecological footprint were used to investigate different lifestyles’ impact to environment. Neural network were also used to carry out multi-perspective environmental impact research from the spatial scale and time scale. The research finds that Dingxi, Gannan and Linxia’s different mode of production has led to different lifestyle, and results in different impact on environment. The governments of the three places should take actions to promote ecological civilization and encourage the establishment of an ecologically-friendly and environmentally-friendly way of life so as to reduce the impact on the ecological environment and realize regional sustainable development.

Acute myeloid leukemia (AML) is an aggressive form of hematological neoplasia characterized by failure of myeloid differentiation. AML is a leading cause of death from leukemia. Cytarabine chemotherapy resistance is a major source of refractory/relapsed AML. A major obstacle to the successful treatment of AML results from residual disease maintained by leukemic stem cells (LSCs), which are mostly resistant to conventional chemotherapy. Here, we determined the effect of a natural compound, Jiyuan oridonin A (JOA), on the differentiation blockade in the M2 subtype [particularly t (8;21)] of AML cells, M3 subtype of AML cells (APL cells), and leukemic stem-like cells both in vitro and in vivo. We found that JOA induced cell differentiation and suppressed the colony formation capacity in various AML cell lines (Kasumi-1, KG-1, MUTZ-8, NB4, and HL-60) without eliciting apoptosis. The mechanism of JOA-induced cell differentiation depends on the specificity of cell type. JOA mediated the differentiation of Kasumi-1 cells by activating the hematopoietic cell lineage signaling pathway, while inhibition of c-MYC was involved in the JOA-induced differentiation of NB4 cells. Moreover, JOA was identified to target leukemic stem-like cells by induced cell differentiation in vivo. These findings demonstrated that JOA could inhibit the proliferation of M2 and M3 subtypes of AML cells and leukemic stem-like cells by overcoming the differentiation blockade, which may offer a novel therapeutic strategy for AML to overcome relapse and drug resistance in patients with AML. Our findings highlight the possibility of using compounds like JOA as a promising differentiation-induced agent for the treatment of AML.

The interaction between a GPCR, such as the vasopressin receptor-2 (V2R), and arrestin depends on the receptors’ phosphorylation pattern. Here authors use FRET and NMR to analyze the phosphorylation patterns of the V2R-arrestin complex and show that phospho-interactions are the key determinants of selective arrestin conformational states and correlated functions.

Characterization of dynamic conformational changes in membrane protein complexes by NMR spectroscopy remains challenging. Here authors report the site-specific incorporation of 4-trimethylsilyl phenylalanine (TMSiPhe) into proteins, which enabled the characterization of multiple conformational states of a phospho-β2 adrenergic receptor/β-arrestin-1 complex in response to different receptor ligands.

Acute myelogenous leukemia (AML) is characterized by blockage of cell differentiation leading to the accumulation of immature cells, which is the most prevalent form of acute leukemia in adults. It is well known that all-trans retinoic acid (ATRA) and arsenic trioxide (ATO) are the preferred drugs for acute promyelocytic leukemia (APL). However, they can lead to irreversible resistance which may be responsible for clinical failure after complete remission (CR). Moreover, the differentiation therapy of ATRA-based treatment has not been effective against AML with t(8;21) translocation. Here we aimed to identify the differentiation effect of OGP46 on AML cell lines (HL-60, NB4, and Kasumi-1) and explore its possible mechanisms. We found that OGP46 has significant inhibitory activity against these cells by triggering cell differentiation with cell-cycle exit at G1/G0 and inhibited the colony-formation capacity of the AML cells. It was shown that OGP46 induced the differentiation of NB4 cells via the transcriptional misregulation in cancer signaling pathway by PML-RARα depletion, while it was attributed to the hematopoietic cell lineage and phagosome pathway in Kasumi-1 cells, which are all critical pathways in cell differentiation. These results highlight that OGP46 is an active agent not only in the APL cell line NB4 but also in AML-M2 cell lines, especially Kasumi-1 with t(8;21) translocation. Therefore, OGP46 may be a potential compound for surmounting the differentiation blockage in AML.

