Structural and evolutionary adaptation of rhoptry kinases and pseudokinases, a family of coccidian virulence factors

BACKGROUND: The widespread protozoan parasite Toxoplasma gondii interferes with host cell functions by exporting the contents of a unique apical organelle, the rhoptry. Among the mix of secreted proteins are an expanded, lineage-specific family of protein kinases termed rhoptry kinases (ROPKs), several of which have been shown to be key virulence factors, including the pseudokinase ROP5. The extent and details of the diversification of this protein family are poorly understood. RESULTS: In this study, we comprehensively catalogued the ROPK family in the genomes of Toxoplasma gondii, Neospora caninum and Eimeria tenella, as well as portions of the unfinished genome of Sarcocystis neurona, and classified the identified genes into 42 distinct subfamilies. We systematically compared the rhoptry kinase protein sequences and structures to each other and to the broader superfamily of eukaryotic protein kinases to study the patterns of diversification and neofunctionalization in the ROPK family and its subfamilies. We identified three ROPK sub-clades of particular interest: those bearing a structurally conserved N-terminal extension to the kinase domain (NTE), an E. tenella-specific expansion, and a basal cluster including ROP35 and BPK1 that we term ROPKL. Structural analysis in light of the solved structures ROP2, ROP5, ROP8 and in comparison to typical eukaryotic protein kinases revealed ROPK-specific conservation patterns in two key regions of the kinase domain, surrounding a ROPK-conserved insert in the kinase hinge region and a disulfide bridge in the kinase substrate-binding lobe. We also examined conservation patterns specific to the NTE-bearing clade. We discuss the possible functional consequences of each. CONCLUSIONS: Our work sheds light on several important but previously unrecognized features shared among rhoptry kinases, as well as the essential differences between active and degenerate protein kinases. We identify the most distinctive ROPK-specific features conserved across both active kinases and pseudokinases, and discuss these in terms of sequence motifs, evolutionary context, structural impact and potential functional relevance. By characterizing the proteins that enable these parasites to invade the host cell and co-opt its signaling mechanisms, we provide guidance on potential therapeutic targets for the diseases caused by coccidian parasites

Friction Stir Welding of Aluminium Alloys

This chapter investigates on the characterization of friction stir welded dissimilar aluminium alloys AA2024 with AA5052, AA2024 with AA6061 and AA 5052 with AA6061. Five tool designs were employed with first two dissimilar combinations to analyze the influence of rotation and traverse speed over microstructural and mechanical properties. H13 tool steel was used as tool material with various pin profiles which includes cylindrical, cylindrical-threaded, squared, tapered and stepped types. In the dissimilar welding of AA 2024 with AA 5052, sound welds were produced with stepped pin tool. In the dissimilar welding of AA 2024 with AA 6061, ratio between tool shoulder to diameter of tool pin was the most influential factor. Welded joints failed in the Heat affected zone (HAZ) of 6061 where the hardness values were comparatively less. In dissimilar welding of AA 5052 with AA6061, cylindrical pin tool was used at a constant speed of 710 rpm and at different feed rates of 28 and 40 mm/min. Micro structural examination showed variation of grain size in every zone and their influence on mechanical properties. Correlating mechanical and metallurgical properties, the optimized process parameters of speed and feed were identified to be 710 rpm and 28 mm/min respectively for all attempted dissimilar combinations

Design and Implementation of Fuzzy Logic Controller for Online Computer Controlled Steering System for Navigation of a Teleoperated Agricultural Vehicle

This paper describes design, modeling, simulation, control, and implementation of teleoperated agricultural vehicle using intelligent technique. This vehicle can be used for ploughing, sowing, and soil moisture sensing. Online computer controlled steering system for a vehicle utilizing two independent drive wheels can be used to avoid obstacles and to improve the ability to resist external side forces. To control the steer angles of the nondriven wheels, the mathematical relationships between the drive wheel speeds and the steer angles of the nondriven wheels are used. A fuzzy logic controller is designed to change the drive wheel speeds and to achieve the desired steer angles. Online control of the agricultural vehicle is achieved from a remote place by means of Web Publishing Tool in LabVIEW. IR sensors in the vehicle are used to detect and to avoid the obstacles around. The developed steering angle control algorithm and fuzzy logic controller have been implemented in an agricultural vehicle which depicts that the vehicle performs its operation efficiently and reduces the manpower and becomes advantageous

