Triple Domain Research Articles

Comparative genomic and methylome analysis of non-virulent D74 and virulent Nagasaki Haemophilus parasuis isolates.

Haemophilus parasuis is a respiratory pathogen of swine and the etiological agent of Glässer's disease. H. parasuis isolates can exhibit different virulence capabilities ranging from lethal systemic disease to subclinical carriage. To identify genomic differences between phenotypically distinct strains, we obtained the closed whole-genome sequence annotation and genome-wide methylation patterns for the highly virulent Nagasaki strain and for the non-virulent D74 strain. Evaluation of the virulence-associated genes contained within the genomes of D74 and Nagasaki led to the discovery of a large number of toxin-antitoxin (TA) systems within both genomes. Five predicted hemolysins were identified as unique to Nagasaki and seven putative contact-dependent growth inhibition toxin proteins were identified only in strain D74. Assessment of all potential vtaA genes revealed thirteen present in the Nagasaki genome and three in the D74 genome. Subsequent evaluation of the predicted protein structure revealed that none of the D74 VtaA proteins contain a collagen triple helix repeat domain. Additionally, the predicted protein sequence for two D74 VtaA proteins is substantially longer than any predicted Nagasaki VtaA proteins. Fifteen methylation sequence motifs were identified in D74 and fourteen methylation sequence motifs were identified in Nagasaki using SMRT sequencing analysis. Only one of the methylation sequence motifs was observed in both strains indicative of the diversity between D74 and Nagasaki. Subsequent analysis also revealed diversity in the restriction-modification systems harbored by D74 and Nagasaki. The collective information reported in this study will aid in the development of vaccines and intervention strategies to decrease the prevalence and disease burden caused by H. parasuis.

Condensates in rotating turbulent flows

Using a large number of numerical simulations we examine the steady state of rotating turbulent flows in triple periodic domains, varying the Rossby number $Ro$ (that measures the inverse rotation rate) and the Reynolds number $Re$ (that measures the strength of turbulence). The examined flows are sustained by either a helical or a non-helical Roberts force, that is invariant along the axis of rotation. The forcing acts at a wavenumber $k_{f}$ such that $k_{f}L=4$, where $2\unicode[STIX]{x03C0}L$ is the size of the domain. Different flow behaviours were obtained as the parameters are varied. Above a critical rotation rate the flow becomes quasi-two-dimensional and transfers energy to the largest scales of the system, forming large coherent structures known as condensates. We examine the behaviour of these condensates and their scaling properties close to and away from this critical rotation rate. Close to the critical rotation rate the system transitions supercritically to the condensate state, displaying a bimodal behaviour oscillating randomly between an incoherent-turbulent state and a condensate state. Away from the critical rotation rate, it is shown that two distinct mechanisms can saturate the growth of the large-scale energy. The first mechanism is due to viscous forces and is similar to the saturation mechanism observed for the inverse cascade in two-dimensional flows. The second mechanism is independent of viscosity and relies on the breaking of the two-dimensionalization condition of the rotating flow. The two mechanisms predict different scaling with respect to the control parameters of the system (Rossby and Reynolds), which are tested with the present results of the numerical simulations. A phase space diagram in the $Re,Ro$ parameter plane is sketched.

Numerical simulation of mesoscale surface pressure features with trailing stratiform squall lines using WRF -ARW model over Gangetic West Bengal region

