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GPCR oligomerization plays a critical role in cellular signaling, yet the stoichiometry of the interactions between oligomers and binding ligands in living cells remains a longstanding challenge. Here, by developing a dual-color simultaneous tracking system based on a total internal reflection fluorescence microscope (TIRFM), the CCR5-CCL5 interactions are visualized and quantitatively assessed in real time. Results show that each oligomeric state of CCR5 could bind with CCL5 but with different binding affinities; CCR5 dimers have a 3.5-fold higher binding affinity than the monomers. The dimerization may cause an asymmetric conformational change which makes the first binding pocket have a 3.5-fold higher binding affinity and the second have only a half compared with the monomeric CCR5. This study is the first example to directly scrutinize the CCR5-CCL5 interactions at the single-molecule level on living cell membranes and will offer great potential for the interaction stoichiometry study of diverse surface proteins.

Single-molecule fluorescence imaging is a powerful tool to study protein function by tracking molecular position and distribution, but the precise and rapid identification of dynamic molecules remains challenging due to the heterogeneous distribution and interaction of proteins on the live cell membrane. We now develop a deep-learning (DL)-assisted single-molecule imaging method that can precisely distinguish the monomer and complex for rapid and real-time tracking of protein interaction. This DL-based model, which comprises convolutional layers, max pooling layers, and fully connected layers, is trained to reach an accuracy of >98% for identifying monomer and complex. We use this method to investigate the dynamic process of chemokine receptor CXCR4 on the live cell membrane during the early signaling stage. The results show that, upon ligand activation, the CXCR4 undergoes a dynamic process of forming a receptor complex. We further demonstrate that the CXCR4 complex tends to be internalized at 2.5-fold higher rate into the cell interior than the monomer via the clathrin-dependent pathway. This study is the first example to scrutinize the early signaling process of CXCR4 at the single-molecule level on the live cell membrane. We envision that this DL-assisted imaging method would be a broadly useful technique to study more protein families for elucidating their physiological and pathological functions.

Due to its biochemical and spectroscopic properties, phycocyanobilin (PCB) is of special interest for a variety of applications, but the production of PCB from Spirulina is complicated and costly. Here we report the assembly and regulation of the entire heme biosynthetic pathway for combinatorial biosynthesis of PCB in Escherichia coli. Our results shows that a plasmid containing bicistronic expression of hox1 and pcyA yields 6.64 mg/L PCB, which is higher than that of plasmid with one promoter for both genes (p

The uniformly flower-like WO3/CoWO4/Co nanostructures were synthesized via a facile one-step hydrothermal reaction with solid heteropolyacid of W and Co as precursors and followed by heating process and calcination under the air condition. The as-prepared WO3/CoWO4/Co composite showed a high discharge capacity of 1104 mAhg−1 after 40 cycles at 100 mAg−1, and delivered excellent rate performance of 1074, 685, 509, 335 mAhg−1 at 200,400,800,1600 mAg−1, respectively. The excellent electrochemical performance of WO3/CoWO4/Co nanostructures is attributed to the uniform dispersion of discrete nanoparticles, and it also can be highly optimized by the freestanding combination of Co-doped tungsten trioxide and cobalt tungstate, which helps to provide high specific capacity, high rate cycling and stable life performance.

The endoplasmic reticulum–associated degradation (ERAD) pathway mediates the endoplasmic reticulum–to–cytosol retrotranslocation of defective proteins through protein complexes called retrotranslocons. Defective proteins usually have complex conformations and topologies, and it is unclear how ERAD can thread these conformationally diverse protein substrates through the retrotranslocons. Here, we investigated the substrate conformation flexibility necessary for transport via retrotranslocons on the ERAD-L, ERAD-M, and HIV-encoded protein Vpu-hijacked ERAD branches. To this end, we appended various ERAD substrates with specific domains whose conformations were tunable in flexibility or tightness by binding to appropriate ligands. With this technique, we could define the capacity of specific retrotranslocons in disentangling very tight, less tight but well-folded, and unstructured conformations. The Hrd1 complex, the retrotranslocon on the ERAD-L branch, permitted the passage of substrates with a proteinase K–resistant tight conformation, whereas the E3 ligase gp78-mediated ERAD-M allowed passage only of nearly completely disordered but not well-folded substrates and thus may have the least unfoldase activity. Vpu-mediated ERAD, containing a potential retrotranslocon, could unfold well-folded substrates for successful retrotranslocation. However, substrate retrotranslocation in Vpu-mediated ERAD was blocked by enhanced conformational tightness of the substrate. On the basis of these findings, we propose a mechanism underlying polypeptide movement through the endoplasmic reticulum membrane. We anticipate that our biochemical system paves the way for identifying the factors necessary for the retrotranslocation of membrane proteins.

