The structure of the archaeal nuclease RecJ2 implicates its catalytic mechanism and inability to interact with GINS.

Bacterial RecJ exhibits 5'→3' exonuclease activity that is specific to ssDNA; however, archaeal RecJs show 5' or 3' exonuclease activity. The hyperthermophilic archaea Methanocaldococcus jannaschii encodes the 5'-exonuclease MjRecJ1 and the 3'-exonuclease MjRecJ2. In addition to nuclease activity, archaeal RecJ interacts with GINS, a structural subcomplex of the replicative DNA helicase complex. However, MjRecJ1 and MjRecJ2 do not interact with MjGINS. Here, we report the structural basis for the inability of the MjRecJ2 homologous dimer to interact with MjGINS and its efficient 3' hydrolysis polarity for short dinucleotides. Based on the crystal structure of MjRecJ2, we propose that the interaction surface of the MjRecJ2 dimer overlaps the potential interaction surface for MjGINS and blocks the formation of the MjRecJ2-GINS complex. Exposing the interaction surface of the MjRecJ2 dimer restores its interaction with MjGINS. The cocrystal structures of MjRecJ2 with substrate dideoxynucleotides or product dCMP/CMP show that MjRecJ2 has a short substrate binding patch, which is perpendicular to the longer patch of bacterial RecJ. Our results provide new insights into the function and diversification of archaeal RecJ/Cdc45 proteins.

A distributed data processing scheme based on Hadoop for synchrotron radiation experiments.

With the development of synchrotron radiation sources and high-frame-rate detectors, the amount of experimental data collected at synchrotron radiation beamlines has increased exponentially. As a result, data processing for synchrotron radiation experiments has entered the era of big data. It is becoming increasingly important for beamlines to have the capability to process large-scale data in parallel to keep up with the rapid growth of data. Currently, there is no set of data processing solutions based on the big data technology framework for beamlines. Apache Hadoop is a widely used distributed system architecture for solving the problem of massive data storage and computation. This paper presents a set of distributed data processing schemes for beamlines with experimental data using Hadoop. The Hadoop Distributed File System is utilized as the distributed file storage system, and Hadoop YARN serves as the resource scheduler for the distributed computing cluster. A distributed data processing pipeline that can carry out massively parallel computation is designed and developed using Hadoop Spark. The entire data processing platform adopts a distributed microservice architecture, which makes the system easy to expand, reduces module coupling and improves reliability.

Detectability of Single Spinless Stellar-Mass Black Holes through Gravitational Lensing of Gravitational Waves with Advanced LIGO.

We investigate the detectability of gravitational waves that have been lensed by a spinless stellar-mass black hole, with respect to the advanced LIGO. By solving the full relativistic linear wave equations in the spacetime of a Schwarzschild black hole, we find that the strong gravity can create unique signals in the lensed waveform, particularly during the merger and ring-down stages. The differences in terms of fitting factor between the lensed waveform and best-fitted unlensed general relativity template with spin precession and higher-order multipoles are greater than 5% for the lens black hole mass within 70M_{⊙}

Microfluidic rotating-target device capable of three-degrees-of-freedom motion for efficient in situ serial synchrotron crystallography.

There is an increasing demand for simple and efficient sample delivery technology to match the rapid development of serial crystallography and its wide application in analyzing the structural dynamics of biological macromolecules. Here, a microfluidic rotating-target device is presented, capable of three-degrees-of-freedom motion, including two rotational degrees of freedom and one translational degree of freedom, for sample delivery. Lysozyme crystals were used as a test model with this device to collect serial synchrotron crystallography data and the device was found to be convenient and useful. This device enables in situ diffraction from crystals in a microfluidic channel without the need for crystal harvesting. The circular motion ensures that the delivery speed can be adjusted over a wide range, showing its good compatibility with different light sources. Moreover, the three-degrees-of-freedom motion guarantees the full utilization of crystals. Hence, sample consumption is greatly reduced, and only 0.1 mg of protein is consumed in collecting a complete dataset.

