Novel protein ligase based on dual split intein.

Inteins are unique single-turnover enzymes that can excise themselves from the precursor protein without the aid of any external cofactors or energy. In most cases, inteins are covalently linked with the extein sequences and protein splicing happens spontaneously. In this study, a novel protein ligation system was developed based on two atypical split inteins without cross reaction, in which the large segments of one S1 and one S11 split intein fusion protein acted as a protein ligase, the small segments (only several amino acids long) was fused to the N-extein and C-extein, respectively. The splicing activity was demonstrated in E. coli and in vitro with different extein sequences, which showed ∼15% splicing efficiency in vitro. The protein trans-splicing in vitro was further optimized, and possible reaction explanations were explored. As a proof of concept, we expect this approach to expand the scope of trans-splicing-based protein engineering and provide new clues for intein based protein ligase.

Corner-Sharing Tetrahedrally Coordinated W-V Dual Active Sites on Cu2 V2 O7 for Photoelectrochemical Water Oxidation.

The sluggish four-electron oxygen evolving reaction is one of the key limitations of photoelectrochemical water decomposition. Optimizing the binding of active sites to oxygen in water and promoting the conversion of *O to *OOH are the key to enhancing oxygen evolution reaction. In this work, W-doped Cu2 V2 O7 (CVO) constructs corner-sharing tetrahedrally coordinated W-V dual active sites to induce the generation of electron deficiency active centers, promote the adsorption of ─OH, and accelerate the transformation of *O to *OOH for water splitting. The photocurrent obtained by the W-modified CVO photoanode is 0.97 mA cm-2 at 1.23 V versus RHE, which is much superior to that of the reported CVO. Experimental and theoretical results show that the excellent catalytic performance may be attributed to the formation of synergistic dual active sites between W and V atoms, and the introduction of W ions reduces the charge migration distance and prolongs the lifetime of photogenerated carriers. Meanwhile, the electronic structure in the center of the d-band is modulated, which leads to the redistribution of the electron density in CVO and lowers the energy barrier for the conversion of the rate-limiting step *O to *OOH.

Protective effects of berberine against ß-amyloid-induced neurotoxicity in HT22 cells via the Nrf2/HO-1 pathway.

Neuronal apoptosis has been found to have a pivotal role in the course of Alzheimer's disease (AD). Berberine (BBR), a potent antioxidant, occurs in plants such as Berberis, Phellodendron chinense, and Hydrastis canadensis. In this study, a neuronal apoptotic model was established in vitro using HT22 cells induced by Aß25-35 to explore whether BBR contributes to protecting neurons against Aß25-35-induced neurotoxicity, as well as its potential mechanisms. BBR was applied to HT22 cells for 1 h prior to exposing the cells to Aß25-35 for 24 h. A CCK-8 assay was utilized to assess cell viability, and Annexin V - fluorescein isothiocyanate (FITC)/propidium iodide and Hoechst 33342 fluorescence staining were used to measure the rate of cell apoptosis. Existing scientific literature was also reviewed to further determine the effects of BBR on ROS production and mitochondrial function in HT22 cells. Furthermore, the expressions of proteins, including cytochrome C, cleaved caspase-3, p-p65, p65, and Nrf2/HO-1 antioxidant axis were assessed by Western blotting. The data indicated that BBR markedly improved cell viability, inhibited apoptosis and intracellular ROS levels, improved mitochondrial membrane potentials, decreased the rate of p-p65/p65, cytochrome C, and cleaved caspase-3, and intensified the activity of Nrf2/HO-1 antioxidants in HT22 cells. Overall, the findings indicated that BBR provides a certain level of neuroprotectiveness in HT22 cells exposed to Aß25-35 via relieving oxidative stress, as well as by restraining the mitochondrial pathway of cellular apoptosis. In addition, the restraint of NF-κB activity and sensitization of the Nrf2/HO-1 antioxidant axis, which together are intimately involved in the neuroprotection of BBR, may be possible mechanisms accounting for its effectiveness against Aß25-35in vitro.