A new small-molecule donor with a dithieno[2,3-d:2′,3′-d′]-benzo[1,2-b:4,5-b′]-dithiophene (DTBDT) core and both alkyl and alkylthio substituents is designed and synthesized to improve the miscibility between DTBDT-based small molecules and [6,6]-phenyl-C71-butyric acid methyl ester (PC71BM). The alkyl substituent on the 4-position and the alkylthio substituent on the 5-position of the substituted thiophene are expected to improve intermolecular interactions and prevent severe aggregation of the small molecules. The new small molecule, DTBDT-S-C8-TTR, exhibits a homogenous blend morphology with small domains and edge-on-oriented crystalline structures in blends with PC71BM, and give a maximum power conversion efficiency (PCE) of 8.43%. To recover the crystallinity of the DTBDT-S-C8-TTR small molecules weakened after being blended with PC71BM, a solvent vapor annealing (SVA) treatment is performed. The SVA-treated blend films reveal well-developed crystalline domains with interconnected fibrillar structures. This blend morphology allows efficient charge carrier transport in blends and leads to increased PCEs. The maximum PCE of 9.18% achieved using DTBDT-S-C8-TTR suggests that substituting both alkylthio and alkyl groups into DTBDT can yield small-molecule-based organic photovoltaics (OPVs) displaying improved photovoltaic performances.

An ultra-high fluorescence enhancement for two dyes on photonic crystal films was achieved to construct a two-color immuno-dot blot assay. This assay was demonstrated to simultaneously detect chemokine receptor co-expressed in cancer cells and reveal their co-operative and subtle changes after binding with respective ligands and drugs.

Dimerization and oligomerization of G-protein coupled receptors (GPCRs) have emerged as important characters during their trans-membrane signal transduction. However, until now the relationship between GPCR dimerization and their trans-membrane signal transduction function is still uncovered. Here, using pertussis toxin (PTX) to decouple the receptor from G protein complex and with single-molecule imaging, we show that in the presence of agonist, cells treated with PTX showed a decrease in the number of dimers and oligomers on the cell surface compared with untreated ones, which suggests that oligomeric status of CXCR4 could be significantly influenced by the decoupling of G protein complex during its signal transduction process. Moreover, with chlorpromazine (CPZ) to inhibit internalization of CXCR4, it was found that after SDF-1α stimulation, cells treated with CPZ showed more dimers and oligomers on the cell surface than untreated ones, which suggest that dimers and oligomers of CXCR4 tend to internalize more easily than monomers. Taken together, our results demonstrate that dimerization and oligomerization of CXCR4 is closely related with its G protein mediated pathway and β-arrestin mediated internalization process, and would play an important role in regulating its signal transduction functions.

Polymer thermal degradation in high-temperature reservoirs is a critical factor to concern during polymer flooding numerical simulation. Based on the continuum assumption, partial differential equations were established to describe polymer viscosity changes. The existing viscosity model was modified to develop the numerical simulation model for polymer thermal degradation in high-temperature oil reservoirs. Results show the modified viscosity model describes the actual physiochemical phenomenon that mass concentration remains constant while viscosity declines during thermal degradation. The modified viscosity model improves the efficiency for polymer thermal degradation simulation and provides theoretical basis for parameter design and risk assessment of polymer flooding projects.

Since all of the organophosphorus pesticides (OPP) inhibit the cholinesterases with a common mechanism, it is still challenging to detect OPP selectively with inhibition-based biosensors. This study focuses on the conversion of a typical inhibition biosensing to a selective substrate-like biosensing. The interaction of quinalphos with plant-esterase involves not only a decrease in enzyme activity but also a heterolytic bond cleavage of quinalphos. The leaving group eliminated from quinalphos is an ideal biomarker due to its specificity in most OPP. Thus, using 2-hydroxyquinoxaline (HQO), the leaving group of quinalphos, as the biomarker and meso-tetra (4-sulfonatophenyl) porphine (TPPS4) as an optical probe, quinalphos can be selectively detected. The molecular recognition between TPPS4 and HQO leads to a considerable sensitivity of the detection. The spectral responses of TPPS4 show a linear dependence on quinalphos concentration in the presence of plant-esterase within the 0.01-1 mg kg(-1) range. The detection limit is 0.01 mg kg(-1), well below the maximum residue limits (MRLs) defined by European Union (0.05 mg kg(-1)) and China (0.2 mg kg(-1)).

