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Sample records for protein features involved

  1. Structural features of protein folding nuclei.

    PubMed

    Garbuzynskiy, S O; Kondratova, M S

    2008-03-05

    A crucial event of protein folding is the formation of a folding nucleus. We demonstrate the presence of a considerable coincidence between the location of folding nuclei and the location of so-called "root structural motifs", which have unique overall folds and handedness. In the case of proteins with a single root structural motif, the involvement in the formation of a folding nucleus is in average significantly higher for amino acids residues that are in root structural motifs, compared to residues in other parts of the protein. The tests carried out revealed that the observed difference is statistically reliable. Thus, a structural feature that corresponds to the protein folding nucleus is now found.

  2. Evaporative cooling feature selection for genotypic data involving interactions

    PubMed Central

    McKinney, B.A.; Reif, D.M.; White, B.C.; Crowe, J.E.; Moore, J.H.

    2007-01-01

    Motivation: The development of genome-wide capabilities for genotyping has led to the practical problem of identifying the minimum subset of genetic variants relevant to the classification of a phenotype. This challenge is especially difficult in the presence of attribute interactions, noise and small sample size. Methods: Analogous to the physical mechanism of evaporation, we introduce an evaporative cooling (EC) feature selection algorithm that seeks to obtain a subset of attributes with the optimum information temperature (i.e. the least noise). EC uses an attribute quality measure analogous to thermodynamic free energy that combines Relief-F and mutual information to evaporate (i.e. remove) noise features, leaving behind a subset of attributes that contain DNA sequence variations associated with a given phenotype. Results: EC is able to identify functional sequence variations that involve interactions (epistasis) between other sequence variations that influence their association with the phenotype. This ability is demonstrated on simulated genotypic data with attribute interactions and on real genotypic data from individuals who experienced adverse events following smallpox vaccination. The EC formalism allows us to combine information entropy, energy and temperature into a single information free energy attribute quality measure that balances interaction and main effects. Availability: Open source software, written in Java, is freely available upon request. Contact: brett.mckinney@gmail.com PMID:17586549

  3. Singular Features of Trypanosomatids' Phosphotransferases Involved in Cell Energy Management

    PubMed Central

    Pereira, Claudio A.; Bouvier, León A.; Cámara, María de los Milagros; Miranda, Mariana R.

    2011-01-01

    Trypanosomatids are responsible for economically important veterinary affections and severe human diseases. In Africa, Trypanosoma brucei causes sleeping sickness or African trypanosomiasis, while in America, Trypanosoma cruzi is the etiological agent of Chagas disease. These parasites have complex life cycles which involve a wide variety of environments with very different compositions, physicochemical properties, and availability of metabolites. As the environment changes there is a need to maintain the nucleoside homeostasis, requiring a quick and regulated response. Most of the enzymes required for energy management are phosphotransferases. These enzymes present a nitrogenous group or a phosphate as acceptors, and the most clear examples are arginine kinase, nucleoside diphosphate kinase, and adenylate kinase. Trypanosoma and Leishmania have the largest number of phosphotransferase isoforms ever found in a single cell; some of them are absent in mammals, suggesting that these enzymes are required in many cellular compartments associated to different biological processes. The presence of such number of phosphotransferases support the hypothesis of the existence of an intracellular enzymatic phosphotransfer network that communicates the spatially separated intracellular ATP consumption and production processes. All these unique features make phosphotransferases a promising start point for rational drug design for the treatment of human trypanosomiasis. PMID:21603267

  4. Prediction of DNA-binding proteins from relational features

    PubMed Central

    2012-01-01

    Background The process of protein-DNA binding has an essential role in the biological processing of genetic information. We use relational machine learning to predict DNA-binding propensity of proteins from their structures. Automatically discovered structural features are able to capture some characteristic spatial configurations of amino acids in proteins. Results Prediction based only on structural relational features already achieves competitive results to existing methods based on physicochemical properties on several protein datasets. Predictive performance is further improved when structural features are combined with physicochemical features. Moreover, the structural features provide some insights not revealed by physicochemical features. Our method is able to detect common spatial substructures. We demonstrate this in experiments with zinc finger proteins. Conclusions We introduced a novel approach for DNA-binding propensity prediction using relational machine learning which could potentially be used also for protein function prediction in general. PMID:23146001

  5. A Novel Feature Extraction Method with Feature Selection to Identify Golgi-Resident Protein Types from Imbalanced Data

    PubMed Central

    Yang, Runtao; Zhang, Chengjin; Gao, Rui; Zhang, Lina

    2016-01-01

    The Golgi Apparatus (GA) is a major collection and dispatch station for numerous proteins destined for secretion, plasma membranes and lysosomes. The dysfunction of GA proteins can result in neurodegenerative diseases. Therefore, accurate identification of protein subGolgi localizations may assist in drug development and understanding the mechanisms of the GA involved in various cellular processes. In this paper, a new computational method is proposed for identifying cis-Golgi proteins from trans-Golgi proteins. Based on the concept of Common Spatial Patterns (CSP), a novel feature extraction technique is developed to extract evolutionary information from protein sequences. To deal with the imbalanced benchmark dataset, the Synthetic Minority Over-sampling Technique (SMOTE) is adopted. A feature selection method called Random Forest-Recursive Feature Elimination (RF-RFE) is employed to search the optimal features from the CSP based features and g-gap dipeptide composition. Based on the optimal features, a Random Forest (RF) module is used to distinguish cis-Golgi proteins from trans-Golgi proteins. Through the jackknife cross-validation, the proposed method achieves a promising performance with a sensitivity of 0.889, a specificity of 0.880, an accuracy of 0.885, and a Matthew’s Correlation Coefficient (MCC) of 0.765, which remarkably outperforms previous methods. Moreover, when tested on a common independent dataset, our method also achieves a significantly improved performance. These results highlight the promising performance of the proposed method to identify Golgi-resident protein types. Furthermore, the CSP based feature extraction method may provide guidelines for protein function predictions. PMID:26861308

  6. A Novel Feature Extraction Method with Feature Selection to Identify Golgi-Resident Protein Types from Imbalanced Data.

    PubMed

    Yang, Runtao; Zhang, Chengjin; Gao, Rui; Zhang, Lina

    2016-02-06

    The Golgi Apparatus (GA) is a major collection and dispatch station for numerous proteins destined for secretion, plasma membranes and lysosomes. The dysfunction of GA proteins can result in neurodegenerative diseases. Therefore, accurate identification of protein subGolgi localizations may assist in drug development and understanding the mechanisms of the GA involved in various cellular processes. In this paper, a new computational method is proposed for identifying cis-Golgi proteins from trans-Golgi proteins. Based on the concept of Common Spatial Patterns (CSP), a novel feature extraction technique is developed to extract evolutionary information from protein sequences. To deal with the imbalanced benchmark dataset, the Synthetic Minority Over-sampling Technique (SMOTE) is adopted. A feature selection method called Random Forest-Recursive Feature Elimination (RF-RFE) is employed to search the optimal features from the CSP based features and g-gap dipeptide composition. Based on the optimal features, a Random Forest (RF) module is used to distinguish cis-Golgi proteins from trans-Golgi proteins. Through the jackknife cross-validation, the proposed method achieves a promising performance with a sensitivity of 0.889, a specificity of 0.880, an accuracy of 0.885, and a Matthew's Correlation Coefficient (MCC) of 0.765, which remarkably outperforms previous methods. Moreover, when tested on a common independent dataset, our method also achieves a significantly improved performance. These results highlight the promising performance of the proposed method to identify Golgi-resident protein types. Furthermore, the CSP based feature extraction method may provide guidelines for protein function predictions.

  7. Genes and Pathways Involved in Adult Onset Disorders Featuring Muscle Mitochondrial DNA Instability

    PubMed Central

    Ahmed, Naghia; Ronchi, Dario; Comi, Giacomo Pietro

    2015-01-01

    Replication and maintenance of mtDNA entirely relies on a set of proteins encoded by the nuclear genome, which include members of the core replicative machinery, proteins involved in the homeostasis of mitochondrial dNTPs pools or deputed to the control of mitochondrial dynamics and morphology. Mutations in their coding genes have been observed in familial and sporadic forms of pediatric and adult-onset clinical phenotypes featuring mtDNA instability. The list of defects involved in these disorders has recently expanded, including mutations in the exo-/endo-nuclease flap-processing proteins MGME1 and DNA2, supporting the notion that an enzymatic DNA repair system actively takes place in mitochondria. The results obtained in the last few years acknowledge the contribution of next-generation sequencing methods in the identification of new disease loci in small groups of patients and even single probands. Although heterogeneous, these genes can be conveniently classified according to the pathway to which they belong. The definition of the molecular and biochemical features of these pathways might be helpful for fundamental knowledge of these disorders, to accelerate genetic diagnosis of patients and the development of rational therapies. In this review, we discuss the molecular findings disclosed in adult patients with muscle pathology hallmarked by mtDNA instability. PMID:26251896

  8. Genes and Pathways Involved in Adult Onset Disorders Featuring Muscle Mitochondrial DNA Instability.

    PubMed

    Ahmed, Naghia; Ronchi, Dario; Comi, Giacomo Pietro

    2015-08-05

    Replication and maintenance of mtDNA entirely relies on a set of proteins encoded by the nuclear genome, which include members of the core replicative machinery, proteins involved in the homeostasis of mitochondrial dNTPs pools or deputed to the control of mitochondrial dynamics and morphology. Mutations in their coding genes have been observed in familial and sporadic forms of pediatric and adult-onset clinical phenotypes featuring mtDNA instability. The list of defects involved in these disorders has recently expanded, including mutations in the exo-/endo-nuclease flap-processing proteins MGME1 and DNA2, supporting the notion that an enzymatic DNA repair system actively takes place in mitochondria. The results obtained in the last few years acknowledge the contribution of next-generation sequencing methods in the identification of new disease loci in small groups of patients and even single probands. Although heterogeneous, these genes can be conveniently classified according to the pathway to which they belong. The definition of the molecular and biochemical features of these pathways might be helpful for fundamental knowledge of these disorders, to accelerate genetic diagnosis of patients and the development of rational therapies. In this review, we discuss the molecular findings disclosed in adult patients with muscle pathology hallmarked by mtDNA instability.

  9. Van der Waals Interactions Involving Proteins

    NASA Technical Reports Server (NTRS)

    Roth, Charles M.; Neal, Brian L.; Lenhoff, Abraham M.

    1996-01-01

    Van der Waals (dispersion) forces contribute to interactions of proteins with other molecules or with surfaces, but because of the structural complexity of protein molecules, the magnitude of these effects is usually estimated based on idealized models of the molecular geometry, e.g., spheres or spheroids. The calculations reported here seek to account for both the geometric irregularity of protein molecules and the material properties of the interacting media. Whereas the latter are found to fall in the generally accepted range, the molecular shape is shown to cause the magnitudes of the interactions to differ significantly from those calculated using idealized models. with important consequences. First, the roughness of the molecular surface leads to much lower average interaction energies for both protein-protein and protein-surface cases relative to calculations in which the protein molecule is approximated as a sphere. These results indicate that a form of steric stabilization may be an important effect in protein solutions. Underlying this behavior is appreciable orientational dependence, one reflection of which is that molecules of complementary shape are found to exhibit very strong attractive dispersion interactions. Although this has been widely discussed previously in the context of molecular recognition processes, the broader implications of these phenomena may also be important at larger molecular separations, e.g., in the dynamics of aggregation, precipitation, and crystal growth.

  10. Van der Waals interactions involving proteins.

    PubMed Central

    Roth, C M; Neal, B L; Lenhoff, A M

    1996-01-01

    Van der Waals (dispersion) forces contribute to interactions of proteins with other molecules or with surfaces, but because of the structural complexity of protein molecules, the magnitude of these effects is usually estimated based on idealized models of the molecular geometry, e.g., spheres or spheroids. The calculations reported here seek to account for both the geometric irregularity of protein molecules and the material properties of the interacting media. Whereas the latter are found to fall in the generally accepted range, the molecular shape is shown to cause the magnitudes of the interactions to differ significantly from those calculated using idealized models, with important consequences. First, the roughness of the molecular surface leads to much lower average interaction energies for both protein-protein and protein-surface cases relative to calculations in which the protein molecule is approximated as a sphere. These results indicate that a form of steric stabilization may be an important effect in protein solutions. Underlying this behavior is appreciable orientational dependence, one reflection of which is that molecules of complementary shape are found to exhibit very strong attractive dispersion interactions. Although this has been widely discussed previously in the context of molecular recognition processes, the broader implications of these phenomena may also be important at larger molecular separations, e.g., in the dynamics of aggregation, precipitation, and crystal growth. Images FIGURE 3 PMID:8789115

  11. Feature Fusion Based SVM Classifier for Protein Subcellular Localization Prediction.

    PubMed

    Rahman, Julia; Mondal, Md Nazrul Islam; Islam, Md Khaled Ben; Hasan, Md Al Mehedi

    2016-12-18

    For the importance of protein subcellular localization in different branches of life science and drug discovery, researchers have focused their attentions on protein subcellular localization prediction. Effective representation of features from protein sequences plays a most vital role in protein subcellular localization prediction specially in case of machine learning techniques. Single feature representation-like pseudo amino acid composition (PseAAC), physiochemical property models (PPM), and amino acid index distribution (AAID) contains insufficient information from protein sequences. To deal with such problems, we have proposed two feature fusion representations, AAIDPAAC and PPMPAAC, to work with Support Vector Machine classifiers, which fused PseAAC with PPM and AAID accordingly. We have evaluated the performance for both single and fused feature representation of a Gram-negative bacterial dataset. We have got at least 3% more actual accuracy by AAIDPAAC and 2% more locative accuracy by PPMPAAC than single feature representation.

  12. Visual Features Involving Motion Seen from Airport Control Towers

    NASA Technical Reports Server (NTRS)

    Ellis, Stephen R.; Liston, Dorion

    2010-01-01

    Visual motion cues are used by tower controllers to support both visual and anticipated separation. Some of these cues are tabulated as part of the overall set of visual features used in towers to separate aircraft. An initial analyses of one motion cue, landing deceleration, is provided as a basis for evaluating how controllers detect and use it for spacing aircraft on or near the surface. Understanding cues like it will help determine if they can be safely used in a remote/virtual tower in which their presentation may be visually degraded.

  13. Proteins involved in meiotic recombination: a role in male infertility?

    PubMed

    Sanderson, Matthew L; Hassold, Terry J; Carrell, Douglas T

    2008-01-01

    Meiotic recombination results in the formation of crossovers, by which genetic information is exchanged between homologous chromosomes during prophase I of meiosis. Recombination is a complex process involving many proteins. Alterations in the genes involved in recombination may result in infertility. Molecular studies have improved our understanding of the roles and mechanisms of the proteins and protein complexes involved in recombination, some of which have function in mitotic cells as well as meiotic cells. Human gene sequencing studies have been performed for some of these genes and have provided further information on the phenotypes observed in some infertile individuals. However, further studies are needed to help elucidate the particular role(s) of a given protein and to increase our understanding of these protein systems. This review will focus on our current understanding of proteins involved in meiotic recombination from a genomic perspective, summarizing our current understanding of known mutations and single nucleotide polymorphisms that may affect male fertility by altering meiotic recombination.

  14. Sturgeon Osteocalcin Shares Structural Features with Matrix Gla Protein

    PubMed Central

    Viegas, Carla S. B.; Simes, Dina C.; Williamson, Matthew K.; Cavaco, Sofia; Laizé, Vincent; Price, Paul A.; Cancela, M. Leonor

    2013-01-01

    Osteocalcin (OC) and matrix Gla protein (MGP) are considered evolutionarily related because they share key structural features, although they have been described to exert different functions. In this work, we report the identification and characterization of both OC and MGP from the Adriatic sturgeon, a ray-finned fish characterized by a slow evolution and the retention of many ancestral features. Sturgeon MGP shows a primary structure, post-translation modifications, and patterns of mRNA/protein distribution and accumulation typical of known MGPs, and it contains seven possible Gla residues that would make the sturgeon protein the most γ-carboxylated among known MGPs. In contrast, sturgeon OC was found to present a hybrid structure. Indeed, although exhibiting protein domains typical of known OCs, it also contains structural features usually found in MGPs (e.g. a putative phosphorylated propeptide). Moreover, patterns of OC gene expression and protein accumulation overlap with those reported for MGP; OC was detected in bone cells and mineralized structures but also in soft and cartilaginous tissues. We propose that, in a context of a reduced rate of evolution, sturgeon OC has retained structural features of the ancestral protein that emerged millions of years ago from the duplication of an ancient MGP gene and may exhibit intermediate functional features. PMID:23884418

  15. Novel protein-protein interaction family proteins involved in chloroplast movement response.

    PubMed

    Kodama, Yutaka; Suetsugu, Noriyuki; Wada, Masamitsu

    2011-04-01

    To optimize photosynthetic activity, chloroplasts change their intracellular location in response to ambient light conditions; chloroplasts move toward low intensity light to maximize light capture, and away from high intensity light to avoid photodamage. Although several proteins have been reported to be involved in the chloroplast photorelocation movement response, any physical interaction among them was not found so far. We recently found a physical interaction between two plant-specific coiled-coil proteins, WEB1 (Weak Chloroplast Movement under Blue Light 1) and PMI2 (Plastid Movement Impaired 2), that were identified to regulate chloroplast movement velocity. Since the both coiled-coil regions of WEB1 and PMI2 were classified into an uncharacterized protein family having DUF827 (DUF: Domain of Unknown Function) domain, it was the first report that DUF827 proteins could mediate protein-protein interaction. In this mini-review article, we discuss regarding molecular function of WEB1 and PMI2, and also define a novel protein family composed of WEB1, PMI2 and WEB1/PMI2-like proteins for protein-protein interaction in land plants.

  16. Involvement of PCH family proteins in cytokinesis and actin distribution.

    PubMed

    Lippincott, J; Li, R

    2000-04-15

    Pombe Cdc15 homology (PCH) proteins constitute an extensive protein family whose members have been found in diverse eukaryotic organisms. These proteins are characterized by the presence of several conserved sequence and structural motifs. Recent studies in yeast and mammalian cultured cells have implicated these proteins in actin-based processes, in particular, cytokinesis. Here we review the recent findings on the in vivo localization, function, and binding partners of PCH family members. We also provide new microscopy data regarding the in vivo dynamics of a budding yeast PCH protein involved in cytokinesis.

  17. Methods for Mapping of Interaction Networks Involving Membrane Proteins

    SciTech Connect

    Hooker, Brian S.; Bigelow, Diana J.; Lin, Chiann Tso

    2007-11-23

    Numerous approaches have been taken to study protein interactions, such as tagged protein complex isolation followed by mass spectrometry, yeast two-hybrid methods, fluorescence resonance energy transfer, surface plasmon resonance, site-directed mutagenesis, and crystallography. Membrane protein interactions pose significant challenges due to the need to solubilize membranes without disrupting protein-protein interactions. Traditionally, analysis of isolated protein complexes by high-resolution 2D gel electrophoresis has been the main method used to obtain an overall picture of proteome constituents and interactions. However, this method is time consuming, labor intensive, detects only abundant proteins and is not suitable for the coverage required to elucidate large interaction networks. In this review, we discuss the application of various methods to elucidate interactions involving membrane proteins. These techniques include methods for the direct isolation of single complexes or interactors as well as methods for characterization of entire subcellular and cellular interactomes.

  18. A Prediction Model for Membrane Proteins Using Moments Based Features

    PubMed Central

    Butt, Ahmad Hassan; Khan, Sher Afzal; Jamil, Hamza; Rasool, Nouman; Khan, Yaser Daanial

    2016-01-01

    The most expedient unit of the human body is its cell. Encapsulated within the cell are many infinitesimal entities and molecules which are protected by a cell membrane. The proteins that are associated with this lipid based bilayer cell membrane are known as membrane proteins and are considered to play a significant role. These membrane proteins exhibit their effect in cellular activities inside and outside of the cell. According to the scientists in pharmaceutical organizations, these membrane proteins perform key task in drug interactions. In this study, a technique is presented that is based on various computationally intelligent methods used for the prediction of membrane protein without the experimental use of mass spectrometry. Statistical moments were used to extract features and furthermore a Multilayer Neural Network was trained using backpropagation for the prediction of membrane proteins. Results show that the proposed technique performs better than existing methodologies. PMID:26966690

  19. Molecular Simulation Studies of Proteins Involved in Parkinson's Disease

    NASA Astrophysics Data System (ADS)

    Carloni, Paolo

    2007-12-01

    This contribution describes two recent computational studies related to proteins involved in Parkinson's Disease (PD). The first focuses on the interplay between dopamine and α-synuclein (AS), which plays a central role in PD (unpublished results). The second deals with the protein DJ-1, whose mutations are present in patients suffering from familiar PD [1]. Computational methods are used to investigate the relationship between such mutations and the protein oligomeric state, which may be important for the progression of the disease.

  20. Key proteins involved in insulin vesicle exocytosis and secretion

    PubMed Central

    Xiong, Qian-Yin; Yu, Cui; Zhang, Yao; Ling, Liefeng; Wang, Lizhuo; Gao, Jia-Lin

    2017-01-01

    In vivo insulin secretion is predominantly affected by blood glucose concentration, blood concentration of amino acids, gastrointestinal hormones and free nerve functional status, in addition to other factors. Insulin is one of the most important hormones in the body, and its secretion is precisely controlled by nutrients, neurotransmitters and hormones. The insulin exocytosis process is similar to the neurotransmitter release mechanism. There are various types of proteins and lipids that participate in the insulin secretory vesicle fusion process, such as soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) protein, Ras-related proteins and vacuolar-type H+-ATPase (V-ATPase). Notably, the SNARE protein is the molecular basis of exocytotic activity. In the current review, the role of the vesicle membrane proteins (synaptobrevins, vesicle associated membrane proteins and target membrane proteins) and auxiliary proteins (Rab proteins and Munc-18 proteins) in vesicle fusion activity were summarized. A summary of these key proteins involved in insulin granule secretion will facilitate understanding of the pathogenesis of diabetes. PMID:28357064

  1. Extracting features from protein sequences to improve deep extreme learning machine for protein fold recognition.

    PubMed

    Ibrahim, Wisam; Abadeh, Mohammad Saniee

    2017-03-27

    Protein fold recognition is an important problem in bioinformatics to predict three-dimensional structure of a protein. One of the most challenging tasks in protein fold recognition problem is the extraction of efficient features from the amino-acid sequences to obtain better classifiers. In this paper, we have proposed six descriptors to extract features from protein sequences. These descriptors are applied in the first stage of a three-stage framework PCA-DELM-LDA to extract feature vectors from the amino-acid sequences. Principal Component Analysis PCA has been implemented to reduce the number of extracted features. The extracted feature vectors have been used with original features to improve the performance of the Deep Extreme Learning Machine DELM in the second stage. Four new features have been extracted from the second stage and used in the third stage by Linear Discriminant Analysis LDA to classify the instances into 27 folds. The proposed framework is implemented on the independent and combined feature sets in SCOP datasets. The experimental results show that extracted feature vectors in the first stage could improve the performance of DELM in extracting new useful features in second stage.

  2. Borderline personality disorder features and history of childhood maltreatment in mothers involved with child protective services.

    PubMed

    Perepletchikova, Francheska; Ansell, Emily; Axelrod, Seth

    2012-05-01

    This study examines the history of childhood maltreatment and Borderline Personality Disorder (BPD) symptoms in mothers whose children were removed from the home by Child Protective Services (CPS) to identify potential targets for future intervention efforts. Forty-one mothers of children removed from the home due to abuse and/or neglect and 58 community-control mothers without CPS involvement were assessed for history of childhood maltreatment, alcohol and drug use, and BPD features. CPS-involved mothers scored significantly higher on measures of childhood maltreatment history and BPD features than did control mothers. The highest BPD scores were associated with the most severe histories of mothers' childhood maltreatment. In total, 50% of CPS-involved mothers reported elevated BPD features, compared with 15% of control mothers. Further, 19% of CPS-involved mothers had self-reported scores consistent with a BPD diagnosis, compared with 4% of control mothers. BPD features rather than maltreatment history per se predicted maternal involvement with CPS, controlling for alcohol and drug use predictors. The present data suggest that evidence-based treatments to address BPD symptoms may be indicated for some CPS-involved parents.

  3. Weighted feature value based Drug Target Protein prediction.

    PubMed

    Hyun, Bo-ra; Jung, Hwiesung; Jang, Woo-Hyuk; Jung, Suk Hoon; Han, Dong-Soo

    2008-01-01

    Drug discovery is a long process in which only a few successful new therapeutic discoveries are made and identification of drug target candidate proteins requires considerable time and efforts. However, the accumulation of information on drugs has made it possible to devise new computational methods for classifying drug target candidates. In this paper, we devise a Drug Target Protein (DT-P) classification method by the summation of weighted features which is extracted from known DT-P. The method is validated using Bayesian decision theory and SVM, and it was revealed to achieve high specificity of 89.5% with 88% accuracy.