Efficiency comparison with fovea-sparing internal limiting membrane peeling and complete internal limiting membrane peeling for treating myopic traction maculopathy.

PURPOSE: To explore whether the efficacy of fovea-sparing internal limiting membrane peeling (FS-ILMP) is better than that of complete internal limiting membrane peeling (ILMP). METHODS: This retrospective clinical study included 34 cases (34 eyes) with myopic traction maculopathy collected from June 2017 to February 2019. Twenty-three-gauge (23-G) pars plana vitrectomy (23G PPV) was performed on all patients. In the FS-ILMP group, 18 eyes retained the internal limiting membrane (ILM) of about 1 to 1.5 papillary diameter centered on fovea centralis, while in the standard ILMP group, the ILM was completely removed from 16 eyes. The best corrected visual acuity (BCVA), central foveal thickness (CFT), and other indexes were collected before and 6 months after surgery. RESULTS: There was no significant difference in baseline clinical characteristics between the two groups. CFT and BCVA were significantly improved in both FS-ILMP and standard ILMP group, but the postoperative BCVA of the FS-ILMP group was significantly better than that of the standard ILMP group (P < 0.001). Two cases of subretinal effusion in macula were recorded in the FS-ILMP group, and three eyes in the standard ILMP group developed macular holes after surgery. Although both treatments relieved the mechanical traction of macular fovea, the patients in the FS-ILMP group showed better clinical outcomes in various aspects. CONCLUSION: These results improved our understanding of the clinical application of vitrectomy combined with preservation of ILM upon the fovea centralis, which might lay a foundation for in-depth study on the treatment of myopic traction maculopathy.

Method for determination of optical rotatory dispersion curve by using a polarization axis finder.

We report a new, to the best of our knowledge, measurement method of optical rotatory dispersion (ORD) based on a polarization axis finder (PAF) and digital image processing. The collimated quasi-monochromatic light is output by a monochromator and passes through a polarizer and the chiral samples, and a PAF is employed to modulate the light beam spatially to form an "hourglass" intensity pattern, which is utilized to determine the polarization direction of linearly polarized light. After being recorded by a digital camera, the modulated patterns with and without samples are analyzed by our specially designed digital image processing program to calculate the optical rotation (OR) angle of the chiral substance at a certain wavelength, and the ORD curve can be obtained by fitting a series of OR angles with different wavelengths. The ORD curves of two standard quartz plates with opposite chirality and a glucose solution with different concentrations are measured experimentally, and the fitted coefficients and measurement results agree well with their theoretical values, which indicates that the presented method is an effective and accurate way to determine the ORD of chiral substances conveniently.

Variation in quality of the different prepared formulation granules of rhubarb was evaluated by quantitative analysis of multicomponents with single marker.

Rhei Radix et Rhizoma formula granule (RRFG) , Winy Rhei Radix et Rhizoma formula granule(WRFG) and Rhubarb charcoal formula granules (RCFG) are the three most popular and effective formula granules of rhubarb in China and anthraquinone components are their main active ingredients. In order to discuss the difference in anthraquinone components of these three drugs, a simultaneous quantitative analysis method of multicomponents by single-marker (QAMS) was developed. Emodin was chosed as the internal reference standard, the relative correction factors (RCFs) of aloe-emodin, rhein, chrysophanol and physcion were established and the contents of the four components were calculated based on the RCFs, respectively. Meanwhile, the contents of these components was determined by external standard method (ESM) and compared with QAMS to verify its rationality, feasibility and repeatability. The results showed that there were no significant difference between QAMS and ESM (RSDs≤2.26%). The contents of anthraquinone components showed a wide variation in these three drugs. All of 5 components were higher in RRFG than that in the others and RCFG had the lowest content. This method was successfully applied for the evaluation on difference of these drugs and the wide variation in anthraquinone components indicated there were different pharmacodynamic basis.