Molecular fluorescence blinking provides a simple and attractive way to achieve super-resolution localization via conventional fluorescence microscopy. However, success in super-resolution imaging relies heavily on their blinking characteristics. We here report easily prepared and photostable nanoparticles, carbon dots (CDs), with desirable fluorescence blinking for high-density super-resolution imaging. The CDs exhibit a low duty cycle (∼0.003) and high photon output (∼8000) per switching event, as well as show much higher resistance to photobleaching than Alexa 647 or Cy5 typically used in single molecule localization microscopy. The stable blinking of CDs allows to perform high-density localization imaging at a resolution of 25 nm by sequentially recording the particle positions. The CD-based super-resolution imaging is further demonstrated by rendering CD-stained tubular peptide self-assemblies, CD-packed clusters with well-defined patterns, and CD-stained microtubules in a cell. Furthermore, this method has been validated as a valuable tool to detect the clustering and distribution of protein receptors in the plasma membrane that are not discerned with normal fluorescence imaging.

Background Chemokines and their cognate receptors play important role in the control of leukocyte chemotaxis, HIV entry and other inflammatory diseases. Developing an effcient method to investigate the functional expression of chemokines and its interactions with specific receptors will be helpful to asses the structural and functional characteristics as well as the design of new approach to therapeutic intervention. Results By making systematic optimization study of expression conditions, soluble and functional production of chemokine C-C motif ligand 8 (CCL8) in Escherichia coli (E. coli) has been achieved with approx. 1.5 mg protein/l culture. Quartz crystal microbalance (QCM) analysis exhibited that the purified CCL8 could bind with C-C chemokine receptor type 3 (CCR3) with dissociation equilibrium constant (K D) as 1.2 × 10−7 M in vitro. Obvious internalization of CCR3 in vivo could be detected in 1 h when exposed to 100 nM of CCL8. Compared with chemokine C-C motif ligand 11 (CCL11) and chemokine C-C motif ligand 24 (CCL24), a weaker chemotactic effect of CCR3 expressing cells was observed when induced by CCL8 with same concentration. Conclusion This study delivers a simple and applicable way to produce functional chemokines in E. coli. The results clearly confirms that CCL8 can interact with chemokine receptor CCR3, therefore, it is promising area to develop drugs for the treatment of related diseases. Electronic supplementary material The online version of this article (10.1186/s12865-017-0237-5) contains supplementary material, which is available to authorized users.

Produced water from polymer flooding was much more difficult to be treated than that from water flooding. The mechanisms of polymer stabilizing the oil-in-water emulsion were the decreases of interfacial tension and electric conductivity, and the increases of zeta potential and bulk phase viscosity. Inorganic flocculant, the combination of inorganic flocculant and polymer degradation agent, and the combination organic flocculant and coagulant were used to break the oil-in-water emulsion stabilized with polymer. It was the most effective way to use the combination of organic flocculant and coagulant for treating polymer stabilizing oil-in-water emulsion.

Cytocompatibility of high nitrogen nickel-free stainless steel (HNS) with different nitrogen content was evaluated and compared with a conventional austenitic stainless steel 317L. The MTT assay (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) was performed on MG63 osteoblasts to assess the cytotoxicity. The expression of selected marker typical of differentiated osteoblasts, such as alkaline phosphatase activity (AKP), was also monitored in MG63 cells cultured on the tested materials. As a result, HNS had higher cell growth than 317L; meanwhile the cytocompatibility was increased with increasing nitrogen content. Furthermore, HNS enhanced osteoblasts differentiation, as confirmed by AKP activity. Overall these facts indicated that HNS had higher cytocompatibility than 317L and the nitrogen content contributed to the higher cytocompatibility of HNS. The influence of nitrogen on surface energy further explained the cytocompatibility of HNS.