Health-related quality of life (HRQoL) is an important aspect in the management of patients with hepatitis B (HB), which remains a serious health problem in China. There have been relatively few HRQoL studies involving Chinese patients with HB. The aim of this study was to analyze HRQoL in patients diagnosed with HB living in Zhejiang Province, China. A cross-sectional sample of 98 patients with chronic HB (CHB), 56 patients with advanced HB that have developed cirrhosis, and 48 healthy controls (HCs), all from Zhejiang Province, was used in this study. HRQoL was assessed using Short-Form 36 (SF-36) version 2, European quality of life questionnaire-5 dimensions (EQ-5D), and chronic liver disease questionnaire (CLDQ). Intergroup score differences were detected with U tests. Factors with a significant effect on HRQoL were identified with Spearman correlational analyses. Patients with HB (both groups) had lower SF-36 scores than HCs (p

Due to the abuse of antibiotics, antibiotic residues can be detected in both natural environment and various industrial products, posing threat to the environment and human health. Here we describe the design and implementation of an engineered Escherichia coli capable of degrading tetracycline (Tc)-one of the commonly used antibiotics once on humans and now on poultry, cattle and fisheries. A Tc-degrading enzyme, TetX, from the obligate anaerobe Bacteroides fragilis was cloned and recombinantly expressed in E. coli and fully characterized, including its Km and kcat value. We quantitatively evaluated its activity both in vitro and in vivo by UV–Vis spectrometer and LC-MS. Moreover, we used a tetracycline inducible amplification circuit including T7 RNA polymerase and its specific promoter PT7 to enhance the expression level of TetX, and studied the dose-response of TetX under different inducer concentrations. Since the deployment of genetically modified organisms (GMOs) outside laboratory brings about safety concerns, it is necessary to explore the possibility of integrating a kill-switch. Toxin-Antitoxin (TA) systems were used to construct a mutually dependent host-plasmid platform and biocontainment systems in various academic and industrious situations. We selected nine TA systems from various bacteria strains and measured the toxicity of toxins (T) and the detoxifying activity of cognate antitoxins (A) to validate their potential to be used to build a kill-switch. These results prove the possibility of using engineered microorganisms to tackle antibiotic residues in environment efficiently and safely. Keywords: TetX, Antibiotic residue, Toxin-antitoxin system, Kill-switch

The effectiveness of the Wolf Pack Algorithm (WPA) in high-dimensional discrete optimization problems has been verified in previous studies; however, it usually takes too long to obtain the best solution. This paper proposes the Multi-Population Parallel Wolf Pack Algorithm (MPPWPA), in which the size of the wolf population is reduced by dividing the population into multiple sub-populations that optimize independently at the same time. Using the approximate average division method, the population is divided into multiple equal mass sub-populations whose better individuals constitute an elite sub-population. Through the elite-mass population distribution, those better individuals are optimized twice by the elite sub-population and mass sub-populations, which can accelerate the convergence. In order to maintain the population diversity, population pretreatment is proposed. The sub-populations migrate according to a constant migration probability and the migration of sub-populations are equivalent to the re-division of the confluent population. Finally, the proposed algorithm is carried out in a synchronous parallel system. Through the simulation experiments on the task assignment of the UAV swarm in three scenarios whose dimensions of solution space are 8, 30 and 150, the MPPWPA is verified as being effective in improving the optimization performance.

To perform air missions with an unmanned aerial vehicle (UAV) swarm is a significant trend in warfare. The task assignment among the UAV swarm is one of the key issues in such missions. This paper proposes PSO-GA-DWPA (discrete wolf pack algorithm with the principles of particle swarm optimization and genetic algorithm) to solve the task assignment of a UAV swarm with fast convergence speed. The PSO-GA-DWPA is confirmed with three different ground-attack scenarios by experiments. The comparative results show that the improved algorithm not only converges faster than the original WPA and PSO, but it also exhibits excellent search quality in high-dimensional space.