Expression regulation of cold-inducible protein RBM3 by FAK/Src signaling for neuroprotection against rotenone under mild hypothermia.

Mild hypothermia is a well-established technique for alleviating neurological injuries in clinical surgery. RNA-binding protein motif 3 (RBM3) has been identified as a crucial factor in mediating hypothermic neuroprotection, providing its induction as a promising strategy for mimicking therapeutic hypothermia. However, little is known about molecular control of RBM3 and signaling pathways affected by hypothermia. In the present study, human SH-SY5Y neuroblastoma cells were used as a neural cell model. Screening of signaling pathways showed that cold exposure led to inactivation of ERK and AMPK pathways, and activation of FAK and PLCγ pathways, with activities of p38, JNK and AKT pathways moderately changed. Next, various small molecule inhibitors specific to these signaling pathways were applied. Interestingly, only FAK-specific inhibitor exhibited a significant inhibitory effect on hypothermia-induced RBM3 gene transcription and protein expression. Likewise, FAK silencing using siRNA technique significantly abrogated the induction of RBM3 by hypothermia. Moreover, FAK inhibition accounted for an inactivation of Src, a known kinase downstream of FAK. Next, either the silencing of Src by siRNA or its inactivation by a chemical inhibitor, strongly blocked the induction of RBM3 by cooling. Notably, in HEK293 and PC12 cells, FAK/Src activation was also shown to be indispensable for hypothermia-stimulated RBM3 expression. Lastly, the CCK8 and Western blot assays showed that both FAK/Src inacitivation and their knockdown substantially abrogate the neuroprotective effects of mild hypothermia against rotenone in SH-SY5Y cells. These data suggest that FAK/Src signaling axis regulates the transcription of Rbm3 gene and mediates neuroprotective effects of mild hypothermia.

Interfacial Linkage and Carbon Encapsulation Enable Full Solution-Printed Perovskite Photovoltaics with Prolonged Lifespan.

Simplified perovskite solar cells (PSCs) were fabricated with the perovskite layer sandwiched and encapsulated between carbon-based electron transport layer (ETL) and counter electrode (CE) by a fully blade-coated process. A self-assembled monolayer of amphiphilic silane (AS) molecules on transparent conducting oxide (TCO) substrate appeals to the fullerene ETL deposition and preserves its integrity against the solvent damage. The AS serves as a "molecular glue" to strengthen the adhesion toughness at the TCO/ETL interface via robust chemical interaction and bonding, facilitating the interfacial charge extraction, increasing PCEs by 77 % and reducing hysteresis. A PCE of 18.64 % was achieved for the fully printed devices, one of the highest reported for carbon-based PSCs. AS-assisted interfacial linkage and carbon-material-assisted self-encapsulation enhance the stability of the PSCs, which did not experience performance degradation when stored at ambient conditions for over 3000 h.

Long noncoding RNA MEG3 silencing protects against hypoxia-induced pheochromocytoma-12 cell injury through inhibition of TIMP2 promoter methylation.