In the present study, an attempt has been made to investigate the simulation of mesoscale surface pressure patterns like pre-squall mesolow, mesohigh and wake low associated with leading convective line-trailing stratiform (TS) squall lines over Gangetic West Bengal (GWB). For this purpose, a two way interactive triple nested domain with high resolution WRF model having2 km grid length in the innermost domain is used. The model simulated results are compared with the available in-situ observations obtained as a part of Severe Thunderstorm: Observations and Regional Modeling (STORM) programme, reflectivity products of Doppler Weather Radar (DWR) Kolkata and TRMM rainfall. Three TS squall lines (15 May 2009, 5 May 2010 and 7 May 2010) are chosen during pre-monsoon thunderstorm season for this study. The model simulated results of diurnal variation of temperature, relative humidity, wind speed and direction at the station Kharagpur in GWB region reveal a sudden fall in temperature, increase in the amount of relative humidity and sudden rise in wind speed during the arrival of the storms. Such results are well comparable with the observations though there are some leading or lagging of time in respect of actual occurrences of such events. The study indicates that the model is able to predict the occurrences of three typical surface pressure features namely: pre-squall mesolow, meso high and wake low. The predicted surface parameters like accumulated rainfall, maximum reflectivity and vertical profiles (temperature, relative humidity and winds) are well accorded with the observations. The convective and stratiform precipitation region of the TS squall lines are well represented by the model. A strong downdraft is observed to be a contributory factor for formation of mesohigh in the convective region of the squall line. Wake low is observed to reside in the stratiform rain region and the descending dry air at this place has triggered the wake low through adiabatic warming. This study has established the usefulness of the high resolution model in predicting trailing stratiform squall lines and its associated features over the study region.

H3K14ac is linked to methylation of H3K9 by the triple Tudor domain of SETDB1

SETDB1 is an essential H3K9 methyltransferase involved in silencing of retroviruses and gene regulation. We show here that its triple Tudor domain (3TD) specifically binds to doubly modified histone H3 containing K14 acetylation and K9 methylation. Crystal structures of 3TD in complex with H3K14ac/K9me peptides reveal that peptide binding and K14ac recognition occurs at the interface between Tudor domains (TD) TD2 and TD3. Structural and biochemical data demonstrate a pocket switch mechanism in histone code reading, because K9me1 or K9me2 is preferentially recognized by the aromatic cage of TD3, while K9me3 selectively binds to TD2. Mutations in the K14ac/K9me binding sites change the sub-nuclear localization of 3TD. ChIP-seq analyses show that SETDB1 is enriched at H3K9me3 regions and K9me3/K14ac is enriched at SETDB1 binding sites overlapping with LINE elements, suggesting that recruitment of the SETDB1 complex to K14ac/K9me regions has a role in silencing of active genomic regions.

Knockdown of Trio by CRISPR/Cas9 suppresses migration and invasion of cervical cancer cells.

Triple functional domain protein (Trio) is an evolutionarily conserved protein with guanine nucleotide exchange factors that regulate different physiological processes in some types of cancer. However, the expression and function of Trio in cervical cancer are still unknown. The purpose of this study was to detect the expression of Trio in cervical cancer tissues and to evaluate its clinical value. Furthermore, the effects of the Trio on the migration and invasion of cervical cancer cells and its mechanism were investigated invitro. The results of the present study revealed that Trio expression levels were significantly higher in most of the clinical cervical cancer samples than in adjacent tissues. The clinicopathological significance of Trio expression was also analyzed, and the results revealed that high expression levels in cervical cancer were correlated with lymph node metastasis (P=0.005). The CRISPR/Cas9 system was used to knockdown the endogenous Trio. The inhibition of Trio significantly decreased the migration and invasion abilities of cervical cancer cells. Meanwhile, levels of RhoA/ROCK signaling factors (RhoA, Rock, and p-LIMK), which contributed to cell migration and invasion, were decreased along with the inhibition of Trio. Therefore, Trio may regulate the migration and invasion of cervical cancer through the RhoA/ROCK signaling pathway.