Proper motility enhances rumen fermentation and microbial protein synthesis with decreased saturation of dissolved gases in rumen simulation technique.

The physiological function of the reticulorumen plays an essential role in ruminant nutrition, and detailed knowledge of rumen motility can further advance understanding of ruminant nutrition and physiology. Rumen motility was simulated by setting different stirrer rotation speeds in a rumen simulation technique (RUSITEC) system. The aim of this study was to investigate the effects of rotation speeds on rumen fermentation, saturation factor of dissolved gases, hydrogen (H2) and methane (CH4) emissions, microbial protein synthesis, and selected microbial population using RUSITEC. The experiment was performed according to a balanced 3 × 3 Latin square design, and each period included 7 d for adaptation and 3 d for sampling. Three motility treatments included 5, 15, and 25 rpm rotation speeds. Daily total gas and H2 and CH4 emissions had quadratic responses to the increasing rotation speed and were highest at 15 rpm. Quadratic and linear responses (highest at 5 rpm) to increasing rotation speed were observed for saturation factors of H2 and CH4, liquid-dissolved H2 and CH4 concentrations, and headspace concentration of H2 in the gas phase, whereas increasing rotation speed linearly decreased saturation factors of CO2 and liquid-dissolved CO2 concentration. Quadratic and linear responses to increasing rotation speed were observed for molar percentages of acetate, ammonia, and microbial protein concentration, whereas increasing rotation speed quadratically increased pH and decreased total volatile fatty acid concentration and acetate-to-propionate ratio. The 15-rpm rotation speed had the highest values of total volatile fatty acids, acetate molar percentage, and microbial protein concentration. Quadratic and linear responses to increasing rotation speed were observed for copy numbers of solid-associated fungi and fluid-associated bacteria, fungi, and protozoa, while increasing rotation speed linearly increased copy numbers of solid-associated protozoa. Rotation at 15 rpm increased populations of fungi and protozoa in the solid rumen contents and the population of bacteria and fungi in the liquid rumen contents. In summary, this study provides insights on the biofunction of proper rumen motility (i.e., at a rotation speed of 15 rpm), such as improving feed fermentation, increasing gas emissions with decreased dissolved gas concentrations and saturation factors, and promoting microbial colonization and microbial protein synthesis, although further increase in rotation speed (i.e., to 25 rpm) decreases feed fermentation and microbial protein synthesis.

Anti-NSCLC activity in vitro of Hsp90N inhibitor KW-2478 and complex crystal structure determination of Hsp90N-KW-2478.

KW-2478 is a promising anti-cancer lead compound targeting to the molecular chaperone heat shock protein 90 N (Hsp90N). Absence of complex crystal structure of Hsp90N-KW-2478, however, hampered further structure optimization of KW-2478 and understanding on the molecular interaction mechanism. Herein, a high-resolution complex crystal structure of Hsp90N-KW-2478 was determined by X-ray diffraction (XRD, resolution limit: 1.59 Å; PDB ID: 6LT8) and their molecular interaction was analyzed in detail, which suggested that KW-2478 perfectly bound in the N-terminal ATP-binding pocket of Hsp90 to disable its molecular chaperone function, therefore suppressed or killed cancer cells. The results from thermal shift assay (TSA, ΔTm, 18.82 ± 0.51 °C) and isothermal titration calorimetry (ITC, Kd, 7.30 ± 2.20 nM) suggested that there is an intense binding force and favorable thermodynamic changes during the process of KW-2478 binding with Hsp90N. Additionally, KW-2478 exhibited favorable anti-NSCLC activity in vitro, as it inhibited cell proliferation (IC50, 8.16 µM for A549; 14.29 µM for H1975) and migration, induced cell cycle arrest and promoted apoptosis. Thirty-six novel KW-2478 derivatives were designed, based on the complex crystal structure and molecular interaction analysis of Hsp90N-KW-2478 complex. Among them, twenty-two derivatives exhibited increased binding force with Hsp90N evaluated by molecular docking assay. The results would provide new guidance for anti-NSCLC new drug development based on the lead compound KW-2478.