An evolutionary perspective on the kinome of malaria parasites

Malaria parasites belong to an ancient lineage that diverged very early from the main branch of eukaryotes. The approximately 90-member plasmodial kinome includes a majority of eukaryotic protein kinases that clearly cluster within the AGC, CMGC, TKL, CaMK and CK1 groups found in yeast, plants and mammals, testifying to the ancient ancestry of these families. However, several hundred millions years of independent evolution, and the specific pressures brought about by first a photosynthetic and then a parasitic lifestyle, led to the emergence of unique features in the plasmodial kinome. These include taxon-restricted kinase families, and unique peculiarities of individual enzymes even when they have homologues in other eukaryotes. Here, we merge essential aspects of all three malaria-related communications that were presented at the Evolution of Protein Phosphorylation meeting, and propose an integrated discussion of the specific features of the parasite's kinome and phosphoproteome

Role of diagnostic hystero-laparoscopy in the evaluation of female infertility

Background: Infertility affects about 10-15% of reproductive age couples. The main causes of infertility include male factor, ovulatory disorders, tubal factor and endometriosis. This study was conducted to determine the role of Diagnostic hystero-laparoscopy (DHL) in the evaluation of female infertility.Methods: This study was a prospective study done in a tertiary care centre over a period of 1 year on all infertile couples. The exclusion criteria: male factor infertility, hypothyroidism, hyperprolactinemia, acute pelvic inflammatory disease. Diagnostic hysterolaparoscopy was performed in all study patients in pre ovulatory phase, under general anaesthesia. The findings were then documented and analysed.Results: A total number of 90 patients were recruited for the study. The mean age of the patients was 28±3 years. The mean BMI of the study patients was 25±4 kg/m2. The mean duration of infertility was 5.4 years. On laparoscopy, 28 patients had polycystic ovaries (31.1%), 4 patients had pelvic adhesions (4.4%), 2 patients had endometriosis (2.2%) and tubal block was identified in 10 patients (11.11%). 10 patients had abnormal findings on hysteroscopy.Conclusions: Hystero laparoscopy is an effective diagnostic tool in the evaluation of infertility and has to be included in basic diagnostic workup

Assessment of genetic and functional diversity of phosphate solubilizing fluorescent pseudomonads isolated from rhizospheric soil

<p>Abstract</p> <p>Background</p> <p>Phosphorus is an essential macronutrient for the growth of plants. However, in most soils a large portion of phosphorus becomes insoluble and therefore, unavailable to plants. Knowledge on biodiversity of phosphate-solubilizing fluorescent pseudomonads is essential to understand their ecological role and their utilization in sustainable agriculture.</p> <p>Results</p> <p>Of 443 fluorescent pseudomonad strains tested, 80 strains (18%) showed positive for the solubilization of tri-calcium phosphate (Ca₃(PO₄)₂) by the formation of visible dissolution halos on Pikovskaya's agar. These phosphate solubilizing strains showed high variability in utilizing various carbon sources. Numerical taxonomy of the phosphate solubilizing strains based on their carbon source utilization profiles resulted into three major phenons at a 0.76 similarity coefficient level. Genotypic analyses of strains by BOX (bacterial repetitive BOX element)-polymerase chain reaction (PCR) resulted into three distinct genomic clusters and 26 distinct BOX profiles at a 80% similarity level. On the basis of phenotypic characterization and <it>16S rRNA </it>gene phylogenetic analyses strains were identified as <it>Pseudomonas aeruginosa, P. mosselii, P. monteilii, P. plecoglossicida, P. putida, P. fulva </it>and <it>P. fluorescens</it>. These phosphate solubilizing strains also showed the production of plant growth promoting enzymes, hormones and exhibited antagonism against phytopathogenic fungi that attack on various crops. Gene specific primers have identified the putative antibiotic producing strains. These putative strains were grown in fermentation media and production of antibiotics was confirmed by thin layer chromatography (TLC) and high performance liquid chromatography (HPLC).</p> <p>Conclusion</p> <p>Present study revealed a high degree of functional and genetic diversity among the phosphate solubilizing fluorescent pseudomonad bacteria. Due to their innate potential of producing an array of plant growth promoting enzymes, hormones and antifungal metabolites these phosphate solubilizing strains are considered to play a vital role in plant growth promotion, disease suppression and subsequent enhancement of yield.</p