Mannose-binding lectin and its roles in immune responses in grass carp (Ctenopharyngodon idella) against Aeromonas hydrophila

The complement system is a crucial component of the innate immune system that links innate and adaptive immunity via four pathways. Mannose-binding lectin (MBL), the initiating molecule of the lectin pathway, plays a significant role in the innate immune system in mammals and fish. Herein, we identified an MBL homolog (gcMBL) in grass carp (Ctenopharyngodon idella). The full-length 948 bp gcMBL cDNA includes a 741 bp open reading frame encoding a 246 amino acid protein with a signal peptide, collagen triple helix repeat domain, and a C-type lectin-like/link domain. The gcMBL protein shares low similarity with MBL counterparts in other species, and is most closely related to Cyprinus carpio MBL. Transcription of gcMBL was widely distributed in different tissues, and was induced by Aeromonas hydrophila in vivo and in vitro. Expression of gcMBL was also affected by LPS and flagellin stimulation in vitro. In cells over-expressing gcMBL, transcripts of almost all components except gcC5 were up-regulated, and gcMBL, gcIL1β, gcTNF-α, gcIFN, gcCD59, gcC5aR and gcITGβ-2 were significantly up-regulated following exposure to A. hydrophila or stimulation by bacterial PAMPs. Meanwhile, gcMBL deficiency achieved by RNAi down-regulated transcript levels following A. hydrophila challenge, and gcMBL induced NF-κB signalling. These findings indicate a vital role of gcMBL in innate immunity in grass carp.

Neurodevelopmental disease-associated de novo mutations and rare sequence variants affect TRIO GDP/GTP exchange factor activity.

Bipolar disorder, schizophrenia, autism and intellectual disability are complex neurodevelopmental disorders, debilitating millions of people. Therapeutic progress is limited by poor understanding of underlying molecular pathways. Using a targeted search, we identified an enrichment of de novo mutations in the gene encoding the 330-kDa triple functional domain (TRIO) protein associated with neurodevelopmental disorders. By generating multiple TRIO antibodies, we show that the smaller TRIO9 isoform is the major brain protein product, and its levels decrease after birth. TRIO9 contains two guanine nucleotide exchange factor (GEF) domains with distinct specificities: GEF1 activates both Rac1 and RhoG; GEF2 activates RhoA. To understand the impact of disease-associated de novo mutations and other rare sequence variants on TRIO function, we utilized two FRET-based biosensors: a Rac1 biosensor to study mutations in TRIO (T)GEF1, and a RhoA biosensor to study mutations in TGEF2. We discovered that one autism-associated de novo mutation in TGEF1 (K1431M), at the TGEF1/Rac1 interface, markedly decreased its overall activity toward Rac1. A schizophrenia-associated rare sequence variant in TGEF1 (F1538Intron) was substantially less active, normalized to protein level and expressed poorly. Overall, mutations in TGEF1 decreased GEF1 activity toward Rac1. One bipolar disorder-associated rare variant (M2145T) in TGEF2 impaired inhibition by the TGEF2 pleckstrin-homology domain, resulting in dramatically increased TGEF2 activity. Overall, genetic damage to both TGEF domains altered TRIO catalytic activity, decreasing TGEF1 activity and increasing TGEF2 activity. Importantly, both GEF changes are expected to decrease neurite outgrowth, perhaps consistent with their association with neurodevelopmental disorders.

Crystal Structure of the Carboxy-Terminal Region of the Bacteriophage T4 Proximal Long Tail Fiber Protein Gp34.

Long tail fibers of bacteriophage T4 are formed by proteins gp34, gp35, gp36, and gp37, with gp34 located at the phage-proximal end and gp37 at the phage-distal, receptor-binding end. We have solved the structure of the carboxy-terminal region of gp34, consisting of amino acids 894–1289, by single-wavelength anomalous diffraction and extended the structure to amino acids 744–1289 using data collected from crystals containing longer gp34-fragments. The structure reveals three repeats of a mixed α-β fibrous domain in residues 744 to 877. A triple-helical neck connects to an extended triple β-helix domain (amino acids 900–1127) punctuated by two β-prism domains. Next, a β-prism domain decorated with short helices and extended β-helices is present (residues 1146–1238), while the C-terminal end is capped with another short β-helical region and three β-hairpins. The structure provides insight into the stability of the fibrous gp34 protein.

The G2385R risk factor for Parkinson's disease enhances CHIP-dependent intracellular degradation of LRRK2.