Model reference adaptive control (MRAC) schemes are known as an effective method to deal with system uncertainties. High adaptive gains are usually needed in order to achieve fast adaptation. However, this leads to high-frequency oscillation in the control signal and may even make the system unstable. A robust adaptive control architecture was designed in this paper for nonlinear aircraft dynamics facing the challenges of input uncertainty, matched uncertainty, and unmatched uncertainty. By introducing a robust compensator to the MRAC framework, the high-frequency components in the control response were eliminated. The proposed control method was applied to the longitudinal-direction motion control of a nonlinear aircraft system. Flight simulation results demonstrated that the proposed robust adaptive method was able to achieve fast adaptation without high-frequency oscillations, and guaranteed transient performance.

The hydrochloride of the racemic amino acid (2-(7-hydroxycoumarin-4-yl)ethyl)glycine, which can serve as a fluorescent probe in proteins, and two halogen derivatives of it, were synthesized by using a new synthetic protocol in five steps. It is less costly and relatively easy to prepare this kind of fluorescent amino acid with the new synthetic method. Furthermore, it can be applied to synthesize other derivatives of the coumarin amino acid with some specific properties.

The genetic incorporation of a photoreactive unnatural amino acid, p-(2-tetrazole)phenylalanine (p-Tpa), into myoglobin site-specifically in Escherichia coli is described by evolving an orthogonal tTNA/aminoacyl-tRNA synthetase pair and the use of p-Tpa as a bioorthogonal chemical ' handle' for fluorescent labeling of p-Tpa-encoded myoglobin by the photoclick reaction. The structural basis for the biosynthetic incorporation of p-Tpa into proteins is analyzed by solving the X-ray structure of p-Tpa-specific aminoacyl-tRNA synthetase in complex with p-Tpa.

The study explains the NMR structures of various peptide-[Ru.sup.II](porphyrin) complexes. The analysis reveals that heme plays a very important role in the helix orientation of such complexes.

The generation of an orthogonal tRNA/aminoacyl-tRNA synthetase pair that allows the selective introduction of the fluorescent amino acid L-(7-hydroxycoumarin-4-yl) ethylglycine into proteins in Escherichia coli in response to the amber stop codon, TAG is reported. It is observed that in its excited state, 7-hydroxycoumarin is both a strong photoacid, and might facilitate the study of proton-transfer processes in proteins.

The velocity of permanent magnet synchronous motors (PMSMs) must be exactly controlled to promote the development of high-performance drive systems. This work proposed an enhanced adaptive controller to improve the tracking performance of PMSM based on a new higher-order adaptive control mechanism. Firstly, the controller adopted a novel higher-order weighted one-step-ahead criterion function to generate the control law for an equivalent partial form linearization system. This model-free design depended on a pseudo partial derivative (PPD) that was derived online from the input and output (I/O) information of the controlled plant. This approach is especially useful for nonlinear systems with vague dynamics. Secondly, the design guaranteed the stability of the bounded input and output and ensured tracking error monotonic convergence under a restricted set of parameters. Thirdly, the design was simulated and applied on an actual PMSM system to demonstrate the effects of different control parameters. Results show that the approach is fit for motor control and yields satisfactory velocity tracking precision and fault tolerance along with the increased order. Moreover, the approach involves lesser calculation efforts for parameter estimation and simplifies the controller design. This study can meet the demand of velocity tracking and demonstrates the effective applications of this approach for real nonlinear motor systems that are typically difficult to model and control. [ABSTRACT FROM AUTHOR]

The preparation and spectral study of one copper(II) complex with the formula [Cu(bpca)(PHOD)]ClO41 [bpca = bis(2-pyridylcarbonyl)amide anion, PHOD = 1,10-phenathrolinedione] has been reported in this article. The structure of the complex has been determined by single-crystal X-ray diffraction. The crystal analysis shows that the mononuclear copper(II) complex is in five-coordinated surroundings with a tridentate bpca and a bidentate PHOD ligand, and two kinds of C–H…O hydrogen bonds connect the whole structure into three-dimensional network. Meanwhile, the complex has been characterized by infrared (IR), Uv-vis, Electron Paramagnetic Resonance (EPR) spectra, and cyclic voltammetric studies. [ABSTRACT FROM PUBLISHER]