  4. SCRATCH: a protein structure and structural feature prediction server

    PubMed Central

    Cheng, J.; Randall, A. Z.; Sweredoski, M. J.; Baldi, P.

    2005-01-01

    SCRATCH is a server for predicting protein tertiary structure and structural features. The SCRATCH software suite includes predictors for secondary structure, relative solvent accessibility, disordered regions, domains, disulfide bridges, single mutation stability, residue contacts versus average, individual residue contacts and tertiary structure. The user simply provides an amino acid sequence and selects the desired predictions, then submits to the server. Results are emailed to the user. The server is available at . PMID:15980571

  5. Proteins involved in vesicular transport and membrane fusion.

    PubMed

    Waters, M G; Griff, I C; Rothman, J E

    1991-08-01

    In the past year, new information about proteins involved in vesicular transport has been plentiful. Particularly noteworthy are the complementary findings that Sec17p is required for vesicle consumption in endoplasmic reticulum-to-Golgi transport in yeast and that an analogous activity in mammalian cells, termed SNAP, is required for transport from the cis to the medial cisternae of the Golgi apparatus.

  6. Proteomic analysis of proteins involved in spermiogenesis in mouse.

    PubMed

    Guo, Xuejiang; Shen, Jian; Xia, Zhengrong; Zhang, Rui; Zhang, Ping; Zhao, Chun; Xing, Jun; Chen, Ling; Chen, Wen; Lin, Min; Huo, Ran; Su, Bing; Zhou, Zuomin; Sha, Jiahao

    2010-03-05

    Spermiogenesis is a unique process in mammals during which haploid round spermatids mature into spermatozoa in the testis. Its successful completion is necessary for fertilization and its malfunction is an important cause of male infertility. Here, we report the high-confidence identification of 2116 proteins in mouse haploid germ cells undergoing spermiogenesis: 299 of these were testis-specific and 155 were novel. Analysis of these proteins showed many proteins possibly functioning in unique processes of spermiogenesis. Of the 84 proteins annotated to be involved in vesicle-related events, VAMP4 was shown to be important for acrosome biogenesis by in vivo knockdown experiments. Knockdown of VAMP4 caused defects of acrosomal vesicle fusion and significantly increased head abnormalities in spermatids from testis and sperm from the cauda epididymis. Analysis of chromosomal distribution of the haploid genes showed underrepresentation on the X chromosome and overrepresentation on chromosome 11, which were due to meiotic sex chromosome inactivation and expansion of testis-expressed gene families, respectively. Comparison with transcriptional data showed translational regulation during spermiogenesis. This characterization of proteins involved in spermiogenesis provides an inventory of proteins useful for understanding the mechanisms of male infertility and may provide candidates for drug targets for male contraception and male infertility.

  7. Analysis of membrane proteins in metagenomics: networks of correlated environmental features and protein families.

    PubMed

    Patel, Prianka V; Gianoulis, Tara A; Bjornson, Robert D; Yip, Kevin Y; Engelman, Donald M; Gerstein, Mark B

    2010-07-01

    Recent metagenomics studies have begun to sample the genomic diversity among disparate habitats and relate this variation to features of the environment. Membrane proteins are an intuitive, but thus far overlooked, choice in this type of analysis as they directly interact with the environment, receiving signals from the outside and transporting nutrients. Using global ocean sampling (GOS) data, we found nearly approximately 900,000 membrane proteins in large-scale metagenomic sequence, approximately a fifth of which are completely novel, suggesting a large space of hitherto unexplored protein diversity. Using GPS coordinates for the GOS sites, we extracted additional environmental features via interpolation from the World Ocean Database, the National Center for Ecological Analysis and Synthesis, and empirical models of dust occurrence. This allowed us to study membrane protein variation in terms of natural features, such as phosphate and nitrate concentrations, and also in terms of human impacts, such as pollution and climate change. We show that there is widespread variation in membrane protein content across marine sites, which is correlated with changes in both oceanographic variables and human factors. Furthermore, using these data, we developed an approach, protein families and environment features network (PEN), to quantify and visualize the correlations. PEN identifies small groups of covarying environmental features and membrane protein families, which we call "bimodules." Using this approach, we find that the affinity of phosphate transporters is related to the concentration of phosphate and that the occurrence of iron transporters is connected to the amount of shipping, pollution, and iron-containing dust.

  8. Human neurocysticercosis: immunological features involved in the host's susceptibility to become infected and to develop disease.

    PubMed

    Sciutto, Edda; Cárdenas, Graciela; Adalid-Peralta, Laura; Fragoso, Gladis; Larralde, Carlos; Fleury, Agnes

    2013-06-01

    Human neurocysticercosis (NC) is a clinically and radiologically heterogeneous disease caused by the establishment of Taenia solium larvae in the central nervous system. Herein, the immunological and endocrinological features involved in resistance to infection and severe forms of the disease are reviewed, and their clinical relevance is discussed.

  9. Efficient Feature Selection and Classification of Protein Sequence Data in Bioinformatics

    PubMed Central

    Faye, Ibrahima; Samir, Brahim Belhaouari; Md Said, Abas

    2014-01-01

    Bioinformatics has been an emerging area of research for the last three decades. The ultimate aims of bioinformatics were to store and manage the biological data, and develop and analyze computational tools to enhance their understanding. The size of data accumulated under various sequencing projects is increasing exponentially, which presents difficulties for the experimental methods. To reduce the gap between newly sequenced protein and proteins with known functions, many computational techniques involving classification and clustering algorithms were proposed in the past. The classification of protein sequences into existing superfamilies is helpful in predicting the structure and function of large amount of newly discovered proteins. The existing classification results are unsatisfactory due to a huge size of features obtained through various feature encoding methods. In this work, a statistical metric-based feature selection technique has been proposed in order to reduce the size of the extracted feature vector. The proposed method of protein classification shows significant improvement in terms of performance measure metrics: accuracy, sensitivity, specificity, recall, F-measure, and so forth. PMID:25045727

  10. NEDD8 protein is involved in ubiquitinated inclusion bodies.

    PubMed

    Dil Kuazi, Afroz; Kito, Katsumi; Abe, Yasuhito; Shin, Ryong-Woon; Kamitani, Tetsu; Ueda, Norifumi

    2003-02-01

    Proteolysis by the ubiquitin-proteasome system is considered to play a pathological role in several degenerative diseases that involve ubiquitinated inclusion bodies. In recent years, several ubiquitin-like proteins have been isolated, but it is uncertain whether their roles are associated with protein degradation through the ubiquitin-proteasome system. NEDD8 (neural precursor cell-expressed and developmentally down-regulated gene), which consists of 81 amino acid residues, possesses the highest sequence similarity to ubiquitin. Recent studies have indicated that NEDD8 is covalently ligated to cullin family proteins, which are components of certain ubiquitin E3 ligases, by a pathway analogous to that of ubiquitin. Thus, by focusing on the structural and functional association between NEDD8 and ubiquitin, it would be of interest to know whether the NEDD8 system is involved in pathological disorders of the ubiquitin-proteasome system. This study has examined the immunohistochemical distribution of NEDD8 protein by using a highly purified antibody in normal tissues and in tissues known to contain ubiquitinated inclusions. NEDD8 protein expression was widely observed in most types of tissues. Furthermore, accumulation of the NEDD8 protein was commonly observed in ubiquitinated inclusion bodies, including Lewy bodies in Parkinson's disease, Mallory bodies in alcoholic liver disease, and Rosenthal fibres in astrocytoma. Two of ten cases of neurofibrillary tangles and senile plaques from patients with Alzheimer's disease showed intense staining for NEDD8 as well as for ubiquitin. These findings suggest the possibility that the NEDD8 system is involved in the metabolism of these inclusion bodies via the ubiquitin-proteasome system.

  11. Identifying Unstable Regions of Proteins Involved in Misfolding Diseases

    NASA Astrophysics Data System (ADS)

    Guest, Will; Cashman, Neil; Plotkin, Steven

    2009-05-01

    Protein misfolding is a necessary step in the pathogenesis of many diseases, including Creutzfeldt-Jakob disease (CJD) and familial amyotrophic lateral sclerosis (fALS). Identifying unstable structural elements in their causative proteins elucidates the early events of misfolding and presents targets for inhibition of the disease process. An algorithm was developed to calculate the Gibbs free energy of unfolding for all sequence-contiguous regions of a protein using three methods to parameterize energy changes: a modified G=o model, changes in solvent-accessible surface area, and all-atoms molecular dynamics. The entropic effects of disulfide bonds and post-translational modifications are treated analytically. It incorporates a novel method for finding local dielectric constants inside a protein to accurately handle charge effects. We have predicted the unstable parts of prion protein and superoxide dismutase 1, the proteins involved in CJD and fALS respectively, and have used these regions as epitopes to prepare antibodies that are specific to the misfolded conformation and show promise as therapeutic agents.

  12. First identification of proteins involved in motility of Mycoplasma gallisepticum.

    PubMed

    Indikova, Ivana; Vronka, Martin; Szostak, Michael P

    2014-10-17

    Mycoplasma gallisepticum, the most pathogenic mycoplasma in poultry, is able to glide over solid surfaces. Although this gliding motility was first observed in 1968, no specific protein has yet been shown to be involved in gliding. We examined M. gallisepticum strains and clonal variants for motility and found that the cytadherence proteins GapA and CrmA were required for gliding. Loss of GapA or CrmA resulted in the loss of motility and hemadsorption and led to drastic changes in the characteristic flask-shape of the cells. To identify further genes involved in motility, a transposon mutant library of M. gallisepticum was generated and screened for motility-deficient mutants, using a screening assay based on colony morphology. Motility-deficient mutants had transposon insertions in gapA and the neighbouring downstream gene crmA. In addition, insertions were seen in gene mgc2, immediately upstream of gapA, in two motility-deficient mutants. In contrast to the GapA/CrmA mutants, the mgc2 motility mutants still possessed the ability to hemadsorb. Complementation of these mutants with a mgc2-hexahistidine fusion gene restored the motile phenotype. This is the first report assigning specific M. gallisepticum proteins to involvement in gliding motility.

  13. Proteomic detection of proteins involved in perchlorate and chlorate metabolism.

    PubMed

    Bansal, Reema; Deobald, Lee A; Crawford, Ronald L; Paszczynski, Andrzej J

    2009-09-01

    Mass spectrometry and a time-course cell lysis method were used to study proteins involved in perchlorate and chlorate metabolism in pure bacterial cultures and environmental samples. The bacterial cultures used included Dechlorosoma sp. KJ, Dechloromonas hortensis, Pseudomonas chloritidismutans ASK-1, and Pseudomonas stutzeri. The environmental samples included an anaerobic sludge enrichment culture from a sewage treatment plant, a sample of a biomass-covered activated carbon matrix from a bioreactor used for treating perchlorate-contaminated drinking water, and a waste water effluent sample from a paper mill. The approach focused on detection of perchlorate (and chlorate) reductase and chlorite dismutase proteins, which are the two central enzymes in the perchlorate (or chlorate) reduction pathways. In addition, acetate-metabolizing enzymes in pure bacterial samples and housekeeping proteins from perchlorate (or chlorate)-reducing microorganisms in environmental samples were also identified.

  14. Molecular signaling involving intrinsically disordered proteins in prostate cancer

    PubMed Central

    Russo, Anna; Manna, Sara La; Novellino, Ettore; Malfitano, Anna Maria; Marasco, Daniela

    2016-01-01

    Investigations on cellular protein interaction networks (PINs) reveal that proteins that constitute hubs in a PIN are notably enriched in Intrinsically Disordered Proteins (IDPs) compared to proteins that constitute edges, highlighting the role of IDPs in signaling pathways. Most IDPs rapidly undergo disorder-to-order transitions upon binding to their biological targets to perform their function. Conformational dynamics enables IDPs to be versatile and to interact with a broad range of interactors under normal physiological conditions where their expression is tightly modulated. IDPs are involved in many cellular processes such as cellular signaling, transcriptional regulation, and splicing; thus, their high-specificity/low-affinity interactions play crucial roles in many human diseases including cancer. Prostate cancer (PCa) is one of the leading causes of cancer-related mortality in men worldwide. Therefore, identifying molecular mechanisms of the oncogenic signaling pathways that are involved in prostate carcinogenesis is crucial. In this review, we focus on the aspects of cellular pathways leading to PCa in which IDPs exert a primary role. PMID:27212129

  15. Bap, a Staphylococcus aureus Surface Protein Involved in Biofilm Formation

    PubMed Central

    Cucarella, Carme; Solano, Cristina; Valle, Jaione; Amorena, Beatriz; Lasa, Íñigo; Penadés, José R.

    2001-01-01

    Identification of new genes involved in biofilm formation is needed to understand the molecular basis of strain variation and the pathogenic mechanisms implicated in chronic staphylococcal infections. A biofilm-producing Staphylococcus aureus isolate was used to generate biofilm-negative transposon (Tn917) insertion mutants. Two mutants were found with a significant decrease in attachment to inert surfaces (early adherence), intercellular adhesion, and biofilm formation. The transposon was inserted at the same locus in both mutants. This locus (bap [for biofilm associated protein]) encodes a novel cell wall associated protein of 2,276 amino acids (Bap), which shows global organizational similarities to surface proteins of gram-negative (Pseudomonas aeruginosa and Salmonella enterica serovar Typhi) and gram-positive (Enteroccocus faecalis) microorganisms. Bap's core region represents 52% of the protein and consists of 13 successive nearly identical repeats, each containing 86 amino acids. bap was present in a small fraction of bovine mastitis isolates (5% of the 350 S. aureus isolates tested), but it was absent from the 75 clinical human S. aureus isolates analyzed. All staphylococcal isolates harboring bap were highly adherent and strong biofilm producers. In a mouse infection model bap was involved in pathogenesis, causing a persistent infection. PMID:11292810

  16. [Proteins of human milk involved in immunological processes].

    PubMed

    Lis, Jolanta; Orczyk-Pawiłowicz, Magdalena; Kątnik-Prastowska, Iwona

    2013-05-31

    Human milk contains a lot of components (i.e. proteins, carbohydrates, lipids, inorganic elements) which provide basic nutrients for infants during the first period of their lives. Qualitative composition of milk components of healthy mothers is similar, but their levels change during lactation stages. Colostrum is the fluid secreted during the first days postpartum by mammary epithelial cells. Colostrum is replaced by transitional milk during 5-15 days postpartum and from 15 days postpartum mature milk is produced. Human milk, apart from nutritional components, is a source of biologically active molecules, i.e. immunoglobulins, growth factors, cytokines, acute phase proteins, antiviral and antibacterial proteins. Such components of human milk are responsible for specific biological activities of human milk. This secretion plays an important role in growth and development of newborns. Bioactive molecules present in the milk support the immature immune system of the newborn and also protect against the development of infection. In this article we describe the pathways involved in the production and secretion of human milk, the state of knowledge on the proteome of human milk, and the contents of components of milk during lactation. Moreover, some growth factors and proteins involved in innate and specific immunity, intercellular communication, immunomodulation, and inflammatory processes have been characterized.

  17. Protein Function Prediction using Text-based Features extracted from the Biomedical Literature: The CAFA Challenge

    PubMed Central

    2013-01-01

    % respectively. Results obtained as part of the CAFA evaluation itself on the CAFA dataset are reported as well. Conclusions Our evaluation shows that the text-based classifier consistently outperforms the baseline classifier that is based on prior distribution, and typically has comparable performance to the baseline classifier that uses sequence similarity. Moreover, the results suggest that combining text features with other types of features can potentially lead to improved prediction performance. The preliminary results also suggest that while our text-based classifier can be used to predict both molecular function and biological process in which a protein is involved, the classifier performs significantly better for predicting molecular function than for predicting biological process. A similar trend was observed for other classifiers participating in the CAFA challenge. PMID:23514326

  18. Analysis of proteins involved in biodegradation of crop biomass

    NASA Technical Reports Server (NTRS)

    Crawford, Kamau; Trotman, Audrey

    1998-01-01

    The biodegradation of crop biomass for re-use in crop production is part of the bioregenerative life support concept proposed by the National Aeronautics and Space Administration (NASA) for long duration, manned space exploration. The current research was conducted in the laboratory to evaluate the use of electrophoretic analysis as a means of rapidly assaying for constitutive and induced proteins associated with the bacterial degradation of crop residue. The proteins involved in crop biomass biodegradation are either constitutive or induced. As a result, effluent and cultures were examined to investigate the potential of using electrophoretic techniques as a means of monitoring the biodegradation process. Protein concentration for optimum banding patterns was determined using the Bio-Rad Protein Assay kit. Four bacterial soil isolates were obtained from the G.W. Carver research Farm at Tuskegee University and used in the decomposition of components of plant biomass. The culture, WDSt3A was inoculated into 500 mL of either Tryptic Soy Broth or Nutrient Broth. Incubation, with shaking of each flask was for 96 hours at 30 C. The cultures consistently gave unique banding patterns under denaturing protein electrophoresis conditions, The associated extracellular enzymes also yielded characteristic banding patterns over a 14-day period, when native electrophoresis techniques were used to examine effluent from batch culture bioreactors. The current study evaluated sample preparation and staining protocols to determine the ease of use, reproducibility and reliability, as well as the potential for automation.

  19. Hepatic involvement in HELLP syndrome: an update with emphasis on imaging features.

    PubMed

    Perronne, Laetitia; Dohan, Anthony; Bazeries, Paul; Guerrache, Youcef; Fohlen, Audrey; Rousset, Pascal; Aubé, Christophe; Laurent, Valérie; Morel, Olivier; Boudiaf, Mourad; Hoeffel, Christine; Soyer, Philippe

    2015-10-01

    HELLP syndrome, which consists of hemolysis, elevated liver enzymes, and low platelet count is an unusual complication of pregnancy that is observed in only 10% to 15% of women with preeclampsia. Hepatic involvement in HELLP syndrome may present with various imaging features depending on the specific condition that includes nonspecific abnormalities such as perihepatic free fluid, hepatic steatosis, liver enlargement, and periportal halo that may precede more severe conditions such as hepatic hematoma and hepatic rupture with hemoperitoneum. Maternal clinical symptoms may be nonspecific and easily mistaken for a variety of other conditions that should be recognized. Because hepatic hematoma occurring in association with preeclampsia and HELLP syndrome is a potentially life-threatening complication, prompt depiction is critical and may help reduce morbidity and mortality. This review provides an update on demographics, risk factors, pathophysiology, and clinical features of hepatic complications due to HELLP syndrome along with a special emphasis on the imaging features of these uncommon conditions.

  20. Prediction of microRNAs involved in immune system diseases through network based features.

    PubMed

    Prabahar, Archana; Natarajan, Jeyakumar

    2017-01-01

    MicroRNAs are a class of small non-coding regulatory RNA molecules that modulate the expression of several genes at post-transcriptional level and play a vital role in disease pathogenesis. Recent research shows that a range of miRNAs are involved in the regulation of immunity and its deregulation results in immune mediated diseases such as cancer, inflammation and autoimmune diseases. Computational discovery of these immune miRNAs using a set of specific features is highly desirable. In the current investigation, we present a SVM based classification system which uses a set of novel network based topological and motif features in addition to the baseline sequential and structural features to predict immune specific miRNAs from other non-immune miRNAs. The classifier was trained and tested on a balanced set of equal number of positive and negative examples to show the discriminative power of our network features. Experimental results show that our approach achieves an accuracy of 90.2% and outperforms the classification accuracy of 63.2% reported using the traditional miRNA sequential and structural features. The proposed classifier was further validated with two immune disease sub-class datasets related to multiple sclerosis microarray data and psoriasis RNA-seq data with higher accuracy. These results indicate that our classifier which uses network and motif features along with sequential and structural features will lead to significant improvement in classifying immune miRNAs and hence can be applied to identify other specific classes of miRNAs as an extensible miRNA classification system.

  1. Involvement of heat shock proteins in gluten-sensitive enteropathy.

    PubMed

    Sziksz, Erna; Pap, Domonkos; Veres, Gábor; Fekete, Andrea; Tulassay, Tivadar; Vannay, Ádám

    2014-06-07

    Gluten-sensitive enteropathy, also known as coeliac disease (CD), is an autoimmune disorder occurring in genetically susceptible individuals that damages the small intestine and interferes with the absorption of other nutrients. As it is triggered by dietary gluten and related prolamins present in wheat, rye and barley, the accepted treatment for CD is a strict gluten-free diet. However, a complete exclusion of gluten-containing cereals from the diet is often difficult, and new therapeutic strategies are urgently needed. A class of proteins that have already emerged as drug targets for other autoimmune diseases are the heat shock proteins (HSPs), which are highly conserved stress-induced chaperones that protect cells against harmful extracellular factors. HSPs are expressed in several tissues, including the gastrointestinal tract, and their levels are significantly increased under stress circumstances. HSPs exert immunomodulatory effects, and also play a crucial role in the maintenance of epithelial cell structure and function, as they are responsible for adequate protein folding, influence the degradation of proteins and cell repair processes after damage, and modulate cell signalling, cell proliferation and apoptosis. The present review discusses the involvement of HSPs in the pathophysiology of CD. Furthermore, HSPs may represent a useful therapeutic target for the treatment of CD due to the cytoprotective, immunomodulatory, and anti-apoptotic effects in the intestinal mucosal barrier.

  2. Identification of an additional protein involved in mannan biosynthesis

    PubMed Central

    Wang, Yan; Mortimer, Jennifer C; Davis, Jonathan; Dupree, Paul; Keegstra, Kenneth

    2013-01-01

    Galactomannans comprise a β-1,4-mannan backbone substituted with α-1,6-galactosyl residues. Genes encoding the enzymes that are primarily responsible for backbone synthesis and side-chain addition of galactomannans were previously identified and characterized. To identify additional genes involved in galactomannan biosynthesis, we previously performed deep EST profiling of fenugreek (Trigonella foenum-graecum L.) seed endosperm, which accumulates large quantities of galactomannans as a reserve carbohydrate during seed development. One of the candidate genes encodes a protein that is likely to be a glycosyltransferase. Because this protein is involved in mannan biosynthesis, we named it ‘mannan synthesis-related’ (MSR). Here, we report the characterization of a fenugreek MSR gene (TfMSR) and its two Arabidopsis homologs, AtMSR1 and AtMSR2. TfMSR was highly and specifically expressed in the endosperm. TfMSR, AtMSR1 and AtMSR2 proteins were all determined to be localized to the Golgi by fluorescence confocal microscopy. The level of mannosyl residues in stem glucomannans decreased by approximately 40% for Arabidopsis msr1 single T-DNA insertion mutants and by more than 50% for msr1 msr2 double mutants, but remained unchanged for msr2 single mutants. In addition, in vitro mannan synthase activity from the stems of msr1 single and msr1 msr2 double mutants also decreased. Expression of AtMSR1 or AtMSR2 in the msr1 msr2 double mutant completely or partially restored mannosyl levels. From these results, we conclude that the MSR protein is important for mannan biosynthesis, and offer some ideas about its role. PMID:22966747

  3. Genes and proteins involved in bacterial magnetic particle formation.

    PubMed

    Matsunaga, Tadashi; Okamura, Yoshiko

    2003-11-01

    Magnetic bacteria synthesize intracellular magnetosomes that impart a cellular swimming behaviour referred to as magnetotaxis. The magnetic structures aligned in chains are postulated to function as biological compass needles allowing the bacterium to migrate along redox gradients through the Earth's geomagnetic field lines. Despite the discovery of this unique group of microorganisms 28 years ago, the mechanisms of magnetic crystal biomineralization have yet to be fully elucidated. This review describes the current knowledge of the genes and proteins involved in magnetite formation in magnetic bacteria and the biotechnological applications of biomagnetites in the interdisciplinary fields of nanobiotechnology, medicine and environmental management.

  4. Critical Features of Fragment Libraries for Protein Structure Prediction.

    PubMed

    Trevizani, Raphael; Custódio, Fábio Lima; Dos Santos, Karina Baptista; Dardenne, Laurent Emmanuel

    2017-01-01

    The use of fragment libraries is a popular approach among protein structure prediction methods and has proven to substantially improve the quality of predicted structures. However, some vital aspects of a fragment library that influence the accuracy of modeling a native structure remain to be determined. This study investigates some of these features. Particularly, we analyze the effect of using secondary structure prediction guiding fragments selection, different fragments sizes and the effect of structural clustering of fragments within libraries. To have a clearer view of how these factors affect protein structure prediction, we isolated the process of model building by fragment assembly from some common limitations associated with prediction methods, e.g., imprecise energy functions and optimization algorithms, by employing an exact structure-based objective function under a greedy algorithm. Our results indicate that shorter fragments reproduce the native structure more accurately than the longer. Libraries composed of multiple fragment lengths generate even better structures, where longer fragments show to be more useful at the beginning of the simulations. The use of many different fragment sizes shows little improvement when compared to predictions carried out with libraries that comprise only three different fragment sizes. Models obtained from libraries built using only sequence similarity are, on average, better than those built with a secondary structure prediction bias. However, we found that the use of secondary structure prediction allows greater reduction of the search space, which is invaluable for prediction methods. The results of this study can be critical guidelines for the use of fragment libraries in protein structure prediction.