Autosomal dominant mutations in leucine-rich repeat kinase 2 (LRRK2) are associated with Parkinson's disease (PD). Most pathogenic LRRK2 mutations result in amino acid substitutions in the central ROC (Ras of complex proteins)-C-terminus of ROC-kinase triple domain and affect enzymatic functions of the protein. However, there are several variants in LRRK2, including the risk factor G2385R, that affect PD pathogenesis by unknown mechanisms. Previously, we have shown that G2385R LRRK2 has decreased kinase activity in vitro and altered affinity to LRRK2 interactors. Specifically, we found an increased binding to the chaperone Hsp90 (heat shock protein 90 kDa) that is known to stabilize LRRK2, suggesting that G2385R may have structural effects on LRRK2. In the present study, we further explored the effects of G2385R on LRRK2 in cells. We found that G2385R LRRK2 has lower steady-state intracellular protein levels compared with wild-type LRRK2 due to increased protein turnover of the mutant protein. Mechanistically, this is a consequence of a higher affinity of G2385R compared with the wild-type protein for two proteins involved in proteasomal degradation, Hsc70 and carboxyl-terminus of Hsc70-interacting protein (CHIP). Overexpression of CHIP decreased intracellular protein levels of both G2385R mutant and wild-type LRRK2, while short interfering RNA CHIP knockdown had the opposite effect. We suggest that the G2385R substitution tilts the equilibrium between refolding and proteasomal degradation toward intracellular degradation. The observation of lower steady-state protein levels may explain why G2385R is a risk factor rather than a penetrant variant for inherited PD.

A thermodynamic description of metastable c-TiAlZrN coatings with triple spinodally decomposed domains

A critical thermodynamic assessment of the metastable c-TiAlZrN coatings, which are reported to spinodally decompose into triple domains, i.e., c-TiN, c-AlN, and c-ZrN, was performed via the CALculation of PHAse Diagram (CALPHAD) technique based on the limited experimental data as well as the first-principles computed free energies. The metastable c-TiAlZrN coatings were modeled as a pseudo-ternary phase consisting of c-TiN, c-AlN and c-ZrN species, and described using the substitutional solution model. The thermodynamic descriptions for the three boundary binaries were directly taken from either the CALPHAD assessment or the first-principles results available in the literature except for a re-adjustment of the pseudo-binary c-AlN/c-ZrN system based on the experimental phase equilibria in the pseudo-ternary system. The good agreement between the calculated phase equilibria and the experimental data over the wide temperature range was obtained, validating the reliability of the presently obtained thermodynamic descriptions for the c-TiAlZrN system. Based on the present thermodynamic description, different phase diagrams and thermodynamic properties can be easily predicted. It is anticipated that the present thermodynamic description of the metastable c-TiAlZrN coatings can serve as the important input for the later quantitative description of the microstructure evolution during service life.

Recombinant Human Serum Albumin Containing 3 Copies of Domain I, Has Significant <i>in Vitro</i> Antioxidative Capacity Compared to the Wild-Type

Human serum albumin (HSA), the most abundant protein in serum, functions as carrier of drugs and contributes to maintaining serum colloid osmotic pressure. We report herein on the preparation of a genetic recombinant HSA, in which domains II and III were changed to domain I (triple domain I; TDI). Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) results indicated that the purity of the TDI was equivalent to that of the wild type (WT). Both far- and near-UV circular dichroism (CD) spectra of the TDI showed that its structural characteristics were similar to the WT. Ligand binding capacity was examined by an ultrafiltration method using 3-carboxy-4-methyl-5-propyl-2-furanpropanoic acid (CMPF) and ketoprofen as markers for site I and site II, respectively. The binding capacity of TDI for both ligands was lower than that for the wild type. TDI significantly suppressed the oxidation of dihydrorhodamine 123 (DRD) by H2O2 compared to the WT. Our current results suggest that TDI has great potential for further development as HSA a product having antioxidative functions.

Primary care interventions to improve transition of youth with chronic health conditions from paediatric to adult healthcare: a systematic review

ObjectiveTo determine effective interventions to improve primary care provider involvement in transitioning youth with chronic conditions from paediatric to adult care.DesignSystematic review. Multiple electronic databases were searched including Ovid MEDLINE,...