Structural and Functional Insights into PpgL, a Metal-Independent ß-Propeller Gluconolactonase That Contributes to Pseudomonas aeruginosa Virulence.

Biofilm formation is a critical determinant in the pathopoiesis of Pseudomonas aeruginosa It could significantly increase bacterial resistance to drugs and host defense. Thus, inhibition of biofilm matrix production could be regarded as a promising attempt to prevent colonization of P. aeruginosa and the subsequent infection. PpgL, a periplasmic gluconolactonase, has been reported to be involved in P. aeruginosa quorum-sensing (QS) system regulation. However, the detailed function and catalysis mechanism remain elusive. Here, the crystal structure of PpgL is described in the current study, along with biochemical analysis, revealing that PpgL is a typical ß-propeller enzyme with unique metal-independent lactone hydrolysis activity. Consequently, comparative analysis of seven-bladed propeller lactone-catalyzing enzymes and mutagenesis studies identify the critical sites which contribute to the diverse catalytic and substrate recognition functions. In addition, the reduced biofilm formation and attenuated invasion phenotype resulting from deletion of ppgL confirm the importance of PpgL in P. aeruginosa pathogenesis. These results suggest that PpgL is a potential target for developing new agents against the diseases caused by P. aeruginosa.

A 4-gene prognostic signature predicting survival in hepatocellular carcinoma.

OBJECTIVE: To develop an independent prognostic signature for patients with hepatocellular carcinoma (HCC). METHODS: HCC gene expression profile the cancer genome atlas-liver hepatocellular carcinoma and GSE14520 were used as discovery and test set, respectively. Differentially expressed genes (DEGs) were identified between HCC tissues and adjacent normal liver tissues. Univariate Cox proportional hazards regression analysis was performed to identify DEGs correlated with survival of HCC patients. A 4-gene-based signature was constructed based on a least absolute shrinkage and selection operator Cox penalized regression model. The predictive value of the signature was analyzed and validated. RESULTS: Two hundred sixty-three DEGs were identified between HCC and adjacent liver tissues. After univariate survival analysis, 90 DEGs were found to be significantly correlated with the overall survival (OS) of HCC patients, of which 4 genes (KPNA2, CDC20, SPP1, and TOP2A) with non-zero coefficient were used to construct a prognostic signature. The 4-gene signature was significantly associated with the age (P = 0.046), grade ( P = 0.022), and T stage ( P = 0.023) of HCC patients in the discovery set and it also significantly associated with TNM stage ( P = 0.033), and serum alpha-fetoprotein lever ( P = 0.034). Patients in the 4-gene low-risk group were associated with better OS and recurrence-free survival (RFS) than those in the high-risk group in the discovery and test set. Meanwhile, the 4-gene signature is an independent prognostic factor regarding OS and RFS in the discovery and test set. CONCLUSION: We developed a 4-gene-based signature, which could be a candidate prognostic factor for patients with HCC.

The crystal structure of Pyrococcus furiosus RecJ implicates it as an ancestor of eukaryotic Cdc45.

RecJ nucleases specifically degrade single-stranded (ss) DNA in the 5' to 3' direction. Archaeal RecJ is different from bacterial RecJ in sequence, domain organization, and substrate specificity. The RecJ from archaea Pyrococcus furiosus (PfuRecJ) also hydrolyzes RNA strands in the 3' to 5' direction. Like eukaryotic Cdc45 protein, archaeal RecJ forms a complex with MCM helicase and GINS. Here, we report the crystal structures of PfuRecJ and the complex of PfuRecJ and two CMPs. PfuRecJ bind one or two divalent metal ions in its crystal structure. A channel consisting of several positively charged residues is identified in the complex structure, and might be responsible for binding substrate ssDNA and/or releasing single nucleotide products. The deletion of the complex interaction domain (CID) increases the values of kcat/Km of 5' exonuclease activity on ssDNA and 3' exonuclease activity on ssRNA by 5- and 4-fold, respectively, indicating that the CID functions as a regulator of enzymatic activity. The DHH domain of PfuRecJ interacts with the C-terminal beta-sheet domain of the GINS51 subunit in the tetrameric GINS complex. The relationship of archaeal and bacterial RecJs, as well as eukaryotic Cdc45, is discussed based on biochemical and structural results.