  5. Critical Features of Fragment Libraries for Protein Structure Prediction

    PubMed Central

    dos Santos, Karina Baptista

    2017-01-01

    The use of fragment libraries is a popular approach among protein structure prediction methods and has proven to substantially improve the quality of predicted structures. However, some vital aspects of a fragment library that influence the accuracy of modeling a native structure remain to be determined. This study investigates some of these features. Particularly, we analyze the effect of using secondary structure prediction guiding fragments selection, different fragments sizes and the effect of structural clustering of fragments within libraries. To have a clearer view of how these factors affect protein structure prediction, we isolated the process of model building by fragment assembly from some common limitations associated with prediction methods, e.g., imprecise energy functions and optimization algorithms, by employing an exact structure-based objective function under a greedy algorithm. Our results indicate that shorter fragments reproduce the native structure more accurately than the longer. Libraries composed of multiple fragment lengths generate even better structures, where longer fragments show to be more useful at the beginning of the simulations. The use of many different fragment sizes shows little improvement when compared to predictions carried out with libraries that comprise only three different fragment sizes. Models obtained from libraries built using only sequence similarity are, on average, better than those built with a secondary structure prediction bias. However, we found that the use of secondary structure prediction allows greater reduction of the search space, which is invaluable for prediction methods. The results of this study can be critical guidelines for the use of fragment libraries in protein structure prediction. PMID:28085928

  6. Life under tension: Computational studies of proteins involved in mechanotransduction

    NASA Astrophysics Data System (ADS)

    Sotomayor, Marcos Manuel

    cadherins. Simulations also revealed how calcium ions control cadherin's shape and the availability of key residues involved in cell-cell adhesion, suggesting a conceptual framework for interpreting mutations in cadherin calcium binding motifs causing hereditary deafness. Overall, simulations provided a unique nanoscopic view of the dynamics and function of some of the proteins involved in mechanotransduction.

  7. Possible involvement of poly(A) in protein synthesis.

    PubMed Central

    Jacobson, A; Favreau, M

    1983-01-01

    The experiments of this paper have re-evaluated the possibility that poly(A) is involved in protein synthesis by testing whether purified poly(A) might competitively inhibit in vitro protein synthesis in rabbit reticulocyte extracts. We have found that poly(A) inhibits the rate of translation of many different poly(A)+ mRNAs and that comparable inhibition is not observed with other ribopolymers. Inhibition by poly(A) preferentially affects the translation of adenylated mRNAs and can be overcome by increased mRNA concentrations or by translating mRNPs instead of mRNA. The extent of inhibition is dependent on the size of the competitor poly(A) as well as on the translation activity which a lysate has for poly(A)+ RNA. In light of our results and numerous experiments in the literature, we propose that poly(A) has a function in protein synthesis and that any role in the determination of mRNA stability is indirect. Images PMID:6137807

  8. Arabinogalactan proteins are involved in root hair development in barley

    PubMed Central

    Marzec, Marek; Szarejko, Iwona; Melzer, Michael

    2015-01-01

    The arabinogalactan proteins (AGPs) are involved in a range of plant processes, including cell differentiation and expansion. Here, barley root hair mutants and their wild-type parent cultivars were used, as a model system, to reveal the role of AGPs in root hair development. The treatment of roots with different concentrations of βGlcY (a reagent which binds to all classes of AGPs) inhibited or totally suppressed the development of root hairs in all of the cultivars. Three groups of AGP (recognized by the monoclonal antibodies LM2, LM14, and MAC207) were diversely localized in trichoblasts and atrichoblasts of root hair-producing plants. The relevant epitopes were present in wild-type trichoblast cell walls and cytoplasm, whereas in wild-type atrichoblasts and in all epidermal cells of a root hairless mutant, they were only present in the cytoplasm. In all of cultivars the higher expression of LM2, LM14, and MAC207 was observed in trichoblasts at an early stage of development. Additionally, the LM2 epitope was detected on the surface of primordia and root hair tubes in plants able to generate root hairs. The major conclusion was that the AGPs recognized by LM2, LM14, and MAC207 are involved in the differentiation of barley root epidermal cells, thereby implying a requirement for these AGPs for root hair development in barley. PMID:25465033

  9. Acute myeloid leukemia fusion proteins deregulate genes involved in stem cell maintenance and DNA repair

    PubMed Central

    Alcalay, Myriam; Meani, Natalia; Gelmetti, Vania; Fantozzi, Anna; Fagioli, Marta; Orleth, Annette; Riganelli, Daniela; Sebastiani, Carla; Cappelli, Enrico; Casciari, Cristina; Sciurpi, Maria Teresa; Mariano, Angela Rosa; Minardi, Simone Paolo; Luzi, Lucilla; Muller, Heiko; Di Fiore, Pier Paolo; Frosina, Guido; Pelicci, Pier Giuseppe

    2003-01-01

    Acute myelogenous leukemias (AMLs) are genetically heterogeneous and characterized by chromosomal rearrangements that produce fusion proteins with aberrant transcriptional regulatory activities. Expression of AML fusion proteins in transgenic mice increases the risk of myeloid leukemias, suggesting that they induce a preleukemic state. The underlying molecular and biological mechanisms are, however, unknown. To address this issue, we performed a systematic analysis of fusion protein transcriptional targets. We expressed AML1/ETO, PML/RAR, and PLZF/RAR in U937 hemopoietic precursor cells and measured global gene expression using oligonucleotide chips. We identified 1,555 genes regulated concordantly by at least two fusion proteins that were further validated in patient samples and finally classified according to available functional information. Strikingly, we found that AML fusion proteins induce genes involved in the maintenance of the stem cell phenotype and repress DNA repair genes, mainly of the base excision repair pathway. Functional studies confirmed that ectopic expression of fusion proteins constitutively activates pathways leading to increased stem cell renewal (e.g., the Jagged1/Notch pathway) and provokes accumulation of DNA damage. We propose that expansion of the stem cell compartment and induction of a mutator phenotype are relevant features underlying the leukemic potential of AML-associated fusion proteins. PMID:14660751

  10. Magnifying Endoscopic Features of Follicular Lymphoma Involving the Stomach: A Report of Two Cases

    PubMed Central

    Takata, Katsuyoshi; Kawano, Seiji; Fujii, Nobuharu; Kawahara, Yoshiro; Yoshino, Tadashi; Okada, Hiroyuki

    2016-01-01

    A 70-year-old woman presented with follicular lymphoma involving the stomach, duodenum, jejunum, bone, and lymph nodes. Esophagogastroduodenoscopy revealed multiple depressed lesions in the stomach. Examination with magnifying endoscopy showed branched abnormal vessels along with gastric pits, which were irregularly shaped but were preserved. The second case was a 45-year-old man diagnosed with stage II1 follicular lymphoma with duodenal, ileal, and colorectal involvement, as well as lymphadenopathy of the mesenteric lymph nodes. Esophagogastroduodenoscopy performed six years after the diagnosis revealed multiple erosions in the gastric body and angle. Magnifying endoscopic observation with narrow-band imaging showed that the gastric pits were only partially preserved and were destroyed in most of the stomach. Branched abnormal vessels were also seen. Pathological features were consistent with follicular lymphoma in both cases. The structural differences reported between the two cases appear to reflect distinct pathologies. Disappearance of gastric pits in the latter case seems to result from loss of epithelial cells, probably due to chronic inflammation. In both cases, branched abnormal vasculature was observed. These two cases suggest that magnified observations of abnormal branched microvasculature may facilitate endoscopic detection and recognition of the extent of gastric involvement in patients with follicular lymphoma. PMID:27747111

  11. What induces pocket openings on protein surface patches involved in protein-protein interactions?

    NASA Astrophysics Data System (ADS)

    Eyrisch, Susanne; Helms, Volkhard

    2009-02-01

    We previously showed for the proteins BCL-XL, IL-2, and MDM2 that transient pockets at their protein-protein binding interfaces can be identified by applying the PASS algorithm to molecular dynamics (MD) snapshots. We now investigated which aspects of the natural conformational dynamics of proteins induce the formation of such pockets. The pocket detection protocol was applied to three different conformational ensembles for the same proteins that were extracted from MD simulations of the inhibitor bound crystal conformation in water and the free crystal/NMR structure in water and in methanol. Additional MD simulations studied the impact of backbone mobility. The more efficient CONCOORD or normal mode analysis (NMA) techniques gave significantly smaller pockets than MD simulations, whereas tCONCOORD generated pockets comparable to those observed in MD simulations for two of the three systems. Our findings emphasize the influence of solvent polarity and backbone rearrangements on the formation of pockets on protein surfaces and should be helpful in future generation of transient pockets as putative ligand binding sites at protein-protein interfaces.

  12. Transient protein-protein interface prediction: datasets, features, algorithms, and the RAD-T predictor

    PubMed Central

    2014-01-01

    Background Transient protein-protein interactions (PPIs), which underly most biological processes, are a prime target for therapeutic development. Immense progress has been made towards computational prediction of PPIs using methods such as protein docking and sequence analysis. However, docking generally requires high resolution structures of both of the binding partners and sequence analysis requires that a significant number of recurrent patterns exist for the identification of a potential binding site. Researchers have turned to machine learning to overcome some of the other methods’ restrictions by generalising interface sites with sets of descriptive features. Best practices for dataset generation, features, and learning algorithms have not yet been identified or agreed upon, and an analysis of the overall efficacy of machine learning based PPI predictors is due, in order to highlight potential areas for improvement. Results The presence of unknown interaction sites as a result of limited knowledge about protein interactions in the testing set dramatically reduces prediction accuracy. Greater accuracy in labelling the data by enforcing higher interface site rates per domain resulted in an average 44% improvement across multiple machine learning algorithms. A set of 10 biologically unrelated proteins that were consistently predicted on with high accuracy emerged through our analysis. We identify seven features with the most predictive power over multiple datasets and machine learning algorithms. Through our analysis, we created a new predictor, RAD-T, that outperforms existing non-structurally specializing machine learning protein interface predictors, with an average 59% increase in MCC score on a dataset with a high number of interactions. Conclusion Current methods of evaluating machine-learning based PPI predictors tend to undervalue their performance, which may be artificially decreased by the presence of un-identified interaction sites. Changes to

  13. Clinical features and pathogenesis of Alzheimer's disease: involvement of mitochondria and mitochondrial DNA.

    PubMed

    Mancuso, Michelangelo; Orsucci, Daniele; LoGerfo, Annalisa; Calsolaro, Valeria; Siciliano, Gabriele

    2010-01-01

    Alzheimer's disease (AD) is a neurodegenerative disorder which results in the irreversible loss of cortical neurons, particularly in the associative neocortex and hippocampus. AD is the most common form of dementia in the elderly people. Apart from the neuronal loss, the pathological hallmarks are extracellular senile plaques containing the peptide beta-amyloid (AP) and neurofibrillary tangles. The Af cascade hypothesis remains the main pathogenetic model, as suggested by familiar AD, mainly associated to mutation in amyloid precursor protein and presenilin genes. The remaining 95% of AD patients are mostly sporadic late-onset cases, with a complex aetiology due to interactions between environmental conditions and genetic features of the individual. Mitochondria play a central role in the bioenergetics of the cell and apoptotic cell death. Morphological, biochemical and genetic abnormalities of the mitochondria in several AD tissues have been reported. Impaired mitochondrial respiration, particularly COX deficiency, has been observed in brain, platelets and fibroblasts of AD patients. Somatic mutations in mitochondrial DNA (mtDNA) could cause energy failure and increased oxidative stress. No causative mutations in the mtDNA have been detected and studies on mtDNA polymorphisms are controversial, but the "mitochondrial cascade hypothesis" here revised, could explain many of the biochemical, genetic and pathological features of sporadic AD.

  14. Distinctive serum protein profiles involving abundant proteins in lung cancer patients based upon antibody microarray analysis

    PubMed Central

    Gao, Wei-Min; Kuick, Rork; Orchekowski, Randal P; Misek, David E; Qiu, Ji; Greenberg, Alissa K; Rom, William N; Brenner, Dean E; Omenn, Gilbert S; Haab, Brian B; Hanash, Samir M

    2005-01-01

    Background Cancer serum protein profiling by mass spectrometry has uncovered mass profiles that are potentially diagnostic for several common types of cancer. However, direct mass spectrometric profiling has a limited dynamic range and difficulties in providing the identification of the distinctive proteins. We hypothesized that distinctive profiles may result from the differential expression of relatively abundant serum proteins associated with the host response. Methods Eighty-four antibodies, targeting a wide range of serum proteins, were spotted onto nitrocellulose-coated microscope slides. The abundances of the corresponding proteins were measured in 80 serum samples, from 24 newly diagnosed subjects with lung cancer, 24 healthy controls, and 32 subjects with chronic obstructive pulmonary disease (COPD). Two-color rolling-circle amplification was used to measure protein abundance. Results Seven of the 84 antibodies gave a significant difference (p < 0.01) for the lung cancer patients as compared to healthy controls, as well as compared to COPD patients. Proteins that exhibited higher abundances in the lung cancer samples relative to the control samples included C-reactive protein (CRP; a 13.3 fold increase), serum amyloid A (SAA; a 2.0 fold increase), mucin 1 and α-1-antitrypsin (1.4 fold increases). The increased expression levels of CRP and SAA were validated by Western blot analysis. Leave-one-out cross-validation was used to construct Diagonal Linear Discriminant Analysis (DLDA) classifiers. At a cutoff where all 56 of the non-tumor samples were correctly classified, 15/24 lung tumor patient sera were correctly classified. Conclusion Our results suggest that a distinctive serum protein profile involving abundant proteins may be observed in lung cancer patients relative to healthy subjects or patients with chronic disease and may have utility as part of strategies for detecting lung cancer. PMID:16117833

  15. GALT protein database: querying structural and functional features of GALT enzyme.

    PubMed

    d'Acierno, Antonio; Facchiano, Angelo; Marabotti, Anna

    2014-09-01

    Knowledge of the impact of variations on protein structure can enhance the comprehension of the mechanisms of genetic diseases related to that protein. Here, we present a new version of GALT Protein Database, a Web-accessible data repository for the storage and interrogation of structural effects of variations of the enzyme galactose-1-phosphate uridylyltransferase (GALT), the impairment of which leads to classic Galactosemia, a rare genetic disease. This new version of this database now contains the models of 201 missense variants of GALT enzyme, including heterozygous variants, and it allows users not only to retrieve information about the missense variations affecting this protein, but also to investigate their impact on substrate binding, intersubunit interactions, stability, and other structural features. In addition, it allows the interactive visualization of the models of variants collected into the database. We have developed additional tools to improve the use of the database by nonspecialized users. This Web-accessible database (http://bioinformatica.isa.cnr.it/GALT/GALT2.0) represents a model of tools potentially suitable for application to other proteins that are involved in human pathologies and that are subjected to genetic variations.

  16. Host membrane proteins involved in the replication of tobamovirus RNA.

    PubMed

    Ishibashi, Kazuhiro; Miyashita, Shuhei; Katoh, Etsuko; Ishikawa, Masayuki

    2012-12-01

    Eukaryotic positive-strand RNA viruses replicate their genomes in membrane-bound replication complexes composed of viral replication proteins and negative-strand RNA templates. These replication proteins are programmed to exhibit RNA polymerase and other replication-related activities only in replication complexes to avoid inducing double-stranded RNA-mediated host defenses. Host membrane components (e.g. proteins and lipids) should play important roles in the activation of replication proteins. Two host membrane proteins are components of the replication complex and activate the replication proteins of tobamoviruses. Interaction analyses using deletion mutants constructed based on structural information suggest a conformational change in replication proteins during the formation of a protein complex with RNA 5'-capping activity.

  17. Clinical features and types of articular involvement in patients with psoriatic arthritis.

    PubMed

    Dönmez, Salim; Pamuk, Ömer Nuri; Akker, Mustafa; Ak, Recep

    2015-06-01

    Psoriatic arthritis (PsA) is a psoriasis-associated inflammatory arthritis which causes joint destruction. There are some epidemiologic data about PsA; however, there are no sufficient data from Turkey. Herein, we evaluated the frequency of PsA in the Thrace region of Turkey according to hospital-based data. In addition, we evaluated clinical features and types of joint involvement in PsA patients. We included 172 PsA patients fulfilling CASPAR criteria admitted to the Division of Rheumatology, Trakya University Medical Faculty, between 2003 and 2012. Data from Turkish Statistical Institution was used to calculate the incidence and prevalence of PsA. Patients' demographic features, durations of psoriasis and PsA, number of tender and swollen joints, treatment modalities, laboratory data, and X-ray film findings were recorded from hospital files. The annual incidence of PsA was 2.8/100,000. The mean annual incidence was 3.47/100,000 in females and 2.15/100,000 in males. The overall prevalence of PsA in our region was 27.9/100,000 (95 % confidence interval (CI) 23.7-32.1) in individuals >16 years. The prevalence of PsA was higher in females than in males (34.7/100,000 vs. 21.5/100,000). Polyarthritis was present in 67 (38.9 %), oligoarthritis in 47 (27.3 %), spondyloarthritis in 39 (22.6 %), and distal interphalangeal (DIP) arthritis in 19 (11.0 %) patients. The duration of psoriasis was significantly longer in polyarticular PsA patients than in DIP and oligoarticular groups (p values = 0.016 and 0.018, respectively). The number of swollen joints correlated with age (r = 0.21, p = 0.006), duration of psoriasis (r = 0.20, p = 0.01), number of tender joints (r = 0.92, p ≤ 0.001), ESR (r = 0.24, p = 0.001), and CRP (r = 0.17, p = 0.026). The frequency of PsA in Thrace region is similar to that in low-frequency regions. The most frequent type of involvement was polyarticular, and it correlated with the duration of psoriasis

  18. Nail involvement in adult patients with plaque-type psoriasis: prevalence and clinical features*

    PubMed Central

    Schons, Karen Regina Rosso; Beber, André Avelino Costa; Beck, Maristela de Oliveira; Monticielo, Odirlei André

    2015-01-01

    BACKGROUND: Psoriasis is a disease of worldwide distribution with a prevalence of 1 to 3%. Nail psoriasis is estimated in 50% of patients with psoriasis, and in the presence of joint involvement, it can reach 80%. OBJECTIVE: To study the nail changes - and their clinical implications - presented by patients with psoriasis vulgaris under surveillance in a university hospital from the south of Brazil. METHODS: his cross-sectional study evaluated 65 adult patients from January 2012 to March 2013. Cutaneous severity was assessed according to the Psoriasis Area and Severity Index (PASI). The Nail Psoriasis Severity Index (NAPSI) was used to evaluate patient's nails. The diagnosis of psoriatic arthritis was established according to the Classification Criteria for Psoriatic Arthritis (CASPAR). RESULTS: The prevalence of NP was 46.1%. These patients had a median [interquartilic range (IQR)] NAPSI of 1 (0-15). A total of 63.3% of patients reported aesthetic discomfort or functional impairment related to their nails. Onycholysis was the most common feature (80%). When compared with patients without nail involvement, patients with NP had lower mean age at psoriasis onset [21 (18-41) vs. 43 (30-56) years, p=0,001]; longer disease duration [15.5 (10-24) vs. 6 (2-12) years, p=0.001]; higher PASI [9.2 (5-17) vs. 3.7 (2-10), p=0.044], higher frequency of psoriatic arthritis (43.3 vs. 3.7, p = 0.002) and more often reported family history of psoriasis (40% vs. 7.4%, p = 0.011). CONCLUSION: Onycholysis was the most frequent finding and most patients feel uncomfortable with the psoriatic nail changes that they experience. PMID:26131859

  19. Feature selection and classification of protein-protein complexes based on their binding affinities using machine learning approaches.

    PubMed

    Yugandhar, K; Gromiha, M Michael

    2014-09-01

    Protein-protein interactions are intrinsic to virtually every cellular process. Predicting the binding affinity of protein-protein complexes is one of the challenging problems in computational and molecular biology. In this work, we related sequence features of protein-protein complexes with their binding affinities using machine learning approaches. We set up a database of 185 protein-protein complexes for which the interacting pairs are heterodimers and their experimental binding affinities are available. On the other hand, we have developed a set of 610 features from the sequences of protein complexes and utilized Ranker search method, which is the combination of Attribute evaluator and Ranker method for selecting specific features. We have analyzed several machine learning algorithms to discriminate protein-protein complexes into high and low affinity groups based on their Kd values. Our results showed a 10-fold cross-validation accuracy of 76.1% with the combination of nine features using support vector machines. Further, we observed accuracy of 83.3% on an independent test set of 30 complexes. We suggest that our method would serve as an effective tool for identifying the interacting partners in protein-protein interaction networks and human-pathogen interactions based on the strength of interactions.

  20. DUF581 Is Plant Specific FCS-Like Zinc Finger Involved in Protein-Protein Interaction

    PubMed Central

    K, Muhammed Jamsheer; Laxmi, Ashverya

    2014-01-01

    Zinc fingers are a ubiquitous class of protein domain with considerable variation in structure and function. Zf-FCS is a highly diverged group of C2-C2 zinc finger which is present in animals, prokaryotes and viruses, but not in plants. In this study we identified that a plant specific domain of unknown function, DUF581 is a zf-FCS type zinc finger. Based on HMM-HMM comparison and signature motif similarity we named this domain as FCS-Like Zinc finger (FLZ) domain. A genome wide survey identified that FLZ domain containing genes are bryophytic in origin and this gene family is expanded in spermatophytes. Expression analysis of selected FLZ gene family members of A. thaliana identified an overlapping expression pattern suggesting a possible redundancy in their function. Unlike the zf-FCS domain, the FLZ domain found to be highly conserved in sequence and structure. Using a combination of bioinformatic and protein-protein interaction tools, we identified that FLZ domain is involved in protein-protein interaction. PMID:24901469

  1. Weak conservation of structural features in the interfaces of homologous transient protein-protein complexes.

    PubMed

    Sudha, Govindarajan; Singh, Prashant; Swapna, Lakshmipuram S; Srinivasan, Narayanaswamy

    2015-11-01

    Residue types at the interface of protein-protein complexes (PPCs) are known to be reasonably well conserved. However, we show, using a dataset of known 3-D structures of homologous transient PPCs, that the 3-D location of interfacial residues and their interaction patterns are only moderately and poorly conserved, respectively. Another surprising observation is that a residue at the interface that is conserved is not necessarily in the interface in the homolog. Such differences in homologous complexes are manifested by substitution of the residues that are spatially proximal to the conserved residue and structural differences at the interfaces as well as differences in spatial orientations of the interacting proteins. Conservation of interface location and the interaction pattern at the core of the interfaces is higher than at the periphery of the interface patch. Extents of variability of various structural features reported here for homologous transient PPCs are higher than the variation in homologous permanent homomers. Our findings suggest that straightforward extrapolation of interfacial nature and inter-residue interaction patterns from template to target could lead to serious errors in the modeled complex structure. Understanding the evolution of interfaces provides insights to improve comparative modeling of PPC structures.

  2. Identification of Protein Interactions Involved in Cellular Signaling

    PubMed Central

    Westermarck, Jukka; Ivaska, Johanna; Corthals, Garry L.

    2013-01-01

    Protein-protein interactions drive biological processes. They are critical for all intra- and extracellular functions, and the technologies to analyze them are widely applied throughout the various fields of biological sciences. This study takes an in-depth view of some common principles of cellular regulation and provides a detailed account of approaches required to comprehensively map signaling protein-protein interactions in any particular cellular system or condition. We provide a critical review of the benefits and disadvantages of the yeast two-hybrid method and affinity purification coupled with mass spectrometric procedures for identification of signaling protein-protein interactions. In particular, we emphasize the quantitative and qualitative differences between tandem affinity and one-step purification (such as FLAG and Strep tag) methods. Although applicable to all types of interaction studies, a special section is devoted in this review to aspects that should be considered when attempting to identify signaling protein interactions that often are transient and weak by nature. Finally, we discuss shotgun and quantitative information that can be gleaned by MS-coupled methods for analysis of multiprotein complexes. PMID:23481661

  3. The VHL short variant involves in protein quality control.

    PubMed

    Liu, Yanbin; Yang, Haixia; Zuo, Feifei; Chen, Liang

    2016-09-01

    The von Hippel-Lindau (VHL) is the most important and frequently mutated gene in human clear cell renal cell carcinoma (ccRCC). In contrast to its long counterpart, the internal translational variant of VHL protein (VHLs) is evolutionarily conserved. Herein we present evidence that VHLs associates with ribosome complex via interaction with the large subunit 6 (RPL6). Manipulation of VHLs expression significantly alters protein synthesis, cell size and mitochondrial mass. VHLs deficiency leads to remarkable sensitivity to drug treatments eliciting nascent protein mis-folding and translational errors. The ubiquitination of nascent peptides are dramatically increased upon the ectopic over-expression of VHLs, which simultaneously co-localizes with proteasome and thus may facilitate the ubiquitin-proteasome mediated degradation. In summary, VHLs contributes to protein quality control in addition to its canonical function in maintaining homeostasis of hypoxia-induced factors alpha subunit (HIFα) in response to environmental oxygen supply.