Sensitivity of PBL schemes of the WRF-ARW model in simulating the boundary layer flow parameters for their application to air pollution dispersion modeling over a tropical station

Mesoscale atmospheric circulations play an important role in the transport of air pollution and local air quality issues. The planetary boundary layer (PBL), the thermo-dymamical structure and the flow field play an important role in air pollution dispersion. Hence, the PBL parameters over Nagpur, India are simulated using the ARW v. 3.6.1 mesoscale model. High-resolution simulations are conducted with triple nested domains having a horizontal resolution of 27, 9 and 3km, as well as 27 vertical levels by using the 1×1° NCEP Final Analysis meteorological fields for initial and boundary conditions. Eight fair-weather days in winter and summer (January and April 2009) with no significant synoptic activity were chosen for the study. Sensitivity experiments of the ARW model were conducted with two non-local (Yonsei University [YSU], and Asymmetric Convective Model v. 2 [ACM2]) and three local turbulence kinetic energy (TKE) closure (Mellor-Yamada Nakanishi and Niino Level 2.5 PBL [MYNN2], Mellor-Yamada-Janjic [MYJ], and quasi-normal scale elimination [QNSE]) turbulence diffusion parameterizations, to study the evolution of PBL parameters and the thermodynamical structure during the study period. After validation of the simulated parameters with the available in situ data, it was revealed that the non-local PBL scheme YSU, followed by local scheme MYNN2, could be able to capture the characteristic variations of surface meteorological variables and the thermodynamical structure of the atmosphere. The present results suggest that PBL schemes, namely YSU and MYNN2, performed better in representing the boundary-layer parameters and are useful for air pollution dispersion studies.

The High Affinity Binding Site on Plasminogen Activator Inhibitor-1 (PAI-1) for the Low Density Lipoprotein Receptor-related Protein (LRP1) Is Composed of Four Basic Residues

Plasminogen activator inhibitor 1 (PAI-1) is a serpin inhibitor of the plasminogen activators urokinase-type plasminogen activator (uPA) and tissue plasminogen activator, which binds tightly to the clearance and signaling receptor low density lipoprotein receptor-related protein 1 (LRP1) in both proteinase-complexed and uncomplexed forms. Binding sites for PAI-1 within LRP1 have been localized to CR clusters II and IV. Within cluster II, there is a strong preference for the triple CR domain fragment CR456. Previous mutagenesis studies to identify the binding site on PAI-1 for LRP1 have given conflicting results or implied small binding contributions incompatible with the high affinity PAI-1/LRP1 interaction. Using a highly sensitive solution fluorescence assay, we have examined binding of CR456 to arginine and lysine variants of PAI-1 and definitively identified the binding site as composed of four basic residues, Lys-69, Arg-76, Lys-80, and Lys-88. These are highly conserved among mammalian PAI-1s. Individual mutations result in a 13-800-fold increase in Kd values. We present evidence that binding involves engagement of CR4 by Lys-88, CR5 by Arg-76 and Lys-80, and CR6 by Lys-69, with the strongest interactions to CR5 and CR6. Collectively, the individual binding contributions account quantitatively for the overall PAI-1/LRP1 affinity. We propose that the greater efficiency of PAI-1·uPA complex binding and clearance by LRP1, compared with PAI-1 alone, is due solely to simultaneous binding of the uPA moiety in the complex to its receptor, thereby making binding of the PAI-1 moiety to LRP1 a two-dimensional surface-localized association.