Hypoxia is a common pathological process caused by insufficient oxygen. Long noncoding RNAs (lncRNAs) have been proven to participate in this pathology. Hypoxia is reported to significantly reduce the secretion of tissue inhibitor of metalloproteinase 2 (TIMP2) and TIMP2 induces pheochromocytoma-12 (PC12) cell cycle arrest. Thus, our study aimed to explore the mechanism by which lncRNA maternally expressed gene 3 (MEG3) was implicated in hypoxia-induced PC12 cell injury through TIMP2 promoter methylation. To elucidate the potential biological significance of MEG3 and the regulatory mechanism between MEG3 and TIMP2, a hypoxia-induced PC12 cell injury model was generated. The hypoxia-exposed cells were subjected to a series of overexpression plasmids and short hairpin RNAs, followed by the measurement of levels of MEG3, TIMP2, lactate dehydrogenase (LDH), malondialdehyde (MDA), superoxide dismutase (SOD), reactive oxygen species (ROS), Bcl-2-associated X protein, B-cell lymphoma-2, and caspase-3, as well as the changes in MMP, cell proliferation, apoptosis, and cell cycle progression. On the basis of the findings, MEG3 was upregulated in hypoxia-injured PC12 cells. MEG3 recruited methylation proteins DNMT3a, DNMT3b, and MBD1 and accelerated TIMP2 promoter methylation, which in turn inhibited its expression. Moreover, PC12 cells following MEG3 silencing and TIMP2 overexpression exhibited significantly decreased levels of LDH, MDA, and ROS along with cell apoptosis, yet increased SOD and MMP levels, as well as cell cycle entry to the S phase and cell proliferation. In conclusion, MEG3 silencing suppresses hypoxia-induced PC12 cell injury by inhibiting TIMP2 promoter methylation. This study may provide novel therapeutic targets for hypoxia-induced injury.

Hydrogen Stabilized RhPdH 2D Bimetallene Nanosheets for Efficient Alkaline Hydrogen Evolution.

The design of catalysts with high activity and robust stability for alkaline hydrogen evolution reaction (HER) remains a great challenge. Here, we report an efficient catalyst of two-dimensional bimetallene hydrides, in which H atoms stabilize the rhodium palladium bimetallene. The system exists because of the introduction of H that is in situ chemically released from the formaldehyde solution during the synthesis. This provides a highly stable catalyst based on an unstable combination of metal elements. Density functional theory calculations show the H is confined by electronic interactions and the Miedema rule of reverse stability of the RhPd alloy. The obtained catalyst exhibits outstanding alkaline HER catalytic performance with a low overpotential of 40 mV at 10 mA cm-2 and remarkable stability for over 10 h at 100 mA cm-2. The experimental results show that the confined H improve the activity, while the ultrathin sheet-like morphology yields stability. Our work provides guidance for synthesizing high-activity catalysts by confining heteroatoms into the crystal lattice of bimetallene and also a very novel mechanism for the growth of bimetallene made of highly immiscible components.

Second Harmonic Generation Susceptibilities from Symmetry Adapted Wannier Functions.

Elucidating the orbital level origin of second harmonic generation (SHG) in materials and identifying the local contributions is a long-standing challenge. We report a first principles approach for the SHG where the contributions from individual orbitals or atoms can be evaluated via symmetry adapted Wannier functions without semiempirical parameters. We apply this method to the common SHG materials KBe_{2}BO_{3}F_{2}, KCaCO_{3}F, and ß-BaB_{2}O_{4}, and show that the orbitals on noncentrosymmetric sublattices are responsible for SHG effect and the energies of these orbitals control the magnitude.

Benzoylaconitine Inhibits Production of IL-6 and IL-8 via MAPK, Akt, NF-κB Signaling in IL-1ß-Induced Human Synovial Cells.

Benzoylaconitine (BAC), the main hydrolysate of aconitine, is a lower toxic monoester type alkaloid considered as the pharmacodynamic constituent in Aconitum species. In this study, the effects and mechanisms of BAC on production of inflammatory cytokines interleukin (IL)-6 and IL-8 were investigated in IL-1ß-stimulated human synovial SW982 cells. The SW982 cells were incubated with BAC (0, 5 and 10 µM) before stimulating with IL-1ß (10 ng/mL). The results revealed that BAC suppressed gene and protein expression of IL-6 and IL-8 induced by IL-1ß. BAC decreased activation of mitogen-activated protein kinase (MAPK) and phosphorylation of Akt. BAC also inhibited degradation of inhibitor of kappaB (IκB)-α, phosphorylation and nuclear transposition of p65 protein. The results demonstrate that BAC exerts an anti-inflammatory effect dependent on MAPK, Akt and nuclear factor-κB (NF-κB) pathways in human synovial cells stimulated with IL-1ß, suggesting that BAC may be exploited as a potential therapeutic agent for rheumatoid arthritis (RA) treatment.