  4. Sex Hormones Regulate Cytoskeletal Proteins Involved in Brain Plasticity

    PubMed Central

    Hansberg-Pastor, Valeria; González-Arenas, Aliesha; Piña-Medina, Ana Gabriela; Camacho-Arroyo, Ignacio

    2015-01-01

    In the brain of female mammals, including humans, a number of physiological and behavioral changes occur as a result of sex hormone exposure. Estradiol and progesterone regulate several brain functions, including learning and memory. Sex hormones contribute to shape the central nervous system by modulating the formation and turnover of the interconnections between neurons as well as controlling the function of glial cells. The dynamics of neuron and glial cells morphology depends on the cytoskeleton and its associated proteins. Cytoskeletal proteins are necessary to form neuronal dendrites and dendritic spines, as well as to regulate the diverse functions in astrocytes. The expression pattern of proteins, such as actin, microtubule-associated protein 2, Tau, and glial fibrillary acidic protein, changes in a tissue-specific manner in the brain, particularly when variations in sex hormone levels occur during the estrous or menstrual cycles or pregnancy. Here, we review the changes in structure and organization of neurons and glial cells that require the participation of cytoskeletal proteins whose expression and activity are regulated by estradiol and progesterone. PMID:26635640

  5. Hierarchical learning architecture with automatic feature selection for multiclass protein fold classification.

    PubMed

    Huang, Chuen-Der; Lin, Chin-Teng; Pal, Nikhil Ranjan

    2003-12-01

    The structure classification of proteins plays a very important role in bioinformatics, since the relationships and characteristics among those known proteins can be exploited to predict the structure of new proteins. The success of a classification system depends heavily on two things: the tools being used and the features considered. For the bioinformatics applications, the role of appropriate features has not been paid adequate importance. In this investigation we use three novel ideas for multiclass protein fold classification. First, we use the gating neural network, where each input node is associated with a gate. This network can select important features in an online manner when the learning goes on. At the beginning of the training, all gates are almost closed, i.e., no feature is allowed to enter the network. Through the training, gates corresponding to good features are completely opened while gates corresponding to bad features are closed more tightly, and some gates may be partially open. The second novel idea is to use a hierarchical learning architecture (HLA). The classifier in the first level of HLA classifies the protein features into four major classes: all alpha, all beta, alpha + beta, and alpha/beta. And in the next level we have another set of classifiers, which further classifies the protein features into 27 folds. The third novel idea is to induce the indirect coding features from the amino-acid composition sequence of proteins based on the N-gram concept. This provides us with more representative and discriminative new local features of protein sequences for multiclass protein fold classification. The proposed HLA with new indirect coding features increases the protein fold classification accuracy by about 12%. Moreover, the gating neural network is found to reduce the number of features drastically. Using only half of the original features selected by the gating neural network can reach comparable test accuracy as that using all the

  6. Differentiation of HL60 cells: involvement of protein phosphorylation

    SciTech Connect

    Spearman, T.N.; Fontana, J.A.; Butcher, F.R.; Durham, J.P.

    1986-05-01

    The addition of retinoic acid (RA) to the human promyelocytic leukemic cell line HL60 in culture results in the cessation of growth and the acquisition of a more mature phenotype. Previous work in these laboratories has demonstrated a concomitant increase in the activity of calcium-dependent, phospholipid-sensitive protein kinase (PK-C). HL60 cells were incubated with /sup 32/P-P/sub i/ in the absence and presence of RA, homogenized, and aliquots subjected to two-dimensional electrophoresis. A comparison of autoradiograms made from these gels revealed several phosphoproteins whose radiolabeling was affected by RA. The radiolabeling of one particular phosphoprotein (49kd, pI 4.8) was found to be increased prior to phenotypic evidence of differentiation. It was demonstrated via incubating HL60 cytosol with /sup 32/P -ATP and Ca/sup 2 +/ in the absence and presence of phosphatidylserine and resolving the labeled proteins as above that this protein is phosphorylated by PK-C. The labeling of this protein was also increased by RA in other leukemic cell lines which showed phenotypic evidence of differentiation while no effect was seen in HL60 sublines resistant to RA or in mature neutrophils (the end product of myeloid differentiation). These results suggest that this protein may be an important intermediate in myeloid differentiation.

  7. Accelerated protein evolution and origins of human-specific features: Foxp2 as an example.

    PubMed

    Zhang, Jianzhi; Webb, David M; Podlaha, Ondrej

    2002-12-01

    Genes responsible for human-specific phenotypes may have been under altered selective pressures in human evolution and thus exhibit changes in substitution rate and pattern at the protein sequence level. Using comparative analysis of human, chimpanzee, and mouse protein sequences, we identified two genes (PRM2 and FOXP2) with significantly enhanced evolutionary rates in the hominid lineage. PRM2 is a histone-like protein essential to spermatogenesis and was previously reported to be a likely target of sexual selection in humans and chimpanzees. FOXP2 is a transcription factor involved in speech and language development. Human FOXP2 experienced a >60-fold increase in substitution rate and incorporated two fixed amino acid changes in a broadly defined transcription suppression domain. A survey of a diverse group of placental mammals reveals the uniqueness of the human FOXP2 sequence and a population genetic analysis indicates possible adaptive selection behind the accelerated evolution. Taken together, our results suggest an important role that FOXP2 may have played in the origin of human speech and demonstrate a strategy for identifying candidate genes underlying the emergences of human-specific features.

  8. Classification of protein-protein interaction full-text documents using text and citation network features.

    PubMed

    Kolchinsky, Artemy; Abi-Haidar, Alaa; Kaur, Jasleen; Hamed, Ahmed Abdeen; Rocha, Luis M

    2010-01-01

    We participated (as Team 9) in the Article Classification Task of the Biocreative II.5 Challenge: binary classification of full-text documents relevant for protein-protein interaction. We used two distinct classifiers for the online and offline challenges: 1) the lightweight Variable Trigonometric Threshold (VTT) linear classifier we successfully introduced in BioCreative 2 for binary classification of abstracts and 2) a novel Naive Bayes classifier using features from the citation network of the relevant literature. We supplemented the supplied training data with full-text documents from the MIPS database. The lightweight VTT classifier was very competitive in this new full-text scenario: it was a top-performing submission in this task, taking into account the rank product of the Area Under the interpolated precision and recall Curve, Accuracy, Balanced F-Score, and Matthew's Correlation Coefficient performance measures. The novel citation network classifier for the biomedical text mining domain, while not a top performing classifier in the challenge, performed above the central tendency of all submissions, and therefore indicates a promising new avenue to investigate further in bibliome informatics.

  9. Therapeutic approaches against common structural features of toxic oligomers shared by multiple amyloidogenic proteins.

    PubMed

    Guerrero-Muñoz, Marcos J; Castillo-Carranza, Diana L; Kayed, Rakez

    2014-04-15

    Impaired proteostasis is one of the main features of all amyloid diseases, which are associated with the formation of insoluble aggregates from amyloidogenic proteins. The aggregation process can be caused by overproduction or poor clearance of these proteins. However, numerous reports suggest that amyloid oligomers are the most toxic species, rather than insoluble fibrillar material, in Alzheimer's, Parkinson's, and Prion diseases, among others. Although the exact protein that aggregates varies between amyloid disorders, they all share common structural features that can be used as therapeutic targets. In this review, we focus on therapeutic approaches against shared features of toxic oligomeric structures and future directions.

  10. Characterization and Modulation of Proteins Involved in Sulfur Mustard Vesication

    DTIC Science & Technology

    2000-06-01

    SM may induce apoptosis as well. Recent evidence has revealed that Bcl-2 can complex with both the Caenorhabditis elegans death proteins 3 and 4 (Ced-3...the product of a gene required for programmed Tris-HCl (pH 6.8), and 0.02% bromophenol blue. Samples were re- cell death in Caenorhabditis elegans [17...biochemical or mor- which is required for apoptosis in Caenorhabditis elegans (8). phological changes characteristic of apoptosis when In human

  11. Identification of bacteriophage virion proteins by the ANOVA feature selection and analysis.

    PubMed

    Ding, Hui; Feng, Peng-Mian; Chen, Wei; Lin, Hao

    2014-08-01

    The bacteriophage virion proteins play extremely important roles in the fate of host bacterial cells. Accurate identification of bacteriophage virion proteins is very important for understanding their functions and clarifying the lysis mechanism of bacterial cells. In this study, a new sequence-based method was developed to identify phage virion proteins. In the new method, the protein sequences were initially formulated by the g-gap dipeptide compositions. Subsequently, the analysis of variance (ANOVA) with incremental feature selection (IFS) was used to search for the optimal feature set. It was observed that, in jackknife cross-validation, the optimal feature set including 160 optimized features can produce the maximum accuracy of 85.02%. By performing feature analysis, we found that the correlation between two amino acids with one gap was more important than other correlations for phage virion protein prediction and that some of the 1-gap dipeptides were important and mainly contributed to the virion protein prediction. This analysis will provide novel insights into the function of phage virion proteins. On the basis of the proposed method, an online web-server, PVPred, was established and can be freely accessed from the website (http://lin.uestc.edu.cn/server/PVPred). We believe that the PVPred will become a powerful tool to study phage virion proteins and to guide the related experimental validations.

  12. Centrophilin: a novel mitotic spindle protein involved in microtubule nucleation

    PubMed Central

    1991-01-01

    A novel protein has been identified which may serve a key function in nucleating spindle microtubule growth in mitosis. This protein, called centrophilin, is sequentially relocated from the centromeres to the centrosomes to the midbody in a manner dependent on the mitotic phase. Centrophilin was initially detected by immunofluorescence with a monoclonal, primate-specific antibody (2D3) raised against kinetochore- enriched chromosome extract from HeLa cells (Valdivia, M. M., and B. R. Brinkley. 1985. J. Cell Biol. 101:1124-1134). Centrophilin forms prominent crescents at the poles of the metaphase spindle, gradually diminishes during anaphase, and bands the equatorial ends of midbody microtubules in telophase. The formation and breakdown of the spindle and midbody correlates in time and space with the aggregation and disaggregation of centrophilin foci. Immunogold EM reveals that centrophilin is a major component of pericentriolar material in metaphase. During recovery from microtubule inhibition, centrophilin foci act as nucleation sites for the assembly of spindle tubules. The 2D3 probe recognizes two high molecular mass polypeptides, 180 and 210 kD, on immunoblots of whole HeLa cell extract. Taken together, these data and the available literature on microtubule dynamics point inevitably to a singular model for control of spindle tubule turnover. PMID:1991791

  13. Prediction of residue-residue contact matrix for protein-protein interaction with Fisher score features and deep learning.

    PubMed

    Du, Tianchuan; Liao, Li; Wu, Cathy H; Sun, Bilin

    2016-11-01

    Protein-protein interactions play essential roles in many biological processes. Acquiring knowledge of the residue-residue contact information of two interacting proteins is not only helpful in annotating functions for proteins, but also critical for structure-based drug design. The prediction of the protein residue-residue contact matrix of the interfacial regions is challenging. In this work, we introduced deep learning techniques (specifically, stacked autoencoders) to build deep neural network models to tackled the residue-residue contact prediction problem. In tandem with interaction profile Hidden Markov Models, which was used first to extract Fisher score features from protein sequences, stacked autoencoders were deployed to extract and learn hidden abstract features. The deep learning model showed significant improvement over the traditional machine learning model, Support Vector Machines (SVM), with the overall accuracy increased by 15% from 65.40% to 80.82%. We showed that the stacked autoencoders could extract novel features, which can be utilized by deep neural networks and other classifiers to enhance learning, out of the Fisher score features. It is further shown that deep neural networks have significant advantages over SVM in making use of the newly extracted features.

  14. Radiographic features of esophageal involvement in chronic graft-vs. -host disease

    SciTech Connect

    McDonald, G.B.; Sullivan, K.M.; Plumley, T.F.

    1984-03-01

    Chronic graft-vs.-host disease (GVHD) is an important late complication of allogeneic bone-marrow transplantation. It resembles several naturally occurring autoimmune diseases and involves the skin, mouth, eyes, liver, and esophagus. The radiographic findings of 14 symptomatic patients with chronic GVHD involving the esophagus were reviewed and found to include webs, ringlike narrowings, and tapering strictures in the mid and upper esophagus. Esophagoscopy revealed characteristic desquamation in the 13 patients studied, but barium studies detected this lesion in only one patient. Knowledge of the site and characteristics of esophageal involvement with chronic GVHD assists the radiologic evaluation of this disorder.

  15. A Method of Protein Model Classification and Retrieval Using Bag-of-Visual-Features

    PubMed Central

    Ma, Ziping; Kang, Baosheng; Lu, Ke

    2014-01-01

    In this paper we propose a novel visual method for protein model classification and retrieval. Different from the conventional methods, the key idea of the proposed method is to extract image features of proteins and measure the visual similarity between proteins. Firstly, the multiview images are captured by vertices and planes of a given octahedron surrounding the protein. Secondly, the local features are extracted from each image of the different views by the SURF algorithm and are vector quantized into visual words using a visual codebook. Finally, KLD is employed to calculate the similarity distance between two feature vectors. Experimental results show that the proposed method has encouraging performances for protein retrieval and categorization as shown in the comparison with other methods. PMID:25258644

  16. An automated approach to network features of protein structure ensembles

    PubMed Central

    Bhattacharyya, Moitrayee; Bhat, Chanda R; Vishveshwara, Saraswathi

    2013-01-01

    Network theory applied to protein structures provides insights into numerous problems of biological relevance. The explosion in structural data available from PDB and simulations establishes a need to introduce a standalone-efficient program that assembles network concepts/parameters under one hood in an automated manner. Herein, we discuss the development/application of an exhaustive, user-friendly, standalone program package named PSN-Ensemble, which can handle structural ensembles generated through molecular dynamics (MD) simulation/NMR studies or from multiple X-ray structures. The novelty in network construction lies in the explicit consideration of side-chain interactions among amino acids. The program evaluates network parameters dealing with topological organization and long-range allosteric communication. The introduction of a flexible weighing scheme in terms of residue pairwise cross-correlation/interaction energy in PSN-Ensemble brings in dynamical/chemical knowledge into the network representation. Also, the results are mapped on a graphical display of the structure, allowing an easy access of network analysis to a general biological community. The potential of PSN-Ensemble toward examining structural ensemble is exemplified using MD trajectories of an ubiquitin-conjugating enzyme (UbcH5b). Furthermore, insights derived from network parameters evaluated using PSN-Ensemble for single-static structures of active/inactive states of β2-adrenergic receptor and the ternary tRNA complexes of tyrosyl tRNA synthetases (from organisms across kingdoms) are discussed. PSN-Ensemble is freely available from http://vishgraph.mbu.iisc.ernet.in/PSN-Ensemble/psn_index.html. PMID:23934896

  17. Thoracic involvement in Behçet's disease: pathologic, clinical, and imaging features.

    PubMed

    Tunaci, A; Berkmen, Y M; Gökmen, E

    1995-01-01

    Behçet's disease is a rare form of vasculitis of obscure etiology. Any large or small artery, vein, or organ may be involved in an unpredictable combination. Intrathoracic manifestations of Behçet's disease consist mainly of thromboembolism of the superior vena cava and/or other mediastinal veins; aneurysms of the aorta and pulmonary arteries; pulmonary infarct and hemorrhage; pleural effusion; and, rarely, myocardial or pericardial involvement, cor pulmonale, and mediastinal or hilar lymphadenopathy. Chest radiography is the best diagnostic method for evaluating thoracic involvement in Behçet's disease. Because aneurysms may develop at the arterial puncture sites and veins may be quickly thrombosed after injection of contrast material, angiography and venography should be avoided whenever possible. Although no comparative studies are available, CT and MR angiography appear to be imaging techniques of choice for evaluating vascular involvement. Pulmonary parenchymal alterations depicted on CT scan have not been fully explored.

  18. Pseudomonas syringae pv. actinidiae draft genomes comparison reveal strain-specific features involved in adaptation and virulence to Actinidia species.

    PubMed

    Marcelletti, Simone; Ferrante, Patrizia; Petriccione, Milena; Firrao, Giuseppe; Scortichini, Marco

    2011-01-01

    A recent re-emerging bacterial canker disease incited by Pseudomonas syringae pv. actinidiae (Psa) is causing severe economic losses to Actinidia chinensis and A. deliciosa cultivations in southern Europe, New Zealand, Chile and South Korea. Little is known about the genetic features of this pathovar. We generated genome-wide Illumina sequence data from two Psa strains causing outbreaks of bacterial canker on the A. deliciosa cv. Hayward in Japan (J-Psa, type-strain of the pathovar) and in Italy (I-Psa) in 1984 and 1992, respectively as well as from a Psa strain (I2-Psa) isolated at the beginning of the recent epidemic on A. chinensis cv. Hort16A in Italy. All strains were isolated from typical leaf spot symptoms. The phylogenetic relationships revealed that Psa is more closely related to P. s. pv. theae than to P. avellanae within genomospecies 8. Comparative genomic analyses revealed both relevant intrapathovar variations and putative pathovar-specific genomic regions in Psa. The genomic sequences of J-Psa and I-Psa were very similar. Conversely, the I2-Psa genome encodes four additional effector protein genes, lacks a 50 kb plasmid and the phaseolotoxin gene cluster, argK-tox but has acquired a 160 kb plasmid and putative prophage sequences. Several lines of evidence from the analysis of the genome sequences support the hypothesis that this strain did not evolve from the Psa population that caused the epidemics in 1984-1992 in Japan and Italy but rather is the product of a recent independent evolution of the pathovar actinidiae for infecting Actinidia spp. All Psa strains share the genetic potential for copper resistance, antibiotic detoxification, high affinity iron acquisition and detoxification of nitric oxide of plant origin. Similar to other sequenced phytopathogenic pseudomonads associated with woody plant species, the Psa strains isolated from leaves also display a set of genes involved in the catabolism of plant-derived aromatic compounds.

  19. Pseudomonas syringae pv. actinidiae Draft Genomes Comparison Reveal Strain-Specific Features Involved in Adaptation and Virulence to Actinidia Species

    PubMed Central

    Marcelletti, Simone; Ferrante, Patrizia; Petriccione, Milena; Firrao, Giuseppe; Scortichini, Marco

    2011-01-01

    A recent re-emerging bacterial canker disease incited by Pseudomonas syringae pv. actinidiae (Psa) is causing severe economic losses to Actinidia chinensis and A. deliciosa cultivations in southern Europe, New Zealand, Chile and South Korea. Little is known about the genetic features of this pathovar. We generated genome-wide Illumina sequence data from two Psa strains causing outbreaks of bacterial canker on the A. deliciosa cv. Hayward in Japan (J-Psa, type-strain of the pathovar) and in Italy (I-Psa) in 1984 and 1992, respectively as well as from a Psa strain (I2-Psa) isolated at the beginning of the recent epidemic on A. chinensis cv. Hort16A in Italy. All strains were isolated from typical leaf spot symptoms. The phylogenetic relationships revealed that Psa is more closely related to P. s. pv. theae than to P. avellanae within genomospecies 8. Comparative genomic analyses revealed both relevant intrapathovar variations and putative pathovar-specific genomic regions in Psa. The genomic sequences of J-Psa and I-Psa were very similar. Conversely, the I2-Psa genome encodes four additional effector protein genes, lacks a 50 kb plasmid and the phaseolotoxin gene cluster, argK-tox but has acquired a 160 kb plasmid and putative prophage sequences. Several lines of evidence from the analysis of the genome sequences support the hypothesis that this strain did not evolve from the Psa population that caused the epidemics in 1984–1992 in Japan and Italy but rather is the product of a recent independent evolution of the pathovar actinidiae for infecting Actinidia spp. All Psa strains share the genetic potential for copper resistance, antibiotic detoxification, high affinity iron acquisition and detoxification of nitric oxide of plant origin. Similar to other sequenced phytopathogenic pseudomonads associated with woody plant species, the Psa strains isolated from leaves also display a set of genes involved in the catabolism of plant-derived aromatic compounds. PMID

  20. Prediction of protein secondary structure using probability based features and a hybrid system.

    PubMed

    Ghanty, Pradip; Pal, Nikhil R; Mudi, Rajani K

    2013-10-01

    In this paper, we propose some co-occurrence probability-based features for prediction of protein secondary structure. The features are extracted using occurrence/nonoccurrence of secondary structures in the protein sequences. We explore two types of features: position-specific (based on position of amino acid on fragments of protein sequences) as well as position-independent (independent of amino acid position on fragments of protein sequences). We use a hybrid system, NEUROSVM, consisting of neural networks and support vector machines for classification of secondary structures. We propose two schemes NSVMps and NSVM for protein secondary structure prediction. The NSVMps uses position-specific probability-based features and NEUROSVM classifier whereas NSVM uses the same classifier with position-independent probability-based features. The proposed method falls in the single-sequence category of methods because it does not use any sequence profile information such as position specific scoring matrices (PSSM) derived from PSI-BLAST. Two widely used datasets RS126 and CB513 are used in the experiments. The results obtained using the proposed features and NEUROSVM classifier are better than most of the existing single-sequence prediction methods. Most importantly, the results using NSVMps that are obtained using lower dimensional features, are comparable to those by other existing methods. The NSVMps and NSVM are finally tested on target proteins of the critical assessment of protein structure prediction experiment-9 (CASP9). A larger dataset is used to compare the performance of the proposed methods with that of two recent single-sequence prediction methods. We also investigate the impact of presence of different amino acid residues (in protein sequences) that are responsible for the formation of different secondary structures.

  1. Apolipoprotein A-IV: a protein intimately involved in metabolism

    PubMed Central

    Wang, Fei; Kohan, Alison B.; Lo, Chun-Min; Liu, Min; Howles, Philip; Tso, Patrick

    2015-01-01

    The purpose of this review is to summarize our current understanding of the physiological roles of apoA-IV in metabolism, and to underscore the potential for apoA-IV to be a focus for new therapies aimed at the treatment of diabetes and obesity-related disorders. ApoA-IV is primarily synthesized by the small intestine, attached to chylomicrons by enterocytes, and secreted into intestinal lymph during fat absorption. In circulation, apoA-IV is associated with HDL and chylomicron remnants, but a large portion is lipoprotein free. Due to its anti-oxidative and anti-inflammatory properties, and because it can mediate reverse-cholesterol transport, proposed functions of circulating apoA-IV have been related to protection from cardiovascular disease. This review, however, focuses primarily on several properties of apoA-IV that impact other metabolic functions related to food intake, obesity, and diabetes. In addition to participating in triglyceride absorption, apoA-IV can act as an acute satiation factor through both peripheral and central routes of action. It also modulates glucose homeostasis through incretin-like effects on insulin secretion, and by moderating hepatic glucose production. While apoA-IV receptors remain to be conclusively identified, the latter modes of action suggest that this protein holds therapeutic promise for treating metabolic disease. PMID:25640749

  2. Prediction of subcellular location apoptosis proteins with ensemble classifier and feature selection.

    PubMed

    Gu, Quan; Ding, Yong-Sheng; Jiang, Xiao-Ying; Zhang, Tong-Liang

    2010-04-01

    Apoptosis proteins have a central role in the development and the homeostasis of an organism. These proteins are very important for understanding the mechanism of programmed cell death. The function of an apoptosis protein is closely related to its subcellular location. It is crucial to develop powerful tools to predict apoptosis protein locations for rapidly increasing gap between the number of known structural proteins and the number of known sequences in protein databank. In this study, amino acids pair compositions with different spaces are used to construct feature sets for representing sample of protein feature selection approach based on binary particle swarm optimization, which is applied to extract effective feature. Ensemble classifier is used as prediction engine, of which the basic classifier is the fuzzy K-nearest neighbor. Each basic classifier is trained with different feature sets. Two datasets often used in prior works are selected to validate the performance of proposed approach. The results obtained by jackknife test are quite encouraging, indicating that the proposed method might become a potentially useful tool for subcellular location of apoptosis protein, or at least can play a complimentary role to the existing methods in the relevant areas. The supplement information and software written in Matlab are available by contacting the corresponding author.