Thermococcus Eurythermalis Endonuclease IV Can Cleave Various Apurinic/Apyrimidinic Site Analogues in ssDNA and dsDNA.

Endonuclease IV (EndoIV) is a DNA damage-specific endonuclease that mainly hydrolyzes the phosphodiester bond located at 5' of an apurinic/apyrimidinic (AP) site in DNA. EndoIV also possesses 3'-exonuclease activity for removing 3'-blocking groups and normal nucleotides. Here, we report that Thermococcus eurythermalis EndoIV (TeuendoIV) shows AP endonuclease and 3'-exonuclease activities. The effect of AP site structures, positions and clustered patterns on the activity was characterized. The AP endonuclease activity of TeuendoIV can incise DNA 5' to various AP site analogues, including the alkane chain Spacer and polyethylene glycol Spacer. However, the short Spacer C2 strongly inhibits the AP endonuclease activity. The kinetic parameters also support its preference to various AP site analogues. In addition, the efficient cleavage at AP sites requires ≥2 normal nucleotides existing at the 5'-terminus. The 3'-exonuclease activity of TeuendoIV can remove one or more consecutive AP sites at the 3'-terminus. Mutations on the residues for substrate recognition show that binding AP site-containing or complementary strand plays a key role for the hydrolysis of phosphodiester bonds. Our results provide a comprehensive biochemical characterization of the cleavage/removal of AP site analogues and some insight for repairing AP sites in hyperthermophile cells.

Biofunctional Polymer-Lipid Hybrid High-Density Lipoprotein-Mimicking Nanoparticles Loading Anti-miR155 for Combined Antiatherogenic Effects on Macrophages.

A biofunctional polymer-lipid hybrid high-density lipoprotein-mimicking nanoparticle (HNP) loading anti-miR155 was constructed for combined antiatherogenic effects on macrophages. The HNP consisted of an anti-miR155 core condensed by acid-labile polyethylenimine (acid-labile PEI) polymers and a lipid bilayer coat that was decorated with apolipoprotein A-1, termed acid-labile PEI/HNP. The acid-labile PEI was synthesized with low-molecular-weight PEI and glutaraldehyde to reduce the cytotoxicity and facilitate nucleic acids escaping from acidic endolysosomes. The increased silencing efficiency of acid-labile PEI/HNP was ascribed to the clathrin-mediated endocytosis and successful endolysosomal escape. Decreased intracellular reactive oxygen species production and DiI-oxLDL uptake revealed the antioxidant activities of both anti-miR155 and HNP. Cholesterol efflux assay indicated the potential of HNP in reverse cholesterol transport. Collectively, the acid-labile PEI/HNP not only realized the efficacy of anti-miR155 in macrophages but also exerted the antiatherosclerotic biofunction of HNP.

Subhalo Abundance Matching in f(R) Gravity.

Using the liminality N-body simulations of Shi et al., we present the first predictions for galaxy clustering in f(R) gravity using subhalo abundance matching. We find that, for a given galaxy density, even for an f(R) model with f_{R0}=-10^{-6}, for which the cold dark matter clustering is very similar to the cold dark matter model with a cosmological constant (ΛCDM), the predicted clustering of galaxies in the f(R) model is very different from ΛCDM. The deviation can be as large as 40% for samples with mean densities close to that of L_{*} galaxies. This large deviation is testable given the accuracy that future large-scale galaxy surveys aim to achieve. Our result demonstrates that galaxy surveys can provide a stringent test of general relativity on cosmological scales, which is comparable to the tests from local astrophysical observations.

Effective Dark Matter Halo Catalog in f(R) Gravity.