Cellular expression and crystal structure of the murine cytomegalovirus MHC-Iv glycoprotein, m153

Mouse cytomegalovirus (MCMV), a β-herpesvirus that establishes latent and persistent infections in mice, is a valuable model for studying complex virus-host interactions. MCMV encodes the m145 family of putative immunoevasins with predicted MHC-I structure. Functions attributed to some family members include downregulation of host MHC-I (m152) and NKG2D ligands (m145, m152, m155) and interaction with inhibitory or activating NK receptors (m157). We present the cellular, biochemical and structural characterization of m153, which is a heavily glycosylated homodimer, that does not require β2m or peptide, and is expressed at the surface of MCMV-infected cells. Its 2.4 Å crystal structure confirms that this compact molecule preserves an MHC-I-like fold and reveals a novel mode of dimerization, confirmed by site-directed mutagenesis, and a distinctive disulfide-stabilized extended amino terminus. The structure provides a useful framework for comparative analysis of the divergent members of the m145 family

Dynamics of Industrial Buyer Behavior of Loader Backhoe

ABSTRACT: To investigate how the buying behavior of firms work is always needed, since it is an every changing process influenced by many forces in its surroundings. Industrial firms tend to have a smaller number of customers which means that they are easier to identify than customers in consumer markets. Effective marketing strategy relies on understanding consumers and their needs. This understanding must include the industrial buyer if industrial marketing efforts are to be well planned and executed. This study is focused on understanding the buyer behavior of loader backhoe equipment&apos;s among Indian consumers. Primary data was collected from 387 buyers with the help of self-administered questionnaires. The hypotheses are tested with the help of chi-square, regression and ANOVA. From the data analysis it is concluded that the brand selection by the buyers of loader backhoe will influenced by brand awareness, past experience, Reliability, Resale value, Productivity, Service parts and network and less downtime. On considering the various brands JCB has good mind share and market share on loader backhoe equipment&apos;s

Autoantibodies in Scurfy Mice and IPEX Patients Recognize Keratin 14

Scurfy mice have a deletion in the Foxp3 gene, resulting in a failure to generate Foxp3+ regulatory T cells, and they subsequently develop severe CD4+ T-cell-mediated autoimmune inflammation. Multiple organs are involved, but the skin is one of the main organs affected. During the course of disease, Scurfy mice develop autoantibodies; however, the targeted antigens are unknown. In this study, we show that Scurfy mice develop autoantibodies directed against skin antigens. Using western blot analysis, we found that Scurfy serum reacted with proteins in total skin lysate, as well as in a keratinocyte lysate. Most of the Scurfy sera tested identified a major band at 50kDa. Transfer of Scurfy CD4+ T cells into nu/nu mice yielded autoantibodies with similar reactivity. Further analysis using 2D western blots, followed by peptide mass fingerprinting, identified several keratins as targets. To confirm this observation, we chose one of the identified targets, keratin 14, and prepared recombinant proteins encompassing the N-terminal, middle, and C-terminal portions of the keratin 14 protein. Scurfy serum predominantly recognized the C-terminal fragment. Sera from patients with immunodysregulation, polyendocrinopathy, enteropathy, X-linked (IPEX) syndrome, the human disease resulting from FOXP3 mutations, also recognized skin antigens, including keratin 14. Thus, the results of our study indicate that autoantibodies in Scurfy mice and patients with IPEX target keratins