  3. Evaluation of correlation between CT image features and ERCC1 protein expression in assessing lung cancer prognosis

    NASA Astrophysics Data System (ADS)

    Tan, Maxine; Emaminejad, Nastaran; Qian, Wei; Sun, Shenshen; Kang, Yan; Guan, Yubao; Lure, Fleming; Zheng, Bin

    2014-03-01

    Stage I non-small-cell lung cancers (NSCLC) usually have favorable prognosis. However, high percentage of NSCLC patients have cancer relapse after surgery. Accurately predicting cancer prognosis is important to optimally treat and manage the patients to minimize the risk of cancer relapse. Studies have shown that an excision repair crosscomplementing 1 (ERCC1) gene was a potentially useful genetic biomarker to predict prognosis of NSCLC patients. Meanwhile, studies also found that chronic obstructive pulmonary disease (COPD) was highly associated with lung cancer prognosis. In this study, we investigated and evaluated the correlations between COPD image features and ERCC1 gene expression. A database involving 106 NSCLC patients was used. Each patient had a thoracic CT examination and ERCC1 genetic test. We applied a computer-aided detection scheme to segment and quantify COPD image features. A logistic regression method and a multilayer perceptron network were applied to analyze the correlation between the computed COPD image features and ERCC1 protein expression. A multilayer perceptron network (MPN) was also developed to test performance of using COPD-related image features to predict ERCC1 protein expression. A nine feature based logistic regression analysis showed the average COPD feature values in the low and high ERCC1 protein expression groups are significantly different (p < 0.01). Using a five-fold cross validation method, the MPN yielded an area under ROC curve (AUC = 0.669±0.053) in classifying between the low and high ERCC1 expression cases. The study indicates that CT phenotype features are associated with the genetic tests, which may provide supplementary information to help improve accuracy in assessing prognosis of NSCLC patients.

  4. The Werner Syndrome Protein Is Involved in RNA Polymerase II Transcription

    PubMed Central

    Balajee, Adayabalam S.; Machwe, Amrita; May, Alfred; Gray, Matthew D.; Oshima, Junko; Martin, George M.; Nehlin, Jan O.; Brosh, Robert; Orren, David K.; Bohr, Vilhelm A.

    1999-01-01

    Werner syndrome (WS) is a human progeroid syndrome characterized by the early onset of a large number of clinical features associated with the normal aging process. The complex molecular and cellular phenotypes of WS involve characteristic features of genomic instability and accelerated replicative senescence. The gene involved (WRN) was recently cloned, and its gene product (WRNp) was biochemically characterized as a helicase. Helicases play important roles in a variety of DNA transactions, including DNA replication, transcription, repair, and recombination. We have assessed the role of the WRN gene in transcription by analyzing the efficiency of basal transcription in WS lymphoblastoid cell lines that carry homozygous WRN mutations. Transcription was measured in permeabilized cells by [3H]UTP incorporation and in vitro by using a plasmid template containing the RNA polymerase II (RNA pol II)–dependent adenovirus major late promoter. With both of these approaches, we find that the transcription efficiency in different WS cell lines is reduced to 40–60% of the transcription in cells from normal individuals. This defect can be complemented by the addition of normal cell extracts to the chromatin of WS cells. Addition of purified wild-type WRNp but not mutated WRNp to the in vitro transcription assay markedly stimulates RNA pol II–dependent transcription carried out by nuclear extracts. A nonhelicase domain (a direct repeat of 27 amino acids) also appears to have a role in transcription enhancement, as revealed by a yeast hybrid–protein reporter assay. This is further supported by the lack of stimulation of transcription when mutant WRNp lacking this domain was added to the in vitro assay. We have thus used several approaches to show a role for WRNp in RNA pol II transcription, possibly as a transcriptional activator. A deficit in either global or regional transcription in WS cells may be a primary molecular defect responsible for the WS clinical phenotype

  5. Cell-surface Attachment of Bacterial Multienzyme Complexes Involves Highly Dynamic Protein-Protein Anchors*

    PubMed Central

    Cameron, Kate; Najmudin, Shabir; Alves, Victor D.; Bayer, Edward A.; Smith, Steven P.; Bule, Pedro; Waller, Helen; Ferreira, Luís M. A.; Gilbert, Harry J.; Fontes, Carlos M. G. A.

    2015-01-01

    Protein-protein interactions play a pivotal role in the assembly of the cellulosome, one of nature's most intricate nanomachines dedicated to the depolymerization of complex carbohydrates. The integration of cellulosomal components usually occurs through the binding of type I dockerin modules located at the C terminus of the enzymes to cohesin modules located in the primary scaffoldin subunit. Cellulosomes are typically recruited to the cell surface via type II cohesin-dockerin interactions established between primary and cell-surface anchoring scaffoldin subunits. In contrast with type II interactions, type I dockerins usually display a dual binding mode that may allow increased conformational flexibility during cellulosome assembly. Acetivibrio cellulolyticus produces a highly complex cellulosome comprising an unusual adaptor scaffoldin, ScaB, which mediates the interaction between the primary scaffoldin, ScaA, through type II cohesin-dockerin interactions and the anchoring scaffoldin, ScaC, via type I cohesin-dockerin interactions. Here, we report the crystal structure of the type I ScaB dockerin in complex with a type I ScaC cohesin in two distinct orientations. The data show that the ScaB dockerin displays structural symmetry, reflected by the presence of two essentially identical binding surfaces. The complex interface is more extensive than those observed in other type I complexes, which results in an ultra-high affinity interaction (Ka ∼1012 m). A subset of ScaB dockerin residues was also identified as modulating the specificity of type I cohesin-dockerin interactions in A. cellulolyticus. This report reveals that recruitment of cellulosomes onto the cell surface may involve dockerins presenting a dual binding mode to incorporate additional flexibility into the quaternary structure of highly populated multienzyme complexes. PMID:25855788

  6. Sturgeon osteocalcin shares structural features with matrix Gla protein: evolutionary relationship and functional implications.

    PubMed

    Viegas, Carla S B; Simes, Dina C; Williamson, Matthew K; Cavaco, Sofia; Laizé, Vincent; Price, Paul A; Cancela, M Leonor

    2013-09-27

    Osteocalcin (OC) and matrix Gla protein (MGP) are considered evolutionarily related because they share key structural features, although they have been described to exert different functions. In this work, we report the identification and characterization of both OC and MGP from the Adriatic sturgeon, a ray-finned fish characterized by a slow evolution and the retention of many ancestral features. Sturgeon MGP shows a primary structure, post-translation modifications, and patterns of mRNA/protein distribution and accumulation typical of known MGPs, and it contains seven possible Gla residues that would make the sturgeon protein the most γ-carboxylated among known MGPs. In contrast, sturgeon OC was found to present a hybrid structure. Indeed, although exhibiting protein domains typical of known OCs, it also contains structural features usually found in MGPs (e.g. a putative phosphorylated propeptide). Moreover, patterns of OC gene expression and protein accumulation overlap with those reported for MGP; OC was detected in bone cells and mineralized structures but also in soft and cartilaginous tissues. We propose that, in a context of a reduced rate of evolution, sturgeon OC has retained structural features of the ancestral protein that emerged millions of years ago from the duplication of an ancient MGP gene and may exhibit intermediate functional features.

  7. Papillary thyroid carcinoma involving cervical neck lymph nodes: correlations with lymphangiogenesis and ultrasound features.

    PubMed

    Choi, Yoonjung; Park, Kyung Joo; Ryu, Seungho; Kim, Dong-Hoon; Yun, Jisup; Kang, Doo Kyoung; Chun, Mison

    2012-01-01

    Stratification of risk factors for cervical lymph node metastasis (LNM) in thyroid papillary carcinoma is important for providing standards for post-operative adjuvant radio-iodine therapy and for patient prognosis. We investigated pathological factors based on the lymphatic vessel system and radiological features associated with tumor with cervical neck LNM. Among patients who had undergone thyroidectomy confirmed to be papillary thyroid carcinoma, we selected 126 age-sex matched paired patients without cervical LNM (group 1) and with LNM (group 2) to evaluate risk factors. Pathological factors evaluated were size, multiplicity, and extra thyroid extension state, based on the pathological reports using stored data. The lymphatic vessel density (LVD) of each tumor was evaluated by staining for VEGFR-3 and D2-40 and correlated with cervical LNM state. Malignant ultrasound features were evaluated to compare the differences between these two groups. Larger tumor size, multiplicity, extrathyroid extension were more common in group 2 (p<0.05). The median percentage of VEGFR-3 for group 1 was 20 (range 0-30) and D2-40 was 13 (range 7-23) while for group 2, VEGFR-3 was 80 (70-90) and D2-40 was 78 (54-114). LVD measured by intratumoral D2-40 staining was 20.6% and 79.4% for group 1 and group 2, respectively. Intra-tumoral lymphatics measured by D2-40 stain had a strong correlation with cervical LNM (Odds 1.230, CI 1.01.-1.499 p value 0.040). Ultrasound (US) features had no significant differences between the two groups although calcifications tended to be higher in group 2 (84% vs. 76% p=0.264). Lymphatic vessel density and nodule echogenicity were not associated with LNM. Intratumoral lymphangiogenesis was most strongly associated with LNM and thus, could be a useful predictive marker for cervical LNM.

  8. P-proteins in Arabidopsis are heteromeric structures involved in rapid sieve tube sealing.

    PubMed

    Jekat, Stephan B; Ernst, Antonia M; von Bohl, Andreas; Zielonka, Sascia; Twyman, Richard M; Noll, Gundula A; Prüfer, Dirk

    2013-01-01

    Structural phloem proteins (P-proteins) are characteristic components of the sieve elements in all dicotyledonous and many monocotyledonous angiosperms. Tobacco P-proteins were recently confirmed to be encoded by the widespread sieve element occlusion (SEO) gene family, and tobacco SEO proteins were shown to be directly involved in sieve tube sealing thus preventing the loss of photosynthate. Analysis of the two Arabidopsis SEO proteins (AtSEOa and AtSEOb) indicated that the corresponding P-protein subunits do not act in a redundant manner. However, there are still pending questions regarding the interaction properties and specific functions of AtSEOa and AtSEOb as well as the general function of structural P-proteins in Arabidopsis. In this study, we characterized the Arabidopsis P-proteins in more detail. We used in planta bimolecular fluorescence complementation assays to confirm the predicted heteromeric interactions between AtSEOa and AtSEOb. Arabidopsis mutants depleted for one or both AtSEO proteins lacked the typical P-protein structures normally found in sieve elements, underlining the identity of AtSEO proteins as P-proteins and furthermore providing the means to determine the role of Arabidopsis P-proteins in sieve tube sealing. We therefore developed an assay based on phloem exudation. Mutants with reduced AtSEO expression levels lost twice as much photosynthate following injury as comparable wild-type plants, confirming that Arabidopsis P-proteins are indeed involved in sieve tube sealing.

  9. Features of an interactive writing discourse: conversational involvement, conventional knowledge, and internalization in "Morning Message".

    PubMed

    Mariage, T V

    2001-01-01

    This study describes how meaning potentials were constructed in the literacy event known as Morning Message. Morning Message provided teachers and students with opportunities to construct a written text around the experiences of one student. This discourse of writing allowed for the examination of how meaning was orchestrated and scaffolded between the teacher and her students. Three findings are discussed, including the function of a series of conversational involvement moves utilized by the teacher, the specific writing conventions and metamessages afforded in the Morning Message dialogue, and an examination of how the social dialogues of Morning Message may have come to guide independent action as internalized processes on several transfer measures.

  10. Intrinsic fluorescence excitation-emission matrix spectral features of cottonseed protein fractions and the effects of denaturants

    Technology Transfer Automated Retrieval System (TEKTRAN)

    To better understand the functional and physicochemical properties of cottonseed protein, we investigated the intrinsic fluorescence excitation-emission matrix (EEM) spectral features of cottonseed protein isolate (CSPI) and sequentially extracted water (CSPw) and alkali (CSPa) protein fractions, an...

  11. Prediction of structural features and application to outer membrane protein identification

    PubMed Central

    Yan, Renxiang; Wang, Xiaofeng; Huang, Lanqing; Yan, Feidi; Xue, Xiaoyu; Cai, Weiwen

    2015-01-01

    Protein three-dimensional (3D) structures provide insightful information in many fields of biology. One-dimensional properties derived from 3D structures such as secondary structure, residue solvent accessibility, residue depth and backbone torsion angles are helpful to protein function prediction, fold recognition and ab initio folding. Here, we predict various structural features with the assistance of neural network learning. Based on an independent test dataset, protein secondary structure prediction generates an overall Q3 accuracy of ~80%. Meanwhile, the prediction of relative solvent accessibility obtains the highest mean absolute error of 0.164, and prediction of residue depth achieves the lowest mean absolute error of 0.062. We further improve the outer membrane protein identification by including the predicted structural features in a scoring function using a simple profile-to-profile alignment. The results demonstrate that the accuracy of outer membrane protein identification can be improved by ~3% at a 1% false positive level when structural features are incorporated. Finally, our methods are available as two convenient and easy-to-use programs. One is PSSM-2-Features for predicting secondary structure, relative solvent accessibility, residue depth and backbone torsion angles, the other is PPA-OMP for identifying outer membrane proteins from proteomes. PMID:26104144

  12. Prediction of structural features and application to outer membrane protein identification

    NASA Astrophysics Data System (ADS)

    Yan, Renxiang; Wang, Xiaofeng; Huang, Lanqing; Yan, Feidi; Xue, Xiaoyu; Cai, Weiwen

    2015-06-01

    Protein three-dimensional (3D) structures provide insightful information in many fields of biology. One-dimensional properties derived from 3D structures such as secondary structure, residue solvent accessibility, residue depth and backbone torsion angles are helpful to protein function prediction, fold recognition and ab initio folding. Here, we predict various structural features with the assistance of neural network learning. Based on an independent test dataset, protein secondary structure prediction generates an overall Q3 accuracy of ~80%. Meanwhile, the prediction of relative solvent accessibility obtains the highest mean absolute error of 0.164, and prediction of residue depth achieves the lowest mean absolute error of 0.062. We further improve the outer membrane protein identification by including the predicted structural features in a scoring function using a simple profile-to-profile alignment. The results demonstrate that the accuracy of outer membrane protein identification can be improved by ~3% at a 1% false positive level when structural features are incorporated. Finally, our methods are available as two convenient and easy-to-use programs. One is PSSM-2-Features for predicting secondary structure, relative solvent accessibility, residue depth and backbone torsion angles, the other is PPA-OMP for identifying outer membrane proteins from proteomes.

  13. Wetting of nonconserved residue-backbones: A feature indicative of aggregation associated regions of proteins.

    PubMed

    Pradhan, Mohan R; Pal, Arumay; Hu, Zhongqiao; Kannan, Srinivasaraghavan; Chee Keong, Kwoh; Lane, David P; Verma, Chandra S

    2016-02-01

    Aggregation is an irreversible form of protein complexation and often toxic to cells. The process entails partial or major unfolding that is largely driven by hydration. We model the role of hydration in aggregation using "Dehydrons." "Dehydrons" are unsatisfied backbone hydrogen bonds in proteins that seek shielding from water molecules by associating with ligands or proteins. We find that the residues at aggregation interfaces have hydrated backbones, and in contrast to other forms of protein-protein interactions, are under less evolutionary pressure to be conserved. Combining evolutionary conservation of residues and extent of backbone hydration allows us to distinguish regions on proteins associated with aggregation (non-conserved dehydron-residues) from other interaction interfaces (conserved dehydron-residues). This novel feature can complement the existing strategies used to investigate protein aggregation/complexation.

  14. Interfacial interactions involved in the biological assembly of Chandipura virus nucleocapsid protein.

    PubMed

    Sreejith, R; Gulati, Sahil; Gupta, Sanjay

    2013-06-01

    The biological assembly of Chandipura virus nucleocapsid (N) protein has been modeled and the amino acid residues involved in specific intermolecular interactions among N monomers during oligomerisation have been predicted.

  15. Large-scale identification of human protein function using topological features of interaction network

    PubMed Central

    Li, Zhanchao; Liu, Zhiqing; Zhong, Wenqian; Huang, Menghua; Wu, Na; Xie, Yun; Dai, Zong; Zou, Xiaoyong

    2016-01-01

    The annotation of protein function is a vital step to elucidate the essence of life at a molecular level, and it is also meritorious in biomedical and pharmaceutical industry. Developments of sequencing technology result in constant expansion of the gap between the number of the known sequences and their functions. Therefore, it is indispensable to develop a computational method for the annotation of protein function. Herein, a novel method is proposed to identify protein function based on the weighted human protein-protein interaction network and graph theory. The network topology features with local and global information are presented to characterise proteins. The minimum redundancy maximum relevance algorithm is used to select 227 optimized feature subsets and support vector machine technique is utilized to build the prediction models. The performance of current method is assessed through 10-fold cross-validation test, and the range of accuracies is from 67.63% to 100%. Comparing with other annotation methods, the proposed way possesses a 50% improvement in the predictive accuracy. Generally, such network topology features provide insights into the relationship between protein functions and network architectures. The source code of Matlab is freely available on request from the authors. PMID:27849060

  16. Site selectivity for protein tyrosine nitration: insights from features of structure and topological network.

    PubMed

    Cheng, Shangli; Lian, Baofeng; Liang, Juan; Shi, Ting; Xie, Lu; Zhao, Yi-Lei

    2013-11-01

    Tyrosine nitration is a covalent post-translational modification, which regulates protein functions such as hindering tyrosine phosphorylation and affecting essential signal transductions in cells. Based on up-to-date proteomics data, tyrosine nitration appears to be a highly selective process since not all tyrosine residues in proteins or all proteins are nitrated in vivo. Quite a few investigations included the protein structural information from the RCSB PDB database, where near 100,000 high-quality three-dimensional structures are available. In this work, we analyzed the local protein structures and amino acid topological networks of the nitrated and non-nitrated tyrosine sites in nitrated proteins, including neighboring atomic distribution, amino acid pair (AAP) and amino acid triangle (AAT). It has been found that aromatic and aliphatic residues, particularly with large volume, aromatic, aliphatic, or acidic side chains, are disfavored for the nitration. After integrating these structural features and topological network features with traditional sequence features, the predictive model achieves a sensitivity of 63.30% and a specificity of 92.24%, resulting in a much better accuracy compared to the previous models with only protein sequence information. Our investigation implies that the site selectivity may stem from a more open, hydrophilic and high-pH chemical environment around the tyrosine residue.

  17. Large-scale identification of human protein function using topological features of interaction network

    NASA Astrophysics Data System (ADS)

    Li, Zhanchao; Liu, Zhiqing; Zhong, Wenqian; Huang, Menghua; Wu, Na; Xie, Yun; Dai, Zong; Zou, Xiaoyong

    2016-11-01

    The annotation of protein function is a vital step to elucidate the essence of life at a molecular level, and it is also meritorious in biomedical and pharmaceutical industry. Developments of sequencing technology result in constant expansion of the gap between the number of the known sequences and their functions. Therefore, it is indispensable to develop a computational method for the annotation of protein function. Herein, a novel method is proposed to identify protein function based on the weighted human protein-protein interaction network and graph theory. The network topology features with local and global information are presented to characterise proteins. The minimum redundancy maximum relevance algorithm is used to select 227 optimized feature subsets and support vector machine technique is utilized to build the prediction models. The performance of current method is assessed through 10-fold cross-validation test, and the range of accuracies is from 67.63% to 100%. Comparing with other annotation methods, the proposed way possesses a 50% improvement in the predictive accuracy. Generally, such network topology features provide insights into the relationship between protein functions and network architectures. The source code of Matlab is freely available on request from the authors.

  18. Dominant-negative mutants of a yeast G-protein beta subunit identify two functional regions involved in pheromone signalling.

    PubMed Central

    Leberer, E; Dignard, D; Hougan, L; Thomas, D Y; Whiteway, M

    1992-01-01

    The STE4 gene, which encodes the beta subunit of the mating response G-protein in the yeast Saccharomyces cerevisiae, was subjected to a saturation mutagenesis using 'doped' oligodeoxynucleotides. We employed a genetic screen to select dominant-negative STE4 mutants, which when overexpressed from the GAL1 promoter, interfered with the signalling function of the wild type protein. The identified inhibitory amino acid alterations define two small regions that are crucially involved in transmitting the mating signal from G beta to downstream components of the signalling pathway. These results underline the positive signalling role of yeast G beta and assign for the first time the positive signalling function of a G-protein beta subunit to specific structural features. Images PMID:1464310

  19. Identification of the major lipoproteins in crayfish hemolymph as proteins involved in immune recognition and clotting.

    PubMed

    Hall, M; van Heusden, M C; Söderhäll, K

    1995-11-22

    Lipid-containing hemolymph proteins from males of the crayfish Pacifastacus leniusculus were isolated by density gradient ultracentrifugation. Two major lipoproteins, one high density lipoprotein (HDL) and one very high density lipoprotein (VHDL), were characterized. The HDL and the VHDL were found to be identical to two proteins previously studied for their roles in immune recognition and hemolymph clotting, namely the beta-1,3-glucan binding protein and the clotting protein. These results imply that crayfish lipoproteins have dual functions, and that they are involved in immunity, hemolymph clotting, and lipid transport in these animals. Also, the oxygen-transporting protein hemocyanin was found to have a small lipid content.

  20. Involvement of Iron-Containing Proteins in Genome Integrity in Arabidopsis Thaliana

    PubMed Central

    Zhang, Caiguo

    2015-01-01

    The Arabidopsis genome encodes numerous iron-containing proteins such as iron-sulfur (Fe-S) cluster proteins and hemoproteins. These proteins generally utilize iron as a cofactor, and they perform critical roles in photosynthesis, genome stability, electron transfer, and oxidation-reduction reactions. Plants have evolved sophisticated mechanisms to maintain iron homeostasis for the assembly of functional iron-containing proteins, thereby ensuring genome stability, cell development, and plant growth. Over the past few years, our understanding of iron-containing proteins and their functions involved in genome stability has expanded enormously. In this review, I provide the current perspectives on iron homeostasis in Arabidopsis, followed by a summary of iron-containing protein functions involved in genome stability maintenance and a discussion of their possible molecular mechanisms. PMID:27330736

  1. Transcriptional coexpression network reveals the involvement of varying stem cell features with different dysregulations in different gastric cancer subtypes.

    PubMed

    Kalamohan, Kalaivani; Periasamy, Jayaprakash; Bhaskar Rao, Divya; Barnabas, Georgina D; Ponnaiyan, Srigayatri; Ganesan, Kumaresan

    2014-10-01

    Despite the advancements in the cancer therapeutics, gastric cancer ranks as the second most common cancers with high global mortality rate. Integrative functional genomic investigation is a powerful approach to understand the major dysregulations and to identify the potential targets toward the development of targeted therapeutics for various cancers. Intestinal and diffuse type gastric tumors remain the major subtypes and the molecular determinants and drivers of these distinct subtypes remain unidentified. In this investigation, by exploring the network of gene coexpression association in gastric tumors, mRNA expressions of 20,318 genes across 200 gastric tumors were categorized into 21 modules. The genes and the hub genes of the modules show gastric cancer subtype specific expression. The expression patterns of the modules were correlated with intestinal and diffuse subtypes as well as with the differentiation status of gastric tumors. Among these, G1 module has been identified as a major driving force of diffuse type gastric tumors with the features of (i) enriched mesenchymal, mesenchymal stem cell like, and mesenchymal derived multiple lineages, (ii) elevated OCT1 mediated transcription, (iii) involvement of Notch activation, and (iv) reduced polycomb mediated epigenetic repression. G13 module has been identified as key factor in intestinal type gastric tumors and found to have the characteristic features of (i) involvement of embryonic stem cell like properties, (ii) Wnt, MYC and E2F mediated transcription programs, and (iii) involvement of polycomb mediated repression. Thus the differential transcription programs, differential epigenetic regulation and varying stem cell features involved in two major subtypes of gastric cancer were delineated by exploring the gene coexpression network. The identified subtype specific dysregulations could be optimally employed in developing subtype specific therapeutic targeting strategies for gastric cancer.

  2. FALDO: a semantic standard for describing the location of nucleotide and protein feature annotation

    DOE PAGES

    Bolleman, Jerven T.; Mungall, Christopher J.; Strozzi, Francesco; ...