We introduce the idea of an effective dark matter halo catalog in f(R) gravity, which is built using the effective density field. Using a suite of high resolution N-body simulations, we find that the dynamical properties of halos, such as the distribution of density, velocity dispersion, specific angular momentum and spin, in the effective catalog of f(R) gravity closely mimic those in the cold dark matter model with a cosmological constant (ΛCDM). Thus, when using effective halos, an f(R) model can be viewed as a ΛCDM model. This effective catalog therefore provides a convenient way for studying the baryonic physics, the galaxy halo occupation distribution and even semianalytical galaxy formation in f(R) cosmologies.

[Transferability of Hyperspectral Model for Estimating Soil Organic Matter Concerned with Soil Moisture].

Hyperspectral remote sensing, known as the state-of-the-art technology in the field of remote sensing, can be used to retrieve physical and chemical properties of surface objects based on the interactions between electromagnetic waves and the objects. Soil organic matter (SOM) is one of the most important parameters used in the assessment of soil fertility. Quick estimation of SOM with hyperspectral remote sensing technique can provide essential soil data to support the development of precision agriculture. The presence of external parameters, however, may affect the modeling precision, and further handicap the transfer ability of existing model. With the aim to study the effects of soil moisture on the Vis/NIR estimation of soil organic matter, and the capacity of direct standardization(DS)algorithm in the calibration transfer, 95 soil samples collected in the Jianghan plain were rewetted and air-dried. Reflectance of these samples at 13 moisture levels was measured. Results show that the model calibrated using air-dried samples has the highest prediction accuracy. This model, however, was not suitable for SOM prediction of the rewetted samples. Prediction bias and RPD improved from -8.34-3.32 g x kg(-1) and 0.64-2.04 to 0 and 7.01, when DS algorithm was applied to the spectra of the rewetted samples. DS algorithm has been proven to be effective in removing the effects of soil moisture on the Vis/NIR estimation of SOM, ensuring a transferrable model for SOM prediction with soil samples at different moisture levels.

Structural transformation of the amyloidogenic core region of TDP-43 protein initiates its aggregation and cytoplasmic inclusion.

TDP-43 (TAR DNA-binding protein of 43 kDa) is a major deposited protein in amyotrophic lateral sclerosis and frontotemporal dementia with ubiquitin. A great number of genetic mutations identified in the flexible C-terminal region are associated with disease pathologies. We investigated the molecular determinants of TDP-43 aggregation and its underlying mechanisms. We identified a hydrophobic patch (residues 318-343) as the amyloidogenic core essential for TDP-43 aggregation. Biophysical studies demonstrated that the homologous peptide formed a helix-turn-helix structure in solution, whereas it underwent structural transformation from an α-helix to a ß-sheet during aggregation. Mutation or deletion of this core region significantly reduced the aggregation and cytoplasmic inclusions of full-length TDP-43 (or TDP-35 fragment) in cells. Thus, structural transformation of the amyloidogenic core initiates the aggregation and cytoplasmic inclusion formation of TDP-43. This particular core region provides a potential therapeutic target to design small-molecule compounds for mitigating TDP-43 proteinopathies.

Associations between PTPN2 polymorphisms and susceptibility to ulcerative colitis and Crohn's disease: a meta-analysis.