Cold-inducible protein RBM3 mediates hypothermic neuroprotection against neurotoxin rotenone via inhibition on MAPK signalling.

Mild hypothermia and its key product, cold-inducible protein RBM3, possess robust neuroprotective effects against various neurotoxins. However, we previously showed that mild hypothermia fails to attenuate the neurotoxicity from MPP+ , one of typical neurotoxins related to the increasing risk of Parkinson disease (PD). To better understand the role of mild hypothermia and RBM3 in PD progression, another known PD-related neurotoxin, rotenone (ROT) was utilized in this study. Using immunoblotting, cell viability assays and TUNEL staining, we revealed that mild hypothermia (32°C) significantly reduced the apoptosis induced by ROT in human neuroblastoma SH-SY5Y cells, when compared to normothermia (37°C). Meanwhile, the overexpression of RBM3 in SH-SY5Y cells mimicked the neuroprotective effects of mild hypothermia on ROT-induced cytotoxicity. Upon ROT stimulation, MAPK signalling like p38, JNK and ERK, and AMPK and GSK-3ß signalling were activated. When RBM3 was overexpressed, only the activation of p38, JNK and ERK signalling was inhibited, leaving AMPK and GSK-3ß signalling unaffected. Similarly, mild hypothermia also inhibited the activation of MAPKs induced by ROT. Lastly, it was demonstrated that the MAPK (especially p38 and ERK) inhibition by their individual inhibitors significantly decreased the neurotoxicity of ROT in SH-SY5Y cells. In conclusion, these data demonstrate that RBM3 mediates mild hypothermia-related neuroprotection against ROT by inhibiting the MAPK signalling of p38, JNK and ERK.

Lone pair effects on ternary infrared nonlinear optical materials.

First-principles studies of the crystal structures, electronic structures and optical properties of noncentrosymmetrical (NCS) K3AsS4, Li3AsS3, Pb9As4S15 and Ag3AsS3 have been performed by means of density functional theory. Via a theoretical method to compute the intensity of the lone pair stereochemical activity of an As-S group, the correlated mechanism among the crystal structures, the stereo-chemical activity of lone pairs on As and the second harmonic generation (SHG) response has been clarified. The results prove that the SHG response is not only attributed to the lone pair stereochemical activity of the As-S group but also related to the direction of the forming layers in the crystal structure arrangement. Besides, the quantitative method for the stereo-chemical activity of lone pairs is universal, which is valid for other lone pair systems like those containing Pb2+, Bi3+, Sn2+, etc. The findings facilitate the exploration of materials that may exhibit a relatively large second order NLO reaction and can be used in infrared applications.

Module-Guided Design Scheme for Deep-Ultraviolet Nonlinear Optical Materials.

Design of functional materials with targeted properties is a challenge because of the diversity of their potential structures. The functional performances of materials are mainly determined by the chemistry and electronic structure of modules consisting of local atomic groups with special arrangements. Tetrahedral modules are excellent modules for designing deep-ultraviolet/ultraviolet (UV) nonlinear optical (NLO) materials, but they are rarely favored due to their unpredictable optical anisotropy and second harmonic generation (SHG) response. In this work, we have developed a module-guided ab initio approach for evaluating the optical anisotropy of tetrahedral modules. The application of this method indicates that the tetrahedral modules with a specific arrangement will enhance the optical anisotropy of materials. With the functional modules consisting of tetrahedral modules and rare-earth cations, new high-performance rare-earth phosphates were assembled. These materials are promising deep-UV NLO materials because of their appropriate birefringences, large band gaps, moderate SHG responses, and easy to obtain large size crystals.

Efficient polarization direction measurement by utilizing the polarization axis finder and digital image processing.