    2016-06-13

    Nucleotide and protein sequence feature annotations are essential to understand biology on the genomic, transcriptomic, and proteomic level. Using Semantic Web technologies to query biological annotations, there was no standard that described this potentially complex location information as subject-predicate-object triples. In this paper, we have developed an ontology, the Feature Annotation Location Description Ontology (FALDO), to describe the positions of annotated features on linear and circular sequences. FALDO can be used to describe nucleotide features in sequence records, protein annotations, and glycan binding sites, among other features in coordinate systems of the aforementioned “omics” areas. Using the same data formatmore » to represent sequence positions that are independent of file formats allows us to integrate sequence data from multiple sources and data types. The genome browser JBrowse is used to demonstrate accessing multiple SPARQL endpoints to display genomic feature annotations, as well as protein annotations from UniProt mapped to genomic locations. Our ontology allows users to uniformly describe – and potentially merge – sequence annotations from multiple sources. Finally, data sources using FALDO can prospectively be retrieved using federalised SPARQL queries against public SPARQL endpoints and/or local private triple stores.« less

  3. FALDO: a semantic standard for describing the location of nucleotide and protein feature annotation

    SciTech Connect

    Bolleman, Jerven T.; Mungall, Christopher J.; Strozzi, Francesco; Baran, Joachim; Dumontier, Michel; Bonnal, Raoul J. P.; Buels, Robert; Hoehndorf, Robert; Fujisawa, Takatomo; Katayama, Toshiaki; Cock, Peter J. A.

    2016-06-13

    Nucleotide and protein sequence feature annotations are essential to understand biology on the genomic, transcriptomic, and proteomic level. Using Semantic Web technologies to query biological annotations, there was no standard that described this potentially complex location information as subject-predicate-object triples. In this paper, we have developed an ontology, the Feature Annotation Location Description Ontology (FALDO), to describe the positions of annotated features on linear and circular sequences. FALDO can be used to describe nucleotide features in sequence records, protein annotations, and glycan binding sites, among other features in coordinate systems of the aforementioned “omics” areas. Using the same data format to represent sequence positions that are independent of file formats allows us to integrate sequence data from multiple sources and data types. The genome browser JBrowse is used to demonstrate accessing multiple SPARQL endpoints to display genomic feature annotations, as well as protein annotations from UniProt mapped to genomic locations. Our ontology allows users to uniformly describe – and potentially merge – sequence annotations from multiple sources. Finally, data sources using FALDO can prospectively be retrieved using federalised SPARQL queries against public SPARQL endpoints and/or local private triple stores.

  4. Increased functional protein expression using nucleotide sequence features enriched in highly expressed genes in zebrafish.

    PubMed

    Horstick, Eric J; Jordan, Diana C; Bergeron, Sadie A; Tabor, Kathryn M; Serpe, Mihaela; Feldman, Benjamin; Burgess, Harold A

    2015-04-20

    Many genetic manipulations are limited by difficulty in obtaining adequate levels of protein expression. Bioinformatic and experimental studies have identified nucleotide sequence features that may increase expression, however it is difficult to assess the relative influence of these features. Zebrafish embryos are rapidly injected with calibrated doses of mRNA, enabling the effects of multiple sequence changes to be compared in vivo. Using RNAseq and microarray data, we identified a set of genes that are highly expressed in zebrafish embryos and systematically analyzed for enrichment of sequence features correlated with levels of protein expression. We then tested enriched features by embryo microinjection and functional tests of multiple protein reporters. Codon selection, releasing factor recognition sequence and specific introns and 3' untranslated regions each increased protein expression between 1.5- and 3-fold. These results suggested principles for increasing protein yield in zebrafish through biomolecular engineering. We implemented these principles for rational gene design in software for codon selection (CodonZ) and plasmid vectors incorporating the most active non-coding elements. Rational gene design thus significantly boosts expression in zebrafish, and a similar approach will likely elevate expression in other animal models.

  5. Structural feature extraction protocol for classifying reversible membrane binding protein domains.

    PubMed

    Källberg, Morten; Lu, Hui

    2009-01-01

    Machine learning based classification protocols for automated function annotation of protein structures have in many instances proven superior to simpler sequence based procedures. Here we present an automated method for extracting features from protein structures by construction of surface patches to be used in such protocols. The utility of the developed patch-growing procedure is exemplified by its ability to identify reversible membrane binding domains from the C1, C2, and PH families.

  6. Bead Based Proteome Enrichment Enhances Features of the Protein Elution Plate (PEP) for Functional Proteomic Profiling

    PubMed Central

    Wang, Xing; Davies, Michael; Roy, Swapan; Kuruc, Matthew

    2015-01-01

    A novel functional proteomics technology called PEP(Protein Elution Plate) was developed to separate complex proteomes from natural sources and analyze protein functions systematically. The technology takes advantage of the powerful resolution of two-dimensional gel electrophoresis (2-D Gels). The modification of electrophoretic conditions in combination with a high-resolution protein elution plate supports the recovery of functionally active proteins. As 2DE(2-Dimensional Electrophoresis) resolution can be limited by protein load, we investigated the use of bead based enrichment technologies, called AlbuVoid™ and KinaSorb™ to determine their effect on the proteomic features which can be generated from the PEP platform. Using a variety of substrates and enzyme activity assays, we report on the benefits of combining bead based enrichment to improve the signal report and the features generated for Hexokinase, Protein Kinase, Protease, and Alkaline Phosphatase activities. As a result, the PEP technology allows systematic analysis of large enzyme families and can build a comprehensive picture of protein function from a complex proteome, providing biological insights that could otherwise not be observed if only protein abundances were analyzed. PMID:28248280

  7. FASTERp: A Feature Array Search Tool for Estimating Resemblance of Protein Sequences

    SciTech Connect

    Macklin, Derek; Egan, Rob; Wang, Zhong

    2014-03-14

    Metagenome sequencing efforts have provided a large pool of billions of genes for identifying enzymes with desirable biochemical traits. However, homology search with billions of genes in a rapidly growing database has become increasingly computationally impractical. Here we present our pilot efforts to develop a novel alignment-free algorithm for homology search. Specifically, we represent individual proteins as feature vectors that denote the presence or absence of short kmers in the protein sequence. Similarity between feature vectors is then computed using the Tanimoto score, a distance metric that can be rapidly computed on bit string representations of feature vectors. Preliminary results indicate good correlation with optimal alignment algorithms (Spearman r of 0.87, ~;;1,000,000 proteins from Pfam), as well as with heuristic algorithms such as BLAST (Spearman r of 0.86, ~;;1,000,000 proteins). Furthermore, a prototype of FASTERp implemented in Python runs approximately four times faster than BLAST on a small scale dataset (~;;1000 proteins). We are optimizing and scaling to improve FASTERp to enable rapid homology searches against billion-protein databases, thereby enabling more comprehensive gene annotation efforts.

  8. Unusual features of protein interaction in human immunodeficiency virus (HIV) virions.

    PubMed

    Bukrinskaya, A G; Sharova, N K

    1990-01-01

    The treatment of HIV-1 virions with ionic and nonionic detergents (NP 40, octylglucoside, Na deoxycholate) resulted in an effect unusual for enveloped viruses: instead of solubilization of glycoproteins, the core protein p24 was solubilized while envelope glycoproteins with other structural proteins were found in subviral particles. These data are consistent with a model of HIV structural organization in which glycoproteins are included in the matrix formed by the protein p 17 and suggest that p24 is neither involved in the matrix nor closely bound to any viral proteins.

  9. The anterior temporal lobes are critically involved in acquiring new conceptual knowledge: evidence for impaired feature integration in semantic dementia.

    PubMed

    Hoffman, Paul; Evans, Gemma A L; Lambon Ralph, Matthew A

    2014-01-01

    Recent evidence from multiple neuroscience techniques indicates that regions within the anterior temporal lobes (ATLs) are a critical node in the neural network for representing conceptual knowledge, yet their function remains elusive. The hub-and-spoke model holds that ATL regions act as a transmodal conceptual hub, distilling the various sensory-motor features of objects and words into integrated, coherent conceptual representations. Single-cell recordings in monkeys suggest that the ATLs are critically involved in visual associative learning; however, investigations of this region in humans have focused on existing knowledge rather than learning. We studied acquisition of new concepts in semantic dementia patients, who have cortical damage centred on the ventrolateral aspects of the ATLs. Patients learned to assign abstract visual stimuli to two categories. The categories conformed to a family resemblance structure in which no individual stimulus features were fully diagnostic; thus the task required participants to form representations that integrate multiple features into a single concept. Patients were unable to do this, instead responding only on the basis of individual features. The study reveals that integrating disparate sources of information into novel coherent concepts is a critical computational function of the ATLs. This explains the central role of this region in conceptual representation and the catastrophic breakdown of concepts in semantic dementia.

  10. Compositional features are potentially involved in the regulation of gene expression of tumor suppressor genes in human tissues.

    PubMed

    Hajjari, Mohammadreza; Khoshnevisan, Atefeh; Behmanesh, Mehrdad

    2014-12-15

    Different mechanisms regulate the expression level of tissue specific genes in human. Here we report some compositional features such as codon usage bias, amino acid usage bias, codon frequency, and base composition which may be potentially related to mRNA amount of tissue specific tumor suppressor genes. Our findings support the possibility that structural elements in gene and protein may play an important role in the regulation of tumor suppressor genes, development, and tumorigenesis. The data presented here can open broad vistas in the understanding and treatment of a variety of human malignancies.

  11. GUN1 Controls Accumulation of the Plastid Ribosomal Protein S1 at the Protein Level and Interacts with Proteins Involved in Plastid Protein Homeostasis1

    PubMed Central

    Pesaresi, Paolo; Rossi, Fabio; Guljamow, Arthur; Sommer, Frederik; Mühlhaus, Timo; Schroda, Michael; Masiero, Simona; Rothbart, Maxi; Hedtke, Boris

    2016-01-01

    Developmental or metabolic changes in chloroplasts can have profound effects on the rest of the plant cell. Such intracellular responses are associated with signals that originate in chloroplasts and convey information on their physiological status to the nucleus, which leads to large-scale changes in gene expression (retrograde signaling). A screen designed to identify components of retrograde signaling resulted in the discovery of the so-called genomes uncoupled (gun) mutants. Genetic evidence suggests that the chloroplast protein GUN1 integrates signals derived from perturbations in plastid redox state, plastid gene expression, and tetrapyrrole biosynthesis (TPB) in Arabidopsis (Arabidopsis thaliana) seedlings, exerting biogenic control of chloroplast functions. However, the molecular mechanism by which GUN1 integrates retrograde signaling in the chloroplast is unclear. Here we show that GUN1 also operates in adult plants, contributing to operational control of chloroplasts. The gun1 mutation genetically interacts with mutations of genes for the chloroplast ribosomal proteins S1 (PRPS1) and L11. Analysis of gun1 prps1 lines indicates that GUN1 controls PRPS1 accumulation at the protein level. The GUN1 protein physically interacts with proteins involved in chloroplast protein homeostasis based on coimmunoprecipitation experiments. Furthermore, yeast two-hybrid and bimolecular fluorescence complementation experiments suggest that GUN1 might transiently interact with several TPB enzymes, including Mg-chelatase subunit D (CHLD) and two other TPB enzymes known to activate retrograde signaling. Moreover, the association of PRPS1 and CHLD with protein complexes is modulated by GUN1. These findings allow us to speculate that retrograde signaling might involve GUN1-dependent formation of protein complexes. PMID:26823545

  12. GUN1 Controls Accumulation of the Plastid Ribosomal Protein S1 at the Protein Level and Interacts with Proteins Involved in Plastid Protein Homeostasis.

    PubMed

    Tadini, Luca; Pesaresi, Paolo; Kleine, Tatjana; Rossi, Fabio; Guljamow, Arthur; Sommer, Frederik; Mühlhaus, Timo; Schroda, Michael; Masiero, Simona; Pribil, Mathias; Rothbart, Maxi; Hedtke, Boris; Grimm, Bernhard; Leister, Dario

    2016-03-01

    Developmental or metabolic changes in chloroplasts can have profound effects on the rest of the plant cell. Such intracellular responses are associated with signals that originate in chloroplasts and convey information on their physiological status to the nucleus, which leads to large-scale changes in gene expression (retrograde signaling). A screen designed to identify components of retrograde signaling resulted in the discovery of the so-called genomes uncoupled (gun) mutants. Genetic evidence suggests that the chloroplast protein GUN1 integrates signals derived from perturbations in plastid redox state, plastid gene expression, and tetrapyrrole biosynthesis (TPB) in Arabidopsis (Arabidopsis thaliana) seedlings, exerting biogenic control of chloroplast functions. However, the molecular mechanism by which GUN1 integrates retrograde signaling in the chloroplast is unclear. Here we show that GUN1 also operates in adult plants, contributing to operational control of chloroplasts. The gun1 mutation genetically interacts with mutations of genes for the chloroplast ribosomal proteins S1 (PRPS1) and L11. Analysis of gun1 prps1 lines indicates that GUN1 controls PRPS1 accumulation at the protein level. The GUN1 protein physically interacts with proteins involved in chloroplast protein homeostasis based on coimmunoprecipitation experiments. Furthermore, yeast two-hybrid and bimolecular fluorescence complementation experiments suggest that GUN1 might transiently interact with several TPB enzymes, including Mg-chelatase subunit D (CHLD) and two other TPB enzymes known to activate retrograde signaling. Moreover, the association of PRPS1 and CHLD with protein complexes is modulated by GUN1. These findings allow us to speculate that retrograde signaling might involve GUN1-dependent formation of protein complexes.

  13. The crystal structure of the thiocyanate-forming protein from Thlaspi arvense, a kelch protein involved in glucosinolate breakdown.

    PubMed

    Gumz, Frauke; Krausze, Joern; Eisenschmidt, Daniela; Backenköhler, Anita; Barleben, Leif; Brandt, Wolfgang; Wittstock, Ute

    2015-09-01

    Kelch repeat-containing proteins are involved in diverse cellular processes, but only a small subset of plant kelch proteins has been functionally characterized. Thiocyanate-forming protein (TFP) from field-penny cress, Thlaspi arvense (Brassicaceae), is a representative of specifier proteins, a group of kelch proteins involved in plant specialized metabolism. As components of the glucosinolate-myrosinase system of the Brassicaceae, specifier proteins determine the profile of bioactive products formed when plant tissue is disrupted and glucosinolates are hydrolyzed by myrosinases. Here, we describe the crystal structure of TaTFP at a resolution of 1.4 Å. TaTFP crystallized as homodimer. Each monomer forms a six-blade β-propeller with a wide "top" and a narrower "bottom" opening with distinct strand-connecting loops protruding far beyond the lower propeller surface. Molecular modeling and mutational analysis identified residues for glucosinolate aglucone and Fe(2+) cofactor binding within these loops. As the first experimentally determined structure of a plant kelch protein, the crystal structure of TaTFP not only enables more detailed mechanistic studies on glucosinolate breakdown product formation, but also provides a new basis for research on the diverse roles and mechanisms of other kelch proteins in plants.

  14. An Ensemble Method with Hybrid Features to Identify Extracellular Matrix Proteins

    PubMed Central

    Yang, Runtao; Zhang, Chengjin; Gao, Rui; Zhang, Lina

    2015-01-01

    The extracellular matrix (ECM) is a dynamic composite of secreted proteins that play important roles in numerous biological processes such as tissue morphogenesis, differentiation and homeostasis. Furthermore, various diseases are caused by the dysfunction of ECM proteins. Therefore, identifying these important ECM proteins may assist in understanding related biological processes and drug development. In view of the serious imbalance in the training dataset, a Random Forest-based ensemble method with hybrid features is developed in this paper to identify ECM proteins. Hybrid features are employed by incorporating sequence composition, physicochemical properties, evolutionary and structural information. The Information Gain Ratio and Incremental Feature Selection (IGR-IFS) methods are adopted to select the optimal features. Finally, the resulting predictor termed IECMP (Identify ECM Proteins) achieves an balanced accuracy of 86.4% using the 10-fold cross-validation on the training dataset, which is much higher than results obtained by other methods (ECMPRED: 71.0%, ECMPP: 77.8%). Moreover, when tested on a common independent dataset, our method also achieves significantly improved performance over ECMPP and ECMPRED. These results indicate that IECMP is an effective method for ECM protein prediction, which has a more balanced prediction capability for positive and negative samples. It is anticipated that the proposed method will provide significant information to fully decipher the molecular mechanisms of ECM-related biological processes and discover candidate drug targets. For public access, we develop a user-friendly web server for ECM protein identification that is freely accessible at http://iecmp.weka.cc. PMID:25680094

  15. Prediction of Protein Structural Class Based on Gapped-Dipeptides and a Recursive Feature Selection Approach.

    PubMed

    Liu, Taigang; Qin, Yufang; Wang, Yongjie; Wang, Chunhua

    2015-12-24

    The prior knowledge of protein structural class may offer useful clues on understanding its functionality as well as its tertiary structure. Though various significant efforts have been made to find a fast and effective computational approach to address this problem, it is still a challenging topic in the field of bioinformatics. The position-specific score matrix (PSSM) profile has been shown to provide a useful source of information for improving the prediction performance of protein structural class. However, this information has not been adequately explored. To this end, in this study, we present a feature extraction technique which is based on gapped-dipeptides composition computed directly from PSSM. Then, a careful feature selection technique is performed based on support vector machine-recursive feature elimination (SVM-RFE). These optimal features are selected to construct a final predictor. The results of jackknife tests on four working datasets show that our method obtains satisfactory prediction accuracies by extracting features solely based on PSSM and could serve as a very promising tool to predict protein structural class.

  16. Neuron membrane trafficking and protein kinases involved in autism and ADHD.

    PubMed

    Kitagishi, Yasuko; Minami, Akari; Nakanishi, Atsuko; Ogura, Yasunori; Matsuda, Satoru

    2015-01-30

    A brain-enriched multi-domain scaffolding protein, neurobeachin has been identified as a candidate gene for autism patients. Mutations in the synaptic adhesion protein cell adhesion molecule 1 (CADM1) are also associated with autism spectrum disorder, a neurodevelopmental disorder of uncertain molecular origin. Potential roles of neurobeachin and CADM1 have been suggested to a function of vesicle transport in endosomal trafficking. It seems that protein kinase B (AKT) and cyclic adenosine monophosphate (cAMP)-dependent protein kinase A (PKA) have key roles in the neuron membrane trafficking involved in the pathogenesis of autism. Attention deficit hyperactivity disorder (ADHD) is documented to dopaminergic insufficiencies, which is attributed to synaptic dysfunction of dopamine transporter (DAT). AKT is also essential for the DAT cell-surface redistribution. In the present paper, we summarize and discuss the importance of several protein kinases that regulate the membrane trafficking involved in autism and ADHD, suggesting new targets for therapeutic intervention.

  17. The presequence pathway is involved in protein sorting to the mitochondrial outer membrane

    PubMed Central

    Wenz, Lena-Sophie; Opaliński, Łukasz; Schuler, Max-Hinderk; Ellenrieder, Lars; Ieva, Raffaele; Böttinger, Lena; Qiu, Jian; van der Laan, Martin; Wiedemann, Nils; Guiard, Bernard; Pfanner, Nikolaus; Becker, Thomas

    2014-01-01

    The mitochondrial outer membrane contains integral α-helical and β-barrel proteins that are imported from the cytosol. The machineries importing β-barrel proteins have been identified, however, different views exist on the import of α-helical proteins. It has been reported that the biogenesis of Om45, the most abundant signal-anchored protein, does not depend on proteinaceous components, but involves direct insertion into the outer membrane. We show that import of Om45 occurs via the translocase of the outer membrane and the presequence translocase of the inner membrane. Assembly of Om45 in the outer membrane involves the MIM machinery. Om45 thus follows a new mitochondrial biogenesis pathway that uses elements of the presequence import pathway to direct a protein to the outer membrane. PMID:24781695

  18. Self protein-protein interactions are involved in TPPP/p25 mediated microtubule bundling

    PubMed Central

    DeBonis, Salvatore; Neumann, Emmanuelle; Skoufias, Dimitrios A.

    2015-01-01

    TPPP/p25 is a microtubule-associated protein, detected in protein inclusions associated with various neurodegenerative diseases. Deletion analysis data show that TPPP/p25 has two microtubule binding sites, both located in intrinsically disordered domains, one at the N-terminal and the other in the C-terminal domain. In copolymerization assays the full-length protein exhibits microtubule stimulation and bundling activity. In contrast, at the same ratio relative to tubulin, truncated forms of TPPP/p25 exhibit either lower or no microtubule stimulation and no bundling activity, suggesting a cooperative phenomenon which is enhanced by the presence of the two binding sites. The binding characteristics of the N- and C-terminally truncated proteins to taxol-stabilized microtubules are similar to the full-length protein. However, the C-terminally truncated TPPP/p25 shows a lower Bmax for microtubule binding, suggesting that it may bind to a site of tubulin that is masked in microtubules. Bimolecular fluorescent complementation assays in cells expressing combinations of various TPPP/p25 fragments, but not that of the central folded domain, resulted in the generation of a fluorescence signal colocalized with perinuclear microtubule bundles insensitive to microtubule inhibitors. The data suggest that the central folded domain of TPPP/p25 following binding to microtubules can drive s homotypic protein-protein interactions leading to bundled microtubules. PMID:26289831

  19. Predicting bacteriophage proteins located in host cell with feature selection technique.

    PubMed

    Ding, Hui; Liang, Zhi-Yong; Guo, Feng-Biao; Huang, Jian; Chen, Wei; Lin, Hao

    2016-04-01

    A bacteriophage is a virus that can infect a bacterium. The fate of an infected bacterium is determined by the bacteriophage proteins located in the host cell. Thus, reliably identifying bacteriophage proteins located in the host cell is extremely important to understand their functions and discover potential anti-bacterial drugs. Thus, in this paper, a computational method was developed to recognize bacteriophage proteins located in host cells based only on their amino acid sequences. The analysis of variance (ANOVA) combined with incremental feature selection (IFS) was proposed to optimize the feature set. Using a jackknife cross-validation, our method can discriminate between bacteriophage proteins located in a host cell and the bacteriophage proteins not located in a host cell with a maximum overall accuracy of 84.2%, and can further classify bacteriophage proteins located in host cell cytoplasm and in host cell membranes with a maximum overall accuracy of 92.4%. To enhance the value of the practical applications of the method, we built a web server called PHPred (〈http://lin.uestc.edu.cn/server/PHPred〉). We believe that the PHPred will become a powerful tool to study bacteriophage proteins located in host cells and to guide related drug discovery.

  20. Defense-related proteins involved in sugarcane responses to biotic stress.

    PubMed

    Souza, Thais P; Dias, Renata O; Silva-Filho, Marcio C

    2017-02-20

    Sugarcane is one of the most important agricultural crops in the world. However, pathogen infection and herbivore attack cause constant losses in yield. Plants respond to pathogen infection by inducing the expression of several protein types, such as glucanases, chitinases, thaumatins, peptidase inhibitors, defensins, catalases and glycoproteins. Proteins induced by pathogenesis are directly or indirectly involved in plant defense, leading to pathogen death or inducing other plant defense responses. Several of these proteins are induced in sugarcane by different pathogens or insects and have antifungal or insecticidal activity. In this review, defense-related proteins in sugarcane are described, with their putative mechanisms of action, pathogen targets and biotechnological perspectives.

  1. Sieve element occlusion (SEO) genes encode structural phloem proteins involved in wound sealing of the phloem.

    PubMed

    Ernst, Antonia M; Jekat, Stephan B; Zielonka, Sascia; Müller, Boje; Neumann, Ulla; Rüping, Boris; Twyman, Richard M; Krzyzanek, Vladislav; Prüfer, Dirk; Noll, Gundula A

    2012-07-10

    The sieve element occlusion (SEO) gene family originally was delimited to genes encoding structural components of forisomes, which are specialized crystalloid phloem proteins found solely in the Fabaceae. More recently, SEO genes discovered in various non-Fabaceae plants were proposed to encode the common phloem proteins (P-proteins) that plug sieve plates after wounding. We carried out a comprehensive characterization of two tobacco (Nicotiana tabacum) SEO genes (NtSEO). Reporter genes controlled by the NtSEO promoters were expressed specifically in immature sieve elements, and GFP-SEO fusion proteins formed parietal agglomerates in intact sieve elements as well as sieve plate plugs after wounding. NtSEO proteins with and without fluorescent protein tags formed agglomerates similar in structure to native P-protein bodies when transiently coexpressed in Nicotiana benthamiana, and the analysis of these protein complexes by electron microscopy revealed ultrastructural features resembling those of native P-proteins. NtSEO-RNA interference lines were essentially devoid of P-protein structures and lost photoassimilates more rapidly after injury than control plants, thus confirming the role of P-proteins in sieve tube sealing. We therefore provide direct evidence that SEO genes in tobacco encode P-protein subunits that affect translocation. We also found that peptides recently identified in fascicular phloem P-protein plugs from squash (Cucurbita maxima) represent cucurbit members of the SEO family. Our results therefore suggest a common evolutionary origin for P-proteins found in the sieve elements of all dicotyledonous plants and demonstrate the exceptional status of extrafascicular P-proteins in cucurbits.