Polyurea microcapsules containing 33, 50 and 67 wt-% sulphur were prepared by interfacial polymerisation. The characteristics of the fabricated microcapsules were investigated for morphology, thermal property and release property in vulcanising rubber. The morphology showed that microcapsules loading 33 wt-% sulphur were spherical and smooth. The microcapsule sizes decreased going from 33 to 50 to 67 wt-% sulphur. Thermal analysis demonstrated that the microcapsules were thermally stable below 250°C. The process of sulphur released from polyurea microcapsules was put forward in vulcanising nitrile butadiene rubber (NBR) and verified by vulcanising property and tensile strength and elongation of the vulcanised NBR.

The effects of small amounts of cerium and antimony additions on the microstructure and the mechanical properties of AZ91D (Mg-9A1-Zn) based alloy were researched via the expendable pattern casting (EPC) process. The results show that the microstructure is obviously refined and the tensile strength of the AZ91D based alloy at ambient temperature is significantly improved. When compared to AZ91D, the AZ91D-1.0%Ce-0.4%Sb alloy has higher ultimate tensile strength and elongation. Its ultimate tensile strength and elongation are enhanced by 39% and 47%, respectively. The morphology of the tensile fracture of the AZ91D-1.0%Ce-0.4%Sb alloy has more characteristics of quasi-cleavage. This indicates that it has had a larger plastic deformation before failure. The tensile strength and elongation decrease with the increase of Ce and Sb contents because of the coarsening and volume Increase of CeSb and Al 11 Ce 3 phases.

To detect the content of 12 heavy metals in blood and hair sample from a general population of Pearl River Delta area, and to analyze the influence of duration of residence, gender, age, smoking and drinking on the heavy metal content. Use inductively coupled plasma mass spectrometry to detect the content of 12 heavy metals lead (Pb), mercury (Hg), cadmium (Cd), aluminum (Al), arsenic (As), copper (Cu), chrome (Cr), manganese (Mn), nickel (Ni), zinc (Zn), tin (Sn) and antimony (Sb) in blood and hair samples of a total of 50 subjects from a general population, collected by multistage stratified cluster random sampling method. The geometric mean of heavy metal content in blood samples of general population (μg/L): blood aluminum 214.00; blood chrome 92.82; blood manganese 21.43; blood nickel 20.59; blood copper 0.67; blood zinc 11.50; blood arsenic 0.55; blood cadmium 2.45; blood tin 0.00; blood antimony 1.92; blood lead 158.84; and blood mercury 1.19. The geometric mean of heavy metal content in hair samples of general population (μg/g): hair aluminum is 84.65; hair chrome 0.00; hair manganese 2.44; hair nickel 0.61; hair copper 28.49; hair zinc 136.65; hair arsenic 0.75; hair cadmium 0.46; hair tin 1.04; hair antimony 0.05; hair lead 8.97; and hair mercury 0.69. Some heavy metals were correlated with duration of residence, gender, age, smoking and drinking. This was the first time that simultaneously detecting heavy metal content in blood and hair was used to analyze the internal heavy metal burden in resident population of Pearl River Delta area. These data can serve as reference for further research.

OBJECTIVE To evaluate the efficacy of oxymatrine in treating chronic hepatitis C and its mechanism. METHODS Forty-three patient were divided randomly into the treated group (20 cases) and the control group (23 cases). The treated group was given oxymatrine 600 mg per day intramuscularly, and the control group was given the general liver protective agents such as vitamins. The therapeutic course of both groups was 3 months. RESULTS HCVRNA of 8 in 17 cases (47.1%) of the treated group converted to negative, while in 18 cases of the control group, the negative conversion only took place in 1 patient (5.6%), the negative conversion rate was significantly higher in the treated group than that in the control group (P < 0.05). The normalization rates of serum alanine transaminase (ALT) of the treated group after 1 month and 2 months treatment was higher than that of the control group, but after 3 months treatment, the normalization rates of the two groups were not different significantly. Plasma level of soluble interleukin-2 receptor and serum level of collagen type IV in the treated group were lowered significantly after treatment, but in the control group, there were no significant change, the difference between the two groups was significant (P < 0.01, P < 0.05). CONCLUSION Oxymatrine is effective in inhibiting proliferation of HCV, antagonisting liver fibrosis and regulating immune reaction of the host, so it could be a safe, effective drug in treating chronic hepatitis C.