We report a new quantitative measurement method of polarization direction based on the polarization axis finder (PAF) and digital image processing. The PAF acts as an azimuthal analyzer to determine the polarization direction of linearly polarized light and form an "hourglass" intensity pattern, and a digital camera is employed to record the pattern, and then the lightwave's polarization direction is obtained accurately by analyzing the pattern with a specially designed digital image processing algorithm. The quantitative measuring experiments have been carried out by validating the standard quartz plates with known rotation angles, and the measurement accuracy has reached 0.01°.

A Fluorooxosilicophosphate with an Unprecedented SiO2 F4 Species.

The large number and structural beauty of silicates are largely due to the variety of connection mode of SiO4 tetrahedra. SiO6 and SiF6 octahedra are also known and give rise for structural versatility of inorganic silicates. However, to date, the crystal structure of inorganic fluorooxosilicates with oxofluoride SiOx F4-x or SiOx F6-x species are unknown. Now, fluorine was successfully introduced into the silicophosphates, and the first fluorooxosilicophosphate K4 Si3 P2 O7 F12 with an unprecedented SiO2 F4 species was synthesized. The existence of Si-F bonds was verified by comprehensively experimental and theoretical work. Using ab initio and bond valence calculations, the oxofluoride SiOx F6-x species is shown to be stable when oxygen atoms connect to other atoms with strong covalent interactions. This work will contribute to the structural diversity of silicate chemistry by the exploration of the new fluorooxosilicates.

A3Sr2P7O21 (A = Rb, Cs): Two Polyphosphates Based on Different Types of P-O Chains and Ring Structures.

Two polyphosphates containing two types of polymerization of the [PO4] groups, Rb3Sr2P7O21 and Cs3Sr2P7O21, were grown through a spontaneous nucleation method. Single-crystal X-ray diffraction data were collected in order to determine their structures. Interestingly, Rb3Sr2P7O21 is the first example of two kinds of [PO3]∞ linear chains coexisting in one phosphate structure. However, in the structure of Cs3Sr2P7O21, the isolated [P4O12] ring and the 1D [PO3]∞ chain can be observed, which is also rare in phosphates. After careful structural analysis, the alkali-metal cations have an effect on the polymerization of the [PO4] groups and make Rb3Sr2P7O21 and Cs3Sr2P7O21 crystallize in different space groups. What is more, IR spectra, UV-vis-NIR diffuse reflectance spectroscopy data, and first-principles theoretical calculations were adopted to determine the optical properties and the structure-properties relationship of the compounds.

Linear-to-λ-Shape P-O-P Bond Transmutation in Polyphosphates with Infinite (PO3)∞ Chain.

A new metal polyphosphate, α-CsBa2(PO3)5, exhibiting the first example of a linear P-O-P bond angle in a one-dimensional (PO3)∞ chain has been reported. Interestingly, α → ß phase transition occurs in CsBa2(PO3)5 along with the P-O-P bonds varying from linear to λ-shape, suggesting that α-CsBa2(PO3)5 with unfavorable linear P-O-P bonds is more stable at ambient temperature.

A New Deep-Ultraviolet Transparent Orthophosphate LiCs2PO4 with Large Second Harmonic Generation Response.

LiCs2PO4, a new deep-ultraviolet (UV) transparent material, was synthesized by the flux method. The material contains unusual edge-sharing LiO4-PO4 tetrahedra. It exhibits a very short absorption edge of λ = 174 nm and generates the largest powder second harmonic generation (SHG) response for deep-UV phosphates that do not contain additional anionic groups, i.e., 2.6 times that of KH2PO4 (KDP). First-principles electronic structure analyses confirm the experimental results and suggest that the strong SHG response may originate from the aligned nonbonding O-2p orbitals. The discovery and characterization of LiCs2PO4 provide a new insight into the structure-property relationships of phosphate-based nonlinear optical materials with large SHG responses and short absorption edges.