Advances in RNA research and RNA nanotechnology depend on the ability to manipulate and probe RNAwith high precision through chemical approaches, both in vitro and in mammalian cells. However, covalent RNA labeling methods with scope and versatility comparable to those of current protein labeling strategies are underdeveloped. A method is reported for the site- and sequence-specific covalent labeling of RNAs in mammalian cells by using tRNAIle2-agmatidine synthetase (Tias) and click chemistry. The crystal structure of Tias in complex with an azide-bearing agmatine analogue was solved to unravel the structural basis for Tias/substrate recognition. The unique RNA sequence specificity and plastic Tias/substrate recognition enable the site-specific transfer of azide/alkyne groups to an RNA molecule of interest in vitro and in mammalian cells. Subsequent click chemistry reactions facilitate the versatile labeling, functionalization, and visualization of target RNA. [ABSTRACT FROM AUTHOR]

More than one third of all proteins are metalloproteins. They catalyze important reactions such as photosynthesis, nitrogen fixation and CO2 reduction. Metalloproteins such as the olfactory receptors also serve as highly elaborate sensors. Here we review recent developments in functional metalloprotein design using the genetic code expansion approach. We show that, through the site-specific incorporation of metal-chelating unnatural amino acids (UAAs), proton and electron transfer mediators, and UAAs bearing bioorthogonal reaction groups, small soluble proteins can recapitulate and expand the important functions of complex metalloproteins. Further developments along this route may result in cell factories and live-cell sensors with unprecedented efficiency and selectivity. [ABSTRACT FROM AUTHOR]

The article reports on the preparation of a series of helical 15-mer peptides that bind strongly to metalloporphyrins and are determined to be effective in solving their solution nuclear magnetic resonance (NMR) structures. It also highlights the preparation of carbon (C) and oxygen (O) adduct of peptide—Ru(II) porphyrin complexes to deal with the kinetic lability and paramagnetism of metalloporphyrin complexes which proved problematic for iron.

Bacteria adapt to their constantly changing environments largely by transcriptional regulation through the activities of various transcription factors (TFs). However, techniques that monitor TF-promoter interactions in situ in living bacteria are lacking. Herein, we developed a whole-cell TF-promoter binding assay based on the intermolecular FRET between an unnatural amino acid, L-(7-hydroxycoumarin-4-yl) ethylglycine, which labels TFs with bright fluorescence through genetic encoding (donor fluorophore) and the live cell nucleic acid stain SYTO 9 (acceptor fluorophore). We show that this new FRET pair monitors the intricate TF-promoter interactions elicited by various types of signal transduction systems, including one-component (CueR) and two-component systems (BasSR and PhoPQ), in bacteria with high specificity and sensitivity. We demonstrate that robust CouA incorporation and FRET occurrence is achieved in all these regulatory systems based on either the crystal structures of TFs or their simulated structures, if 3D structures of the TFs were unavailable. Furthermore, using CueR and PhoPQ systems as models, we demonstrate that the whole-cell FRET assay is applicable for the identification and validation of complex regulatory circuit and novel modulators of regulatory systems of interest. Finally, we show that the FRET system is applicable for single-cell analysis and monitoring TF activities in Escherichia coli colonizing a Caenorhabditis elegans host. In conclusion, we established a tractable and sensitive TF-promoter binding assay, which not only complements currently available approaches for DNA-protein interactions but also provides novel opportunities for functional annotation of bacterial signal transduction systems and studies of the bacteria-host interface. [ABSTRACT FROM AUTHOR]