  2. Distilling the essential features of a protein surface for improving protein-ligand docking, scoring, and virtual screening

    NASA Astrophysics Data System (ADS)

    Zavodszky, Maria I.; Sanschagrin, Paul C.; Kuhn, Leslie A.; Korde, Rajesh S.

    2002-12-01

    For the successful identification and docking of new ligands to a protein target by virtual screening, the essential features of the protein and ligand surfaces must be captured and distilled in an efficient representation. Since the running time for docking increases exponentially with the number of points representing the protein and each ligand candidate, it is important to place these points where the best interactions can be made between the protein and the ligand. This definition of favorable points of interaction can also guide protein structure-based ligand design, which typically focuses on which chemical groups provide the most energetically favorable contacts. In this paper, we present an alternative method of protein template and ligand interaction point design that identifies the most favorable points for making hydrophobic and hydrogen-bond interactions by using a knowledge base. The knowledge-based protein and ligand representations have been incorporated in version 2.0 of SLIDE and resulted in dockings closer to the crystal structure orientations when screening a set of 57 known thrombin and glutathione S-transferase (GST) ligands against the apo structures of these proteins. There was also improved scoring enrichment of the dockings, meaning better differentiation between the chemically diverse known ligands and a ˜15,000-molecule dataset of randomly-chosen small organic molecules. This approach for identifying the most important points of interaction between proteins and their ligands can equally well be used in other docking and design techniques. While much recent effort has focused on improving scoring functions for protein-ligand docking, our results indicate that improving the representation of the chemistry of proteins and their ligands is another avenue that can lead to significant improvements in the identification, docking, and scoring of ligands.

  3. Autophagy-linked FYVE protein (Alfy) promotes autophagic removal of misfolded proteins involved in amyotrophic lateral sclerosis (ALS).

    PubMed

    Han, Huihui; Wei, Wanyi; Duan, Weisong; Guo, Yansu; Li, Yi; Wang, Jie; Bi, Yue; Li, Chunyan

    2015-03-01

    Autophagy-linked FYVE (Alfy) is a protein implicated in the selective degradation of aggregated proteins. In our present study, we found that Alfy was recruited into the aggregated G93A-SOD1 in transgenic mice with amyotrophic lateral sclerosis (ALS). We demonstrated that Alfy overexpression could decrease the expression of mutant proteins via the autophagosome-lysosome pathway, and thereby, the toxicity of mutant proteins was reduced. The clearance of the mutant proteins in NSC34 cells was significantly inhibited in an Alfy knockdown cellular model. We therefore deduced that Alfy translocalization likely is involved in the pathogenesis of ALS. Alfy may be developed into a useful target for ALS therapy.

  4. FRG1P-mediated aggregation of proteins involved in pre-mRNA processing.

    PubMed

    van Koningsbruggen, Silvana; Straasheijm, Kirsten R; Sterrenburg, Ellen; de Graaf, Natascha; Dauwerse, Hans G; Frants, Rune R; van der Maarel, Silvère M

    2007-02-01

    FRG1 is considered a candidate gene for facioscapulohumeral muscular dystrophy (FSHD) based on its location at chromosome 4qter and its upregulation in FSHD muscle. The FRG1 protein (FRG1P) localizes to nucleoli, Cajal bodies (and speckles), and has been suggested to be a component of the human spliceosome but its exact function is unknown. Recently, transgenic mice overexpressing high levels of FRG1P in skeletal muscle were described to present with muscular dystrophy. Moreover, upregulation of FRG1P was demonstrated to correlate with missplicing of specific pre-mRNAs. In this study, we have combined colocalization studies with yeast two-hybrid screens to identify proteins that associate with FRG1P. We demonstrate that artificially induced nucleolar aggregates of VSV-FRG1P specifically sequester proteins involved in pre-mRNA processing. In addition, we have identified SMN, PABPN1, and FAM71B, a novel speckle and Cajal body protein, as binding partners of FRG1P. All these proteins are, or seem to be, involved in RNA biogenesis. Our data confirm the presence of FRG1P in protein complexes containing human spliceosomes and support a potential role of FRG1P in either splicing or another step in nuclear RNA biogenesis. Intriguingly, among FRG1P-associated proteins are SMN and PABPN1, both being involved in neuromuscular disorders, possibly through RNA biogenesis-related processes.

  5. Tick receptor for outer surface protein A from Ixodes ricinus — the first intrinsically disordered protein involved in vector-microbe recognition

    NASA Astrophysics Data System (ADS)

    Urbanowicz, Anna; Lewandowski, Dominik; Szpotkowski, Kamil; Figlerowicz, Marek

    2016-04-01

    The tick receptor for outer surface protein A (TROSPA) is the only identified factor involved in tick gut colonization by various Borrelia species. TROSPA is localized in the gut epithelium and can recognize and bind the outer surface bacterial protein OspA via an unknown mechanism. Based on earlier reports and our latest observations, we considered that TROSPA would be the first identified intrinsically disordered protein (IDP) involved in the interaction between a vector and a pathogenic microbe. To verify this hypothesis, we performed structural studies of a TROSPA mutant from Ixodes ricinus using both computational and experimental approaches. Irrespective of the method used, we observed that the secondary structure content of the TROSPA polypeptide chain is low. In addition, the collected SAXS data indicated that this protein is highly extended and exists in solution as a set of numerous conformers. These features are all commonly considered hallmarks of IDPs. Taking advantage of our SAXS data, we created structural models of TROSPA and proposed a putative mechanism for the TROSPA-OspA interaction. The disordered nature of TROSPA may explain the ability of a wide spectrum of Borrelia species to colonize the tick gut.

  6. Tick receptor for outer surface protein A from Ixodes ricinus — the first intrinsically disordered protein involved in vector-microbe recognition

    PubMed Central

    Urbanowicz, Anna; Lewandowski, Dominik; Szpotkowski, Kamil; Figlerowicz, Marek

    2016-01-01

    The tick receptor for outer surface protein A (TROSPA) is the only identified factor involved in tick gut colonization by various Borrelia species. TROSPA is localized in the gut epithelium and can recognize and bind the outer surface bacterial protein OspA via an unknown mechanism. Based on earlier reports and our latest observations, we considered that TROSPA would be the first identified intrinsically disordered protein (IDP) involved in the interaction between a vector and a pathogenic microbe. To verify this hypothesis, we performed structural studies of a TROSPA mutant from Ixodes ricinus using both computational and experimental approaches. Irrespective of the method used, we observed that the secondary structure content of the TROSPA polypeptide chain is low. In addition, the collected SAXS data indicated that this protein is highly extended and exists in solution as a set of numerous conformers. These features are all commonly considered hallmarks of IDPs. Taking advantage of our SAXS data, we created structural models of TROSPA and proposed a putative mechanism for the TROSPA-OspA interaction. The disordered nature of TROSPA may explain the ability of a wide spectrum of Borrelia species to colonize the tick gut. PMID:27112540

  7. The Deceptively Simple N170 Reflects Network Information Processing Mechanisms Involving Visual Feature Coding and Transfer Across Hemispheres

    PubMed Central

    Ince, Robin A. A.; Jaworska, Katarzyna; Gross, Joachim; Panzeri, Stefano; van Rijsbergen, Nicola J.; Rousselet, Guillaume A.; Schyns, Philippe G.

    2016-01-01

    A key to understanding visual cognition is to determine “where”, “when”, and “how” brain responses reflect the processing of the specific visual features that modulate categorization behavior—the “what”. The N170 is the earliest Event-Related Potential (ERP) that preferentially responds to faces. Here, we demonstrate that a paradigmatic shift is necessary to interpret the N170 as the product of an information processing network that dynamically codes and transfers face features across hemispheres, rather than as a local stimulus-driven event. Reverse-correlation methods coupled with information-theoretic analyses revealed that visibility of the eyes influences face detection behavior. The N170 initially reflects coding of the behaviorally relevant eye contralateral to the sensor, followed by a causal communication of the other eye from the other hemisphere. These findings demonstrate that the deceptively simple N170 ERP hides a complex network information processing mechanism involving initial coding and subsequent cross-hemispheric transfer of visual features. PMID:27550865

  8. Pressure-temperature folding landscape in proteins involved in neurodegenerative diseases and cancer.

    PubMed

    Cordeiro, Yraima; Foguel, Debora; Silva, Jerson L

    2013-12-15

    High hydrostatic pressure (HHP) is a valuable tool to study processes such as protein folding, protein hydration and protein-protein interactions. HHP is a nondestructive technique because it reversibly affects internal cavities excluded from the solvent present in the hydrophobic core of proteins. HHP allows the solvation of buried amino acid side chains, thus shifting the equilibrium towards states of the studied molecule or molecular ensemble that occupy smaller volumes. HHP has long been used to dissociate multimeric proteins and protein aggregates and allows investigation of intermediate folding states, some of which are formed by proteins involved in human degenerative diseases, such as spongiform encephalopathies and Parkinson's disease, as well as cancer. When coupled with nuclear magnetic resonance and spectroscopic methods such as infrared and fluorescence spectroscopy, HHP treatment facilitates the understanding of protein folding and misfolding processes; the latter is related to protein aggregation into amyloid or amorphous species. In this review, we will address how HHP provides information about intermediate folding states and the aggregation processes of p53, which is related to cancer, and prion proteins, transthyretin and α-synuclein, which are related to human degenerative diseases.

  9. Association of ADAM12-S protein with radiographic features of knee osteoarthritis and bone and cartilage markers.

    PubMed

    Kerna, I; Kisand, K; Laitinen, P; Tamm, A E; Kumm, J; Lintrop, M; Tamm, A O

    2012-02-01

    ADAM12 (A disintegrin and metalloprotease) is one of the candidate genes demonstrating susceptibility to osteoarthritis. The purpose of this study was to investigate the relationship between ADAM12-S protein and radiographic knee osteoarthritis (KOA) and its correlation to several bone and cartilage biomarkers. The ADAM12-S protein was measured in 276 subjects (60% women, aged 32-60 years), including 181 individuals with and 95 without radiographic KOA features. The radiographs were obtained from both tibiofemoral (TF) and patellofemoral (PF) joints. The serum levels of ADAM12-S protein were measured by DELFIA1/AutoDELFIA research kit. The ADAM12-S protein was found in detectable ranges in 43 subjects (16 men), without statistical difference between the two genders. In the whole group, the ADAM12-S was related to radiographic KOA grades in TF (P = 0.004) as well in PF joint (P = 0.003). We also found a correlation between ADAM12-S protein and osteophytes in TF and/or PF joints (P = 0.003). No correlations were found between serum levels of S-CTx-I (C-terminal cross-linked telopeptides of type I collagen) or S-PINP (type I procollagen N-terminal propeptide) and ADAM12-S. Similarly, in the whole group, the ADAM12-S protein was not correlated with U-CTx-II (urinary C-telopeptide fragments of type II collagen); however, in the female group, trend to positive correlation between the investigated biomarkers (P = 0.019) was observed. The ADAM12-S protein could be elevated in some KOA cases, and this elevation correlates with the grades of the disease, mostly owning to development of osteophytes. This finding suggests the possible involvement of the ADAM12-S protein in the pathogenesis of KOA.

  10. Involvement of a small GTP binding protein in HIV-1 release

    PubMed Central

    Audoly, Gilles; Popoff, Michel R; Gluschankof, Pablo

    2005-01-01

    Background There is evidence suggesting that actin binding to HIV-1 encoded proteins, or even actin dynamics themselves, might play a key role in virus budding and/or release from the infected cell. A crucial step in the reorganisation of the actin cytoskeleton is the engagement of various different GTP binding proteins. We have thus studied the involvement of GTP-binding proteins in the final steps of the HIV-1 viral replication cycle. Results Our results demonstrate that virus production is abolished when cellular GTP binding proteins involved in actin polymerisation are inhibited with specific toxins. Conclusion We propose a new HIV budding working model whereby Gag interactions with pre-existing endosomal cellular tracks as well as with a yet non identified element of the actin polymerisation pathway are required in order to allow HIV-1 to be released from the infected cell. PMID:16080789

  11. Proteomic Analysis of Differentially Expressed Proteins Involved in Peel Senescence in Harvested Mandarin Fruit.

    PubMed

    Li, Taotao; Zhang, Jingying; Zhu, Hong; Qu, Hongxia; You, Shulin; Duan, Xuewu; Jiang, Yueming

    2016-01-01

    Mandarin (Citrus reticulata), a non-climacteric fruit, is an economically important fruit worldwide. The mechanism underlying senescence of non-climacteric fruit is poorly understood. In this study, a gel-based proteomic study followed by LC-ESI-MS/MS analysis was carried out to investigate the proteomic changes involved in peel senescence in harvested mandarin "Shatangju" fruit stored for 18 days. Over the course of the storage period, the fruit gradually senesced, accompanied by a decreased respiration rate and increased chlorophyll degradation and disruption of membrane integrity. Sixty-three proteins spots that showed significant differences in abundance were identified. The up-regulated proteins were mainly associated with cell wall degradation, lipid degradation, protein degradation, senescence-related transcription factors, and transcription-related proteins. In contrast, most proteins associated with ATP synthesis and scavenging of reactive oxygen species were significantly down-regulated during peel senescence. Three thioredoxin proteins and three Ca(2+) signaling-related proteins were significantly up-regulated during peel senescence. It is suggested that mandarin peel senescence is associated with energy supply efficiency, decreased antioxidant capability, and increased protein and lipid degradation. In addition, activation of Ca(2+) signaling and transcription factors might be involved in cell wall degradation and primary or secondary metabolism.

  12. Proteomic Analysis of Differentially Expressed Proteins Involved in Peel Senescence in Harvested Mandarin Fruit

    PubMed Central

    Li, Taotao; Zhang, Jingying; Zhu, Hong; Qu, Hongxia; You, Shulin; Duan, Xuewu; Jiang, Yueming

    2016-01-01

    Mandarin (Citrus reticulata), a non-climacteric fruit, is an economically important fruit worldwide. The mechanism underlying senescence of non-climacteric fruit is poorly understood. In this study, a gel-based proteomic study followed by LC-ESI-MS/MS analysis was carried out to investigate the proteomic changes involved in peel senescence in harvested mandarin “Shatangju” fruit stored for 18 days. Over the course of the storage period, the fruit gradually senesced, accompanied by a decreased respiration rate and increased chlorophyll degradation and disruption of membrane integrity. Sixty-three proteins spots that showed significant differences in abundance were identified. The up-regulated proteins were mainly associated with cell wall degradation, lipid degradation, protein degradation, senescence-related transcription factors, and transcription-related proteins. In contrast, most proteins associated with ATP synthesis and scavenging of reactive oxygen species were significantly down-regulated during peel senescence. Three thioredoxin proteins and three Ca2+ signaling-related proteins were significantly up-regulated during peel senescence. It is suggested that mandarin peel senescence is associated with energy supply efficiency, decreased antioxidant capability, and increased protein and lipid degradation. In addition, activation of Ca2+ signaling and transcription factors might be involved in cell wall degradation and primary or secondary metabolism. PMID:27303420

  13. Proteins involved in motility and sperm-egg interaction evolve more rapidly in mouse spermatozoa.

    PubMed

    Vicens, Alberto; Lüke, Lena; Roldan, Eduardo R S

    2014-01-01

    Proteomic studies of spermatozoa have identified a large catalog of integral sperm proteins. Rapid evolution of these proteins may underlie adaptive changes of sperm traits involved in different events leading to fertilization, although the selective forces underlying such rapid evolution are not well understood. A variety of selective forces may differentially affect several steps ending in fertilization, thus resulting in a compartmentalized adaptation of sperm proteins. Here we analyzed the evolution of genes associated to various events in the sperm's life, from sperm formation to sperm-egg interaction. Evolutionary analyses were performed on gene sequences from 17 mouse strains whose genomes have been sequenced. Four of these are derived from wild Mus musculus, M. domesticus, M. castaneus and M. spretus. We found a higher proportion of genes exhibiting a signature of positive selection among those related to sperm motility and sperm-egg interaction. Furthermore, sperm proteins involved in sperm-egg interaction exhibited accelerated evolution in comparison to those involved in other events. Thus, we identified a large set of candidate proteins for future comparative analyses of genotype-phenotype associations in spermatozoa of species subjected to different sexual selection pressures. Adaptive evolution of proteins involved in motility could be driven by sperm competition, since this selective force is known to increase the proportion of motile sperm and their swimming velocity. On the other hand, sperm proteins involved in gamete interaction could be coevolving with their egg partners through episodes of sexual selection or sexual conflict resulting in species-specific sperm-egg interactions and barriers preventing interspecies fertilization.

  14. Protein single-model quality assessment by feature-based probability density functions.

    PubMed

    Cao, Renzhi; Cheng, Jianlin

    2016-04-04

    Protein quality assessment (QA) has played an important role in protein structure prediction. We developed a novel single-model quality assessment method-Qprob. Qprob calculates the absolute error for each protein feature value against the true quality scores (i.e. GDT-TS scores) of protein structural models, and uses them to estimate its probability density distribution for quality assessment. Qprob has been blindly tested on the 11th Critical Assessment of Techniques for Protein Structure Prediction (CASP11) as MULTICOM-NOVEL server. The official CASP result shows that Qprob ranks as one of the top single-model QA methods. In addition, Qprob makes contributions to our protein tertiary structure predictor MULTICOM, which is officially ranked 3rd out of 143 predictors. The good performance shows that Qprob is good at assessing the quality of models of hard targets. These results demonstrate that this new probability density distribution based method is effective for protein single-model quality assessment and is useful for protein structure prediction. The webserver of Qprob is available at: http://calla.rnet.missouri.edu/qprob/. The software is now freely available in the web server of Qprob.

  15. Elucidating Protein Involvement in the Stabilization of the Biogenic Silver Nanoparticles

    NASA Astrophysics Data System (ADS)

    Ballottin, Daniela; Fulaz, Stephanie; Souza, Michele L.; Corio, Paola; Rodrigues, Alexandre G.; Souza, Ana O.; Gaspari, Priscyla M.; Gomes, Alexandre F.; Gozzo, Fábio; Tasic, Ljubica

    2016-06-01

    Silver nanoparticles (AgNPs) have been broadly used as antibacterial and antiviral agents. Further, interests for green AgNP synthesis have increased in recent years and several results for AgNP biological synthesis have been reported using bacteria, fungi and plant extracts. The understanding of the role and nature of fungal proteins, their interaction with AgNPs and the subsequent stabilization of nanosilver is yet to be deeply investigated. Therefore, in an attempt to better understand biogenic AgNP stabilization with the extracellular fungal proteins and to describe these supramolecular interactions between proteins and silver nanoparticles, AgNPs, produced extracellularly by Aspergillus tubingensis—isolated as an endophytic fungus from Rizophora mangle—were characterized in order to study their physical characteristics, identify the involved proteins, and shed light into the interactions among protein-NPs by several techniques. AgNPs of around 35 nm in diameter as measured by TEM and a positive zeta potential of +8.48 mV were obtained. These AgNPs exhibited a surface plasmon resonance (SPR) band at 440 nm, indicating the nanoparticles formation, and another band at 280 nm, attributed to the electronic excitations in tryptophan, tyrosine, and/or phenylalanine residues in fungal proteins. Fungal proteins were covalently bounded to the AgNPs, mainly through S-Ag bonds due to cysteine residues (HS-) and with few N-Ag bonds from H2N- groups, as verified by Raman spectroscopy. Observed supramolecular interactions also occur by electrostatic and other protein-protein interactions. Furthermore, proteins that remain free on AgNP surface may perform hydrogen bonds with other proteins or water increasing thus the capping layer around the AgNPs and consequently expanding the hydrodynamic diameter of the particles (~264 nm, measured by DLS). FTIR results enabled us to state that proteins adsorbed to the AgNPs did not suffer relevant secondary structure alteration upon

  16. Structural Interface Forms and Their Involvement in Stabilization of Multidomain Proteins or Protein Complexes

    PubMed Central

    Dygut, Jacek; Kalinowska, Barbara; Banach, Mateusz; Piwowar, Monika; Konieczny, Leszek; Roterman, Irena

    2016-01-01

    The presented analysis concerns the inter-domain and inter-protein interface in protein complexes. We propose extending the traditional understanding of the protein domain as a function of local compactness with an additional criterion which refers to the presence of a well-defined hydrophobic core. Interface areas in selected homodimers vary with respect to their contribution to share as well as individual (domain-specific) hydrophobic cores. The basic definition of a protein domain, i.e., a structural unit characterized by tighter packing than its immediate environment, is extended in order to acknowledge the role of a structured hydrophobic core, which includes the interface area. The hydrophobic properties of interfaces vary depending on the status of interacting domains—In this context we can distinguish: (1) Shared hydrophobic cores (spanning the whole dimer); (2) Individual hydrophobic cores present in each monomer irrespective of whether the dimer contains a shared core. Analysis of interfaces in dystrophin and utrophin indicates the presence of an additional quasi-domain with a prominent hydrophobic core, consisting of fragments contributed by both monomers. In addition, we have also attempted to determine the relationship between the type of interface (as categorized above) and the biological function of each complex. This analysis is entirely based on the fuzzy oil drop model. PMID:27763556

  17. Structural Interface Forms and Their Involvement in Stabilization of Multidomain Proteins or Protein Complexes.

    PubMed

    Dygut, Jacek; Kalinowska, Barbara; Banach, Mateusz; Piwowar, Monika; Konieczny, Leszek; Roterman, Irena

    2016-10-18

    The presented analysis concerns the inter-domain and inter-protein interface in protein complexes. We propose extending the traditional understanding of the protein domain as a function of local compactness with an additional criterion which refers to the presence of a well-defined hydrophobic core. Interface areas in selected homodimers vary with respect to their contribution to share as well as individual (domain-specific) hydrophobic cores. The basic definition of a protein domain, i.e., a structural unit characterized by tighter packing than its immediate environment, is extended in order to acknowledge the role of a structured hydrophobic core, which includes the interface area. The hydrophobic properties of interfaces vary depending on the status of interacting domains-In this context we can distinguish: (1) Shared hydrophobic cores (spanning the whole dimer); (2) Individual hydrophobic cores present in each monomer irrespective of whether the dimer contains a shared core. Analysis of interfaces in dystrophin and utrophin indicates the presence of an additional quasi-domain with a prominent hydrophobic core, consisting of fragments contributed by both monomers. In addition, we have also attempted to determine the relationship between the type of interface (as categorized above) and the biological function of each complex. This analysis is entirely based on the fuzzy oil drop model.

  18. Computer-based comparison of structural features of envelope protein of Alkhurma hemorrhagic fever virus with the homologous proteins of two closest viruses.

    PubMed

    Mohabatkar, Hassan

    2011-06-01

    The aim of this study was prediction of epitopes and medically important structural properties of protein E of Alkhurma hemorrhagic fever virus (AHFV) and comparing these features with two closely relates viruses, i.e. Kyasanur Forest disease virus (KFDV) and Tick-borne encephalitis virus (TBEV) by bioinformatics tools. Prediction of evolutionary distance, localization, sequence of signal peptides, C, N O glycosylation sites, transmembrane helices (TMHs), cysteine bond positions and B cell and T cell epitopes of E proteins were performed. 2D-MH, Virus-PLoc, Signal-CF, EnsembleGly, MemBrain, DiANNA, BCPREDS and MHCPred servers were applied for the prediction. According to the results, the evolutionary distance of E protein of AHFV and two other viruses was almost equal. In all three proteins of study, residues 1-35 were predicted as signal sequences and one asparagine was predicted to be glycosylated. Results of prediction of transmembrane helices showed one TMH at position 444-467 and the other one at position 476-490. Twelve cysteines were potentially involved to form six disulfide bridges in the proteins. Four parts were predicted as B cell epitopes in E protein of AHFV. One epitope was conserved between three proteins of study. The only conserved major histocompatibility complex (MHC) binding epitope between three viruses was for DRB0401 allele. As there are not much experimental data available about AHFV, computer-aided study and comparison of E protein of this virus with two closely related flaviviruses can help in better understanding of medical properties of the virus.

  19. Polymorphisms and features of cytomegalovirus UL144 and UL146 in congenitally infected neonates with hepatic involvement.