OBJECTIVE: Ulcerative colitis (UC) and Crohn's disease (CD) result from an interaction between genetic and environmental factors. Though several polymorphisms have been identified in PTPN2, their roles in the incidence of UC and CD are conflicting. This meta-analysis was aimed to clarify the impact of these polymorphisms on UC and CD risk. METHOD: PubMed, EMBASE, Cochrane Library and CBM were searched until 23 July 2013 for eligible studies on three PTPN2 polymorphisms: rs2542151, rs1893217 and rs7234029. Data were extracted, and pooled odd ratios (ORs) as well as 95 % confidence intervals (95 % CIs) were calculated. CONCLUSION: The meta-analysis indicated that rs2542151, rs1893217 and rs1893217 were associated with increased CD risk, while the former was associated with increased UC risk. The differences in age of onset and ethnic groups may influence the associations. Gene-gene and gene-environment interactions should be investigated in the future. RESULTS: Seventeen studies with 18,308 cases and 20,406 controls were included. Significant associations were found between rs2542151 polymorphism and CD susceptibility (OR = 1.22, 95 % CI, 1.15-1.30, I (2) = 32 %), as well as between rs2542151 and UC susceptibility (OR = 1.16, 95 % CI, 1.07-1.25, I (2) = 39 %). A similar result was found in Caucasians, but not in Asians. Moreover, a significant increase in CD risk for all carriers of the minor allele of rs1893217 (OR = 1.45, 95 % CI, 1.23-1.70, I (2) = 0 %) and rs7234029 (OR = 1.36, 95 % CI, 1.16-1.59, I (2) = 0 %) were found. For children, the rs1893217 polymorphism appeared to confer susceptibility to CD (OR = 1.56, 95 % CI, 1.28-1.89, I (2) = 0 %).

Growing a single suspended perfect protein crystal in a fully noncontact manner.

Nucleation is a fundamental process that determines the structure, morphology, and properties of crystalline materials, and is difficult to control because it is unpredictable. Here, we demonstrate a new method to control the protein crystal nucleation using a magnetic force, where we manipulate the movement and coalescence of nucleation precursors by adding paramagnetic salt into the crystallization solution to constrain the number and position of nucleation. We found that protein nucleation could be significantly affected by the magnetic force that the gradient magnetic fields generate. When the magnetization force is sufficiently enough, nucleation can be confined to the crystallization solution with no interface contact; therefore, only one crystal nucleus appears, which results in noncontact suspension growth of a single crystal in the crystallization solution system. Under these situations, the nucleation rate significantly decreases due to the coalescence of the dense liquid phase, and the crystal growth rate also decreases due to the suppression of convection, which increases the crystal quality. Our findings provide a new method for the noncontact control of crystal nucleation and demonstrate that externally applied physical environments can be used to affect the liquid-liquid phase separation process.

A quality comparison of protein crystals grown under containerless conditions generated by diamagnetic levitation, silicone oil and agarose gel.

High-quality crystals are key to obtaining accurate three-dimensional structures of proteins using X-ray diffraction techniques. However, obtaining such protein crystals is often a challenge. Several containerless crystallization techniques have been reported to have the ability to improve crystal quality, but it is unknown which is the most favourable way to grow high-quality protein crystals. In this paper, a quality comparison of protein crystals which were grown under three containerless conditions provided by diamagnetic levitation, silicone oil and agarose gel was conducted. A control experiment on a vessel wall was also simultaneously carried out. Seven different proteins were crystallized under the four conditions, and the crystal quality was assessed in terms of the resolution limit, the mosaicity and the Rmerge. It was found that the crystals grown under the three containerless conditions demonstrated better morphology than those of the control. X-ray diffraction data indicated that the quality of the crystals grown under the three containerless conditions was better than that of the control. Of the three containerless crystallization techniques, the diamagnetic levitation technique exhibited the best performance in enhancing crystal quality. This paper is to our knowledge the first report of improvement of crystal quality using a diamagnetic levitation technique. Crystals obtained from agarose gel demonstrated the second best improvement in crystal quality. The study indicated that the diamagnetic levitation technique is indeed a favourable method for growing high-quality protein crystals, and its utilization is thus potentially useful in practical efforts to obtain well diffracting protein crystals.

[Progress in the fragment-based drug discovery].

As an extension of the structure-based drug discovery, fragment-based drug discovery is matured increasingly, and plays an important role in drug development. Fragments in a small library, with lower molecular mass and high "ligand efficiency", are detected by SPR, MS, NMR, X-ray crystallography technologies and other biophysical methods. Then they are considered as starting points for chemical optimization with the guidance of structural biology methods to get good "drug-like" lead and candidate compounds. In this article, we reviewed the current progress of fragment-based drug discovery and detailed a number of examples to illustrate the novel strategies.