A novel C-type lectin with triple carbohydrate recognition domains has critical roles for the hard tick Haemaphysalis longicornis against Gram-negative bacteria

C-type lectins (CLecs) play an important role in innate immunity against invaders. In this study, a novel CLec was identified from Haemaphysalis longicornis ticks (HlCLec). HlCLec contains a signal peptide and a transmembrane region. Interestingly, HlCLec possesses three dissimilar carbohydrate recognition domains (CRDs). Each CRD contains the mutated motif of Ca2+-binding site 2. HlCLec mRNA was up-regulated during blood feeding, and had highest expression in the midgut and ovary. HlCLec localization was also confirmed by immunofluorescent antibody test (IFAT). HlCLec was found on the cell membrane and basal lamina of midgut and ovary. In addition, the recombinant HlCLec and individual CRDs demonstrated direct binding activity to Escherichia coli and Staphylococcus aureus; however, no growth inhibition activity was observed. Furthermore, E. coli injection after silencing of HlCLec caused drastic reduction in survival rate of ticks. These results strongly suggest the key role of HlCLec in tick innate immunity against Gram-negative bacteria.

Wavelets and triple difference as a mathematical method for filtering and mitigation of DGPS errors

The accuracy of GPS position is primarily dependent on the satellite position, signal delay, and various environment noises such as ionospheric delay effects, ephemeris errors, satellite clock errors, multi-path distortion, tropospheric delay effects, and numerical errors. To overcome most of these errors the Differential Global Positioning System (DGPS) is used. Differential Global Positioning System is one of the most popular and reliable techniques for improving GPS accuracy by minimizing correlated errors between reference stations and rovers. The primary motivation of this paper is to improve GPS measurement accuracies by classifying errors in the frequency domain and subsequently separating, mitigating and decreasing them.Wavelet spectral techniques can separate GPS signals into sub-bands where different errors can be separated and mitigated. The main goal of this paper was the development and implementation of DGPS error mitigation techniques using triple difference and wavelet. This paper studies, analyzes and provides new techniques that will help mitigate these errors in the frequency domain. The proposed technique applied to smooth noise for GPS receiver positioning data is based upon the analysis of wavelet transform (WT). The technique is applied using wavelet as a de-noising tool to tackle the high-frequency errors in the triple difference domain and to obtain a de-noised triple difference signal that can be used in a positioning calculation.

Upregulated TRIO expression correlates with a malignant phenotype in human hepatocellular carcinoma.

Triple functional domain protein (TRIO) is an evolutionarily conserved Dbl family guanine nucleotide exchange factors (GEFs) involved in cell proliferation and progression of some types of cancer. However, the expression and prognostic role of TRIO in hepatocellular carcinoma (HCC) have not yet been determined. Therefore, we attempted to determine the impact of TRIO on the clinical outcome of HCC patients to further identify its role in HCC. TRIO expression was examined using quantitative real-time PCR (qRT-PCR) and Western blotting in nonmalignant liver cells, HCC cells, and 93 paired of HCC tissues and adjacent noncancerous tissues. Statistical analyses were used to assess associations between TRIO expression and clinicopathological and prognostic factors. Small interfering RNA (siRNA)-mediated TRIO inhibition was performed in Hep3B and Huh7 cells to elucidate its roles in HCC. 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay was employed to measure cell proliferation, and apoptosis assay was analyzed by flow cytometry, respectively. Adhesion and transwell invasion assay were performed to determine the invasion ability of HCC cells in vitro. TRIO was significantly upregulated in the HCC cell lines and tissues compared with the nonmalignant liver cells and adjacent noncancerous liver tissues. In addition, high TRIO expression level associated with lymph node metastasis (P = 0.0183), clinical tumor node metastasis (TNM) stage (P = 0.0.0106), and decrease in overall survival (OS) (P = 0.017). Knockdown of TRIO on Hep3B and Huh7 cell lines suppressed cell proliferation and migration and induced apoptosis. Furthermore, silencing TRIO expression led to decrease of ras-related C3 botulinum toxin substrate 1 (Rac1), p-P38, B cell lymphoma 2 (BCL-2), and matrix metallopeptidase 9 (MMP-9). Our results demonstrated that TRIO protein expression is elevated and associated with a worse over survival rates in patients with HCC. Aberrant expression of TRIO might play an important role in HCC through promoting cell proliferation and invasion, and TRIO may be a novel therapeutic target for the treatment of HCC.