Protein-tyrosine phosphatase nonreceptor type 22 (PTPN22) is a lymphoid-specific tyrosine phosphatase (LYP), and mutations in the PTPN22 gene are highly correlated with a spectrum of autoimmune diseases. However, compounds and mechanisms that specifically inhibit LYP enzymes to address therapeutic needs to manage these diseases remain to be discovered. Here, we conducted a similarity search of a commercial database for PTPN22 inhibitors and identified several LYP inhibitor scaffolds, which helped identify one highly active inhibitor, NC1. Using noncompetitive inhibition curve and phosphatase assays, we determined NC1's inhibition mode toward PTPN22 and its selectivity toward a panel of phosphatases. We found thatNC1is a noncompetitive LYP inhibitor and observed that it exhibits selectivity against other protein phosphatases and effectively inhibits LYP activity in lymphoid T cells and modulates T-cell receptor signaling. Results from site-directed mutagenesis, fragment- centric topographic mapping, and molecular dynamics simulation experiments suggested that NC1, unlike other known LYP inhibitors, concurrently binds to a "WPD" pocket and a second pocket surrounded by an LYP-specific insert, which contributes to its selectivity against other phosphatases. Moreover, using a newly developed method to incorporate the unnatural amino acid 2-fluorine-tyrosine and 19F NMR spectroscopy, we provide direct evidence that NC1 allosterically regulates LYP activity by restricting WPD-loop movement. In conclusion, our approach has identified a new allosteric binding site in LYP useful for selective LYP inhibitor development; we propose that the 19F NMR probe developed here may also be useful for characterizing allosteric inhibitors of other tyrosine phosphatases. [ABSTRACT FROM AUTHOR]

Cation-π interactions, where protein aromatic residues supply π systems while a positive-charged portion of phospholipid head groups are the cations, have been suggested as important binding modes for peripheral membrane proteins. However, aromatic amino acids can also insert into membranes and hydrophobically interact with lipid tails. Heretofore there has been no facile way to differentiate these two types of interactions. We show that specific incorporation of fluorinated amino acids into proteins can experimentally distinguish cation-π interactions from membrane insertion of the aromatic side chains. Fluorinated aromatic amino acids destabilize the cation-π interactions by altering electrostatics of the aromatic ring, whereas their increased hydrophobicity enhances membrane insertion. Incorporation of pentafluorophenylalanine or difluorotyrosine into a Staphylococcus aureus phosphatidylinositol-specific phospholipase C variant engineered to contain a specific PC-binding site demonstrates the effectiveness of this methodology. Applying this methodology to the plethora of tyrosine residues in Bacillus thuringiensis phosphatidylinositol- specific phospholipase C definitively identifies those involved in cation-π interactions with phosphatidylcholine. This powerful method can easily be used to determine the roles of aromatic residues in other peripheral membrane proteins and in integral membrane proteins. [ABSTRACT FROM AUTHOR]

Peroxisomes play an essential role in maintaining fatty acid homeostasis. Although mitochondria are also known to participate in the catabolism of fatty acids viaβ-oxidation, differences exist between the peroxisomal and mitochondrial β-oxidation. Only peroxisomes, but not mitochondrion, can shorten very long chain fatty acids. Here, we describe the crystal structure of a ternary complex of peroxisomal 2,4-dienoyl CoA reductases (pDCR) with hexadienoyl CoA and NADP, as a prototype for comparison with the mitochondrial 2,4-dienoyl CoA reductase (mDCR) to shed light on the differences between the enzymes from the two organelles at the molecular level. Unexpectedly, the structure of pDCR refined to 1.84 Å resolution reveals the absence of the tyrosine-serine pair seen in the active site of mDCR, which together with a lysine and an asparagine have been deemed a hallmark of the SDR family of enzymes. Instead, aspartate hydrogen-bonded to the Cα hydroxyl via a water molecule seems to perturb the water molecule for protonation of the substrate. Our studies provide the first structural evidence for participation of water in the DCR-catalyzed reactions. Biochemical studies and structural analysis suggest that pDCRs can catalyze the shortening of six-carbon-long substrates in vitro. However, the Km values ofpDCRfor short chain acyl CoAs are at least 6-fold higher than those for substrates with 10 or more aliphatic carbons. Unlike mDCR, hinge movements permit pDCR to process very long chain polyunsaturated fatty acids. [ABSTRACT FROM AUTHOR]