Head-to-head domain wall structures in wide permalloy strips

We analyze the equilibrium micromagnetic domain wall structures encountered in Permalloy strips of a wide range of thicknesses and widths, with strip widths up to several micrometers. By performing an extensive set of micromagnetic simulations, we show that the equilibrium phase diagram of the domain wall structures exhibits in addition to the previously found structures (symmetric and asymmetric transverse walls, vortex wall) also double vortex and triple vortex domain walls for large enough strip widths and thicknesses. Also several metastable domain wall structures are found for wide and/or thick strips. We discuss the details of the relaxation process from random magnetization initial states towards the stable domain wall structure, and show that our results are robust with respect to changes of e.g. the magnitude of the Gilbert damping constant and details of the initial conditions.

Structural characterization of triple transmembrane domain containing fragments of a yeast G protein-coupled receptor in an organic : aqueous environment by solution-state NMR spectroscopy.

This report summarizes recent biophysical and protein expression experiments on polypeptides containing the N-terminus, the first, second, and third transmembrane (TM) domains and the contiguous loops of the α-factor receptor Ste2p, a G protein-coupled receptor. The 131-residue polypeptide Ste2p(G31-R161), TM1-TM3, was investigated by solution NMR in trifluoroethanol/water. TM1-TM3 contains helical TM domains at the predicted locations, supported by continuous sets of medium-range NOEs. In addition, a short helix N-terminal to TM1 was detected, as well as a short helical stretch in the first extracellular loop. Two 161-residue polypeptides, [Ste2p(M1-R161), NT-TM1-TM3], that contain the entire N-terminal sequence, one with a single mutation, were directly expressed and isolated from Escherichia coli in yields as high as 30 mg/L. Based on its increased stability, the L11P mutant will be used in future experiments to determine long-range interactions. The study demonstrated that 3-TM domains of a yeast G protein-coupled receptor can be produced in isotopically labeled form suitable for solution NMR studies. The quality of spectra is superior to data recorded in micelles and allows more rapid data analysis. No tertiary contacts have been determined, and if present, they are likely transient. This observation supports earlier studies by us that secondary structure was retained in smaller fragments, both in organic solvents and in detergent micelles, but that stable tertiary contacts may only be present when the protein is imbedded in lipids.

A weather research and forecasting model evaluation for simulating heavy precipitation over the downstream area of the Yalong River Basin

The forecasting capability of the weather research and forecasting (WRF) model for heavy precipitation in the downstream area of the Yalong River Basin in Southwest China was evaluated for the first time through the simulation of three heavy precipitation events with seven commonly used microphysics parameterization schemes (MPS) (Kessler, Lin et al. (Lin), Single-Moment 3-class (WSM3), Single-Moment 5-class (WSM5), Ferrier, Single-Moment 6-class (WSM6), and New Thompson et al. (NTH)) and three cumulus parameterization schemes (CPS) (Kain-Fritsch (KF), Betts-Miller-Janjic (BMJ), and Grell-Devenyi (GD)). Of the three rainfall events, the first two are typical large-area heavy precipitation events in the Yalong River Basin and consist of several continuous storms. The third one is a heavy precipitation event with only one storm. In this study, a triple nested domain with a 3-km grid resolution inner-most domain over the study area was configured for the WRF model. We employed the probability of detection (POD), false alarm ratio (FAR), BIAS, and equitable threat (ET) scores to compare the spatial distribution of heavy rainfall created by the WRF model with the observations from the gauges in the downstream area of the river basin. The root mean squared errors (RMSEs) at each sub river basin and the whole downstream of Yalong River Basin were also calculated for the evaluation. In addition, it is important to include the computation efficiency when choosing a scheme combination. We recorded the time consumption for each model simulation and made comparisons for selecting the optimum scheme with less time consumption and acceptable prediction accuracy. Through comprehensive comparison, the scheme combination of WSM3 and GD holds a stable performance in leveraging the prediction accuracy and computation efficiency for the heavy precipitation events.