    PubMed

    Guo, Gangqiang; Zhang, Liang; Ye, Sisi; Hu, Yingying; Li, Baoqing; Sun, Xiangwei; Mao, Chenchen; Xu, Jianfeng; Chen, Yiping; Zhang, Lifang; Xue, Xiangyang

    2017-01-01

    Human cytomegalovirus is a significant agent of hepatic involvement in neonates. In this study, we investigated the polymorphisms and features of the viral genes UL144 and UL146 as well as their significance to congenital hepatic involvement. In 79 neonates with congenital cytomegalovirus infection and hepatic involvement, full length UL144 and UL146 were successfully amplified in 73.42% and 60.76% of cases, respectively. Sequencing indicated that both genes were hypervariable. Notably, UL144 genotype B was highly associated with aspartate aminotransferase (P = 0.028) and lactate dehydrogenase (P = 0.046). Similarly, UL146 genotype G1 and G13 were significantly associated with CMV IgM (P = 0.026), CMV IgG (P = 0.034), alanine aminotransferase (P = 0.019), and aspartate aminotransferase (P = 0.032). In conclusion, dominant UL144 (genotype B) and UL146 (genotype G1 and G13) genotypes are associated with elevated levels of enzymes and CMV IgM and IgG of cytomegalovirus infection.

  20. Polymorphisms and features of cytomegalovirus UL144 and UL146 in congenitally infected neonates with hepatic involvement

    PubMed Central

    Ye, Sisi; Hu, Yingying; Li, Baoqing; Sun, Xiangwei; Mao, Chenchen; Xu, Jianfeng; Chen, Yiping; Zhang, Lifang; Xue, Xiangyang

    2017-01-01

    Human cytomegalovirus is a significant agent of hepatic involvement in neonates. In this study, we investigated the polymorphisms and features of the viral genes UL144 and UL146 as well as their significance to congenital hepatic involvement. In 79 neonates with congenital cytomegalovirus infection and hepatic involvement, full length UL144 and UL146 were successfully amplified in 73.42% and 60.76% of cases, respectively. Sequencing indicated that both genes were hypervariable. Notably, UL144 genotype B was highly associated with aspartate aminotransferase (P = 0.028) and lactate dehydrogenase (P = 0.046). Similarly, UL146 genotype G1 and G13 were significantly associated with CMV IgM (P = 0.026), CMV IgG (P = 0.034), alanine aminotransferase (P = 0.019), and aspartate aminotransferase (P = 0.032). In conclusion, dominant UL144 (genotype B) and UL146 (genotype G1 and G13) genotypes are associated with elevated levels of enzymes and CMV IgM and IgG of cytomegalovirus infection. PMID:28222150

  1. A human skeletal muscle interactome centered on proteins involved in muscular dystrophies: LGMD interactome

    PubMed Central

    2013-01-01

    Background The complexity of the skeletal muscle and the identification of numerous human disease-causing mutations in its constitutive proteins make it an interesting tissue for proteomic studies aimed at understanding functional relationships of interacting proteins in both health and diseases. Method We undertook a large-scale study using two-hybrid screens and a human skeletal-muscle cDNA library to establish a proteome-scale map of protein-protein interactions centered on proteins involved in limb-girdle muscular dystrophies (LGMD). LGMD is a group of more than 20 different neuromuscular disorders that principally affect the proximal pelvic and shoulder girdle muscles. Results and conclusion The interaction network we unraveled incorporates 1018 proteins connected by 1492 direct binary interactions and includes 1420 novel protein-protein interactions. Computational, experimental and literature-based analyses were performed to assess the overall quality of this network. Interestingly, LGMD proteins were shown to be highly interconnected, in particular indirectly through sarcomeric proteins. In-depth mining of the LGMD-centered interactome identified new candidate genes for orphan LGMDs and other neuromuscular disorders. The data also suggest the existence of functional links between LGMD2B/dysferlin and gene regulation, between LGMD2C/γ-sarcoglycan and energy control and between LGMD2G/telethonin and maintenance of genome integrity. This dataset represents a valuable resource for future functional investigations. PMID:23414517

  2. Structural Features of Ion Transport and Allosteric Regulation in Sodium-Calcium Exchanger (NCX) Proteins

    PubMed Central

    Giladi, Moshe; Tal, Inbal; Khananshvili, Daniel

    2016-01-01

    Na+/Ca2+ exchanger (NCX) proteins extrude Ca2+ from the cell to maintain cellular homeostasis. Since NCX proteins contribute to numerous physiological and pathophysiological events, their pharmacological targeting has been desired for a long time. This intervention remains challenging owing to our poor understanding of the underlying structure-dynamic mechanisms. Recent structural studies have shed light on the structure-function relationships underlying the ion-transport and allosteric regulation of NCX. The crystal structure of an archaeal NCX (NCX_Mj) along with molecular dynamics simulations and ion flux analyses, have assigned the ion binding sites for 3Na+ and 1Ca2+, which are being transported in separate steps. In contrast with NCX_Mj, eukaryotic NCXs contain the regulatory Ca2+-binding domains, CBD1 and CBD2, which affect the membrane embedded ion-transport domains over a distance of ~80 Å. The Ca2+-dependent regulation is ortholog, isoform, and splice-variant dependent to meet physiological requirements, exhibiting either a positive, negative, or no response to regulatory Ca2+. The crystal structures of the two-domain (CBD12) tandem have revealed a common mechanism involving a Ca2+-driven tethering of CBDs in diverse NCX variants. However, dissociation kinetics of occluded Ca2+ (entrapped at the two-domain interface) depends on the alternative-splicing segment (at CBD2), thereby representing splicing-dependent dynamic coupling of CBDs. The HDX-MS, SAXS, NMR, FRET, equilibrium 45Ca2+ binding and stopped-flow techniques provided insights into the dynamic mechanisms of CBDs. Ca2+ binding to CBD1 results in a population shift, where more constraint conformational states become highly populated without global conformational changes in the alignment of CBDs. This mechanism is common among NCXs. Recent HDX-MS studies have demonstrated that the apo CBD1 and CBD2 are stabilized by interacting with each other, while Ca2+ binding to CBD1 rigidifies local backbone

  3. Predicting Essential Genes and Proteins Based on Machine Learning and Network Topological Features: A Comprehensive Review

    PubMed Central

    Zhang, Xue; Acencio, Marcio Luis; Lemke, Ney

    2016-01-01

    Essential proteins/genes are indispensable to the survival or reproduction of an organism, and the deletion of such essential proteins will result in lethality or infertility. The identification of essential genes is very important not only for understanding the minimal requirements for survival of an organism, but also for finding human disease genes and new drug targets. Experimental methods for identifying essential genes are costly, time-consuming, and laborious. With the accumulation of sequenced genomes data and high-throughput experimental data, many computational methods for identifying essential proteins are proposed, which are useful complements to experimental methods. In this review, we show the state-of-the-art methods for identifying essential genes and proteins based on machine learning and network topological features, point out the progress and limitations of current methods, and discuss the challenges and directions for further research. PMID:27014079

  4. Multiple proteins of White spot syndrome virus involved in recognition of beta-integrin.

    PubMed

    Zhang, Jing-Yan; Liu, Qing-Hui; Huang, Jie

    2014-06-01

    The recognition and attachment of virus to its host cell surface is a critical step for viral infection. Recent research revealed that beta-integrin was involved in White spot syndrome virus (WSSV) infection. In this study, the interaction of beta-integrin with structure proteins of WSSV and motifs involved in WSSV infection was examined. The results showed that envelope proteins VP26, VP31, VP37, VP90 and nucleocapsid protein VP136 interacted with LvInt. RGD-, YGL- and LDV-related peptide functioned as motifs of WSSV proteins binding with beta-integrin. The beta-integrin ligand of RGDT had better blocking effect compared with that of YGL- and LDV-related peptides. In vivo assay indicated that RGD-, LDV- and YGL-related peptides could partially block WSSV infection. These data collectively indicate that multiple proteins were involved in recognition of beta-integrin. Identification of proteins in WSSV that are associated with beta-integrin will assist development of new agents for effective control of the white spot syndrome.

  5. Characterization of protein and carbohydrate mid-IR spectral features in crop residues

    NASA Astrophysics Data System (ADS)

    Xin, Hangshu; Zhang, Yonggen; Wang, Mingjun; Li, Zhongyu; Wang, Zhibo; Yu, Peiqiang

    2014-08-01

    To the best of our knowledge, a few studies have been conducted on inherent structure spectral traits related to biopolymers of crop residues. The objective of this study was to characterize protein and carbohydrate structure spectral features of three field crop residues (rice straw, wheat straw and millet straw) in comparison with two crop vines (peanut vine and pea vine) by using Fourier transform infrared spectroscopy (FTIR) technique with attenuated total reflectance (ATR). Also, multivariate analyses were performed on spectral data sets within the regions mainly related to protein and carbohydrate in this study. The results showed that spectral differences existed in mid-IR peak intensities that are mainly related to protein and carbohydrate among these crop residue samples. With regard to protein spectral profile, peanut vine showed the greatest mid-IR band intensities that are related to protein amide and protein secondary structures, followed by pea vine and the rest three field crop straws. The crop vines had 48-134% higher spectral band intensity than the grain straws in spectral features associated with protein. Similar trends were also found in the bands that are mainly related to structural carbohydrates (such as cellulosic compounds). However, the field crop residues had higher peak intensity in total carbohydrates region than the crop vines. Furthermore, spectral ratios varied among the residue samples, indicating that these five crop residues had different internal structural conformation. However, multivariate spectral analyses showed that structural similarities still exhibited among crop residues in the regions associated with protein biopolymers and carbohydrate. Further study is needed to find out whether there is any relationship between spectroscopic information and nutrition supply in various kinds of crop residue when fed to animals.

  6. Reduction of Cellular Expression Levels Is a Common Feature of Functionally Affected Pendrin (SLC26A4) Protein Variants

    PubMed Central

    de Moraes, Vanessa C S; Bernardinelli, Emanuele; Zocal, Nathalia; Fernandez, Jhonathan A; Nofziger, Charity; Castilho, Arthur M; Sartorato, Edi L; Paulmichl, Markus; Dossena, Silvia

    2016-01-01

    Sequence alterations in the pendrin gene (SLC26A4) leading to functionally affected protein variants are frequently involved in the pathogenesis of syndromic and nonsyndromic deafness. Considering the high number of SLC26A4 sequence alterations reported to date, discriminating between functionally affected and unaffected pendrin protein variants is essential in contributing to determine the genetic cause of deafness in a given patient. In addition, identifying molecular features common to the functionally affected protein variants can be extremely useful to design future molecule-directed therapeutic approaches. Here we show the functional and molecular characterization of six previously uncharacterized pendrin protein variants found in a cohort of 58 Brazilian deaf patients. Two variants (p.T193I and p.L445W) were undetectable in the plasma membrane, completely retained in the endoplasmic reticulum and showed no transport function; four (p.P142L, p.G149R, p.C282Y and p.Q413R) showed reduced function and significant, although heterogeneous, expression levels in the plasma membrane. Importantly, total expression levels of all of the functionally affected protein variants were significantly reduced with respect to the wild-type and a fully functional variant (p.R776C), regardless of their subcellular localization. Interestingly, reduction of expression may also reduce the transport activity of variants with an intrinsic gain of function (p.Q413R). As reduction of overall cellular abundance was identified as a common molecular feature of pendrin variants with affected function, the identification of strategies to prevent reduction in expression levels may represent a crucial step of potential future therapeutic interventions aimed at restoring the transport activity of dysfunctional pendrin variants. PMID:26752218

  7. Not all mitochondrial carrier proteins support permeability transition pore formation: no involvement of uncoupling protein 1.

    PubMed

    Crichton, Paul G; Parker, Nadeene; Vidal-Puig, Antonio J; Brand, Martin D

    2009-12-15

    The mPTP (mitochondrial permeability transition pore) is a non-specific channel that is formed in the mitochondrial inner membrane in response to several stimuli, including elevated levels of matrix calcium. The pore is proposed to be composed of the ANT (adenine nucleotide translocase), voltage-dependent anion channel and cyclophilin D. Knockout studies, however, have demonstrated that ANT is not essential for permeability transition, which has led to the proposal that other members of the mitochondrial carrier protein family may be able to play a similar function to ANT in pore formation. To investigate this possibility, we have studied the permeability transition properties of BAT (brown adipose tissue) mitochondria in which levels of the mitochondrial carrier protein, UCP1 (uncoupling protein 1), can exceed those of ANT. Using an improved spectroscopic assay, we have quantified mPTP formation in de-energized mitochondria from wild-type and Ucp1KO (Ucp1-knockout) mice and assessed the dependence of pore formation on UCP1. When correctly normalized for differences in mitochondrial morphology, we find that calcium-induced mPTP activity is the same in both types of mitochondria, with similar sensitivity to GDP (approximately 50% inhibited), although the portion sensitive to cyclosporin A is higher in mitochondria lacking UCP1 (approximately 80% inhibited, compared with approximately 60% in mitochondria containing UCP1). We conclude that UCP1 is not a component of the cyclosporin A-sensitive mPTP in BAT and that playing a role in mPTP formation is not a general characteristic of the mitochondrial carrier protein family but is, more likely, restricted to specific members including ANT.

  8. Protein-protein interactions involved in the recognition of p27 by E3 ubiquitin ligase.

    PubMed Central

    Xu, Kui; Belunis, Charles; Chu, Wei; Weber, David; Podlaski, Frank; Huang, Kuo-Sen; Reed, Steven I; Vassilev, Lyubomir T

    2003-01-01

    The p27(Kip1) protein is a potent cyclin-dependent kinase inhibitor, the level of which is decreased in many common human cancers as a result of enhanced ubiquitin-dependent degradation. The multiprotein complex SCF(Skp2) has been identified as the ubiquitin ligase that targets p27, but the functional interactions within this complex are not well understood. One component, the F-box protein Skp2, binds p27 when the latter is phosphorylated on Thr(187), thus providing substrate specificity for the ligase. Recently, we and others have shown that the small cell cycle regulatory protein Cks1 plays a critical role in p27 ubiquitination by increasing the binding affinity of Skp2 for p27. Here we report the development of a homogeneous time-resolved fluorescence assay that allows the quantification of the molecular interactions between human recombinant Skp2, Cks1 and a p27-derived peptide phosphorylated on Thr(187). Using this assay, we have determined the dissociation constant of the Skp2-Cks1 complex (K(d) 140 +/- 14 nM) and have shown that Skp2 binds phosphorylated p27 peptide with high affinity only in the presence of Cks1 (K(d) 37 +/- 2 nM). Cks1 does not bind directly to the p27 phosphopeptide or to Skp1, which confirms its suggested role as an allosteric effector of Skp2. PMID:12529174

  9. HOPS: a novel cAMP-dependent shuttling protein involved in protein synthesis regulation.

    PubMed

    Della Fazia, Maria Agnese; Castelli, Marilena; Bartoli, Daniela; Pieroni, Stefania; Pettirossi, Valentina; Piobbico, Danilo; Viola-Magni, Mariapia; Servillo, Giuseppe

    2005-07-15

    The liver has the ability to autonomously regulate growth and mass. Following partial hepatectomy, hormones, growth factors, cytokines and their coupled signal transduction pathways have been implicated in hepatocyte proliferation. To understand the mechanisms responsible for the proliferative response, we studied liver regeneration by characterization of novel genes that are activated in residual hepatocytes. A regenerating liver cDNA library screening was performed with cDNA-subtracted probes derived from regenerating and normal liver. Here, we describe the biology of Hops (for hepatocyte odd protein shuttling). HOPS is a novel shuttling protein that contains an ubiquitin-like domain, a putative NES and a proline-rich region. HOPS is rapidly exported from the nucleus and is overexpressed during liver regeneration. Evidence shows that cAMP governs HOPS export in hepatocytes of normal and regenerating liver and is mediated via CRM-1. We demonstrate that HOPS binds to elongation factor eEF-1A and interferes in protein synthesis. HOPS overexpression in H-35-hepatoma and 3T3-NIH cells strongly reduces proliferation.

  10. RNA-binding proteins involved in post-transcriptional regulation in bacteria

    PubMed Central

    Van Assche, Elke; Van Puyvelde, Sandra; Vanderleyden, Jos; Steenackers, Hans P.

    2015-01-01

    Post-transcriptional regulation is a very important mechanism to control gene expression in changing environments. In the past decade, a lot of interest has been directed toward the role of small RNAs (sRNAs) in bacterial post-transcriptional regulation. However, sRNAs are not the only molecules controlling gene expression at this level, RNA-binding proteins (RBPs) play an important role as well. CsrA and Hfq are the two best studied bacterial proteins of this type, but recently, additional proteins involved in post-transcriptional control have been identified. This review focuses on the general working mechanisms of post-transcriptionally active RBPs, which include (i) adaptation of the susceptibility of mRNAs and sRNAs to RNases, (ii) modulating the accessibility of the ribosome binding site of mRNAs, (iii) recruiting and assisting in the interaction of mRNAs with other molecules and (iv) regulating transcription terminator/antiterminator formation, and gives an overview of both the well-studied and the newly identified proteins that are involved in post-transcriptional regulatory processes. Additionally, the post-transcriptional mechanisms by which the expression or the activity of these proteins is regulated, are described. For many of the newly identified proteins, however, mechanistic questions remain. Most likely, more post-transcriptionally active proteins will be identified in the future. PMID:25784899

  11. The TSG101 protein binds to connexins and is involved in connexin degradation

    SciTech Connect

    Auth, Tanja Schlueter, Sharazad; Urschel, Stephanie; Kussmann, Petra; Sonntag, Stephan; Hoeher, Thorsten; Kreuzberg, Maria M.; Dobrowolski, Radoslaw; Willecke, Klaus

    2009-04-01

    Gap junctions mediate electrical and metabolic communication between cells in almost all tissues and are proposed to play important roles in cellular growth control, differentiation and embryonic development. Gap junctional communication and channel assembly were suggested to be regulated by interaction of connexins with different proteins including kinases and phosphatases. Here, we identified the tumor susceptibility gene 101 (TSG101) protein to bind to the carboxyterminal tail of connexin45 in a yeast two-hybrid protein interaction screen. Glutathione S-transferase pull down experiments and immunoprecipitation revealed that not only connexin45 but also connexin30.2, -36, and -43 carboxyterminal regions were associated with TSG101 protein in pull down analyses and that connexin31, -43 and -45 co-precipitate with endogenous TSG101 protein in lysates from HM1 embryonic stem cells. TSG101 has been shown to be involved in cell cycle control, transcriptional regulation and turnover of endocytosed proteins. Thus, we decided to study the functional role of this interaction. SiRNA mediated knock down of TSG101 in HM1 embryonic stem cells led to increased levels of connexin43 and -45, prolonged half life of these connexins and increased transfer of microinjected Lucifer yellow. Our results suggest that TSG101 is involved in the degradation of connexins via interaction with connexin proteins.

  12. Characterization of a DNA binding protein of bacteriophage PRD1 involved in DNA replication.

    PubMed Central

    Pakula, T M; Caldentey, J; Serrano, M; Gutierrez, C; Hermoso, J M; Salas, M; Bamford, D H

    1990-01-01

    Escherichia coli phage PRD1 protein P12, involved in PRD1 DNA replication in vivo, has been highly purified from E. coli cells harbouring a gene XII-containing plasmid. Protein P12 binds to single-stranded DNA as shown by gel retardation assays and nuclease protection experiments. Binding of protein P12 to single-stranded DNA increases about 14% the contour length of the DNA as revealed by electron microscopy. Binding to single-stranded DNA seems to be cooperative, and it is not sequence specific. Protein P12 also binds to double-stranded DNA although with an affinity 10 times lower than to single-stranded DNA. Using the in vitro phage phi 29 DNA replication system, it is shown that protein P12 stimulates the overall phi 29 DNA replication. Images PMID:2251117

  13. Phylogenomic analysis of the Chlamydomonas genome unmasks proteins potentially involved in photosynthetic function and regulation

    PubMed Central

    Karpowicz, Steven J.; Heinnickel, Mark; Dewez, David; Hamel, Blaise; Dent, Rachel; Niyogi, Krishna K.; Johnson, Xenie; Alric, Jean; Wollman, Francis-André; Li, Huiying; Merchant, Sabeeha S.

    2010-01-01

    Chlamydomonas reinhardtii, a unicellular green alga, has been exploited as a reference organism for identifying proteins and activities associated with the photosynthetic apparatus and the functioning of chloroplasts. Recently, the full genome sequence of Chlamydomonas was generated and a set of gene models, representing all genes on the genome, was developed. Using these gene models, and gene models developed for the genomes of other organisms, a phylogenomic, comparative analysis was performed to identify proteins encoded on the Chlamydomonas genome which were likely involved in chloroplast functions (or specifically associated with the green algal lineage); this set of proteins has been designated the GreenCut. Further analyses of those GreenCut proteins with uncharacterized functions and the generation of mutant strains aberrant for these proteins are beginning to unmask new layers of functionality/regulation that are integrated into the workings of the photosynthetic apparatus. PMID:20490922

  14. Application of intelligent techniques for classification of bacteria using protein sequence-derived features.

    PubMed

    Banerjee, Amit Kumar; Ravi, Vadlamani; Murty, U S N; Sengupta, Neelava; Karuna, Batepatti

    2013-07-01

    Standard molecular experimental methodologies and mathematical procedures often fail to answer many phylogeny and classification related issues. Modern artificial intelligent-based techniques, such as radial basis function, genetic algorithm, artificial neural network, and support vector machines are of ample potential in this regard. Reliance on a large number of essential parameters will aid in enhanced robustness, reliability, and better accuracy as opposed to single molecular parameter. This study was conducted with dataset of computed protein physicochemical properties belonging to 20 different bacterial genera. A total of 57 sequential and structural parameters derived from protein sequences were considered for the initial classification. Feature selection based techniques were employed to find out the most important features influencing the dataset. Various amino acids, hydrophobicity, relative sulfur percentage, and codon number were selected as important parameters during the study. Comparative analyses were performed applying RapidMiner data mining platform. Support vector machine proved to be the best method with maximum accuracy of more than 91%.

  15. Group G streptococcal M protein exhibits structural features analogous to those of class I M protein of group A streptococci.

    PubMed Central

    Collins, C M; Kimura, A; Bisno, A L

    1992-01-01

    We have previously studied a collection of group G streptococcal strains isolated from bacteremic human infections and demonstrated that such strains resist phagocytosis by human polymorphonuclear leukocytes but are type specifically opsonized by homologous antiserum. We have now performed Southern hybridization analysis on genomic DNA from eight blood isolates. All eight isolates showed DNA homology to a group A emm24 gene probe. The M-protein gene of one of the isolates, strain 1750, has now been isolated. This gene (emmG1) encodes a polypeptide of 67 kDa (MG1) which is reactive with antibodies to the partially purified M protein of the parent strain. The predicted amino acid structure of MG1 demonstrates significant identity with the carboxy terminus (C, D, and anchor domains) of M6 and M24 but only limited identity with the amino terminus (variable portion) of these group A M proteins. Southern hybridization of genomic DNA of the eight group G blood isolates with an emmG1 gene probe indicated there were at least four emm alleles associated with these strains. These studies indicate that M proteins of group G streptococci, like those of group A, are genetically heterogeneous. Moreover, MG1 appears to conform to the recently proposed class I structure of M-protein molecules and thus shares certain distinct structural features with the M proteins of well-established rheumatogenic group A streptococcal serotypes. Further comparison of the structures of group G and group A M proteins of throat and skin isolates may cast light on those configurations of the M protein molecules which are and are not critical for the expression of rheumatogenicity. Images PMID:1500178

  16. A Cellulose Synthase-Like Protein Involved in Hyphal Tip Growth and Morphological Differentiation in Streptomyces▿

    PubMed Central

    Xu, Hongbin; Chater, Keith F.; Deng, Zixin; Tao, Meifeng

    2008-01-01

    Cellulose synthase and cellulose synthase-like proteins, responsible for synthesizing β-glucan-containing polysaccharides, play a fundamental role in cellular architectures, such as plant cell and tissue morphogenesis, bacterial biofilm formation, and fruiting-body development. However, the roles of the proteins involved in the developmental process are not well understood. Here, we report that a cellulose synthase-like protein (CslASc) in Streptomyces has a function in hyphal tip growth and morphological differentiation. The cslASc replacement mutant showed pleiotropic defects, including the severe delay of aerial-hyphal formation and altered cell wall morphology. Calcofluor white fluorescence analysis demonstrated that polysaccharide synthesis at hyphal tips was dependent on CslASc. cslASc was constitutively transcribed, and an enhanced green fluorescent protein-CslASc fusion protein was mostly located at the hyphal tips. An extract enriched in morphogenetic chaplin proteins promoted formation of aerial hyphae by the mutant. Furthermore, a two-hybrid experiment indicated that the glycosyltransferase domain of CslASc interacted with the tropomyosin-like polarity-determining DivIVA protein, suggesting that the tip-located DivIVA governed tip recruitment of the CslASc membrane protein. These results imply that the cellulose synthase-like protein couples extracellular and cytoskeletal components functioning in tip growth and cell development. PMID:18487344

  17. Involvement of a tissue-specific RNA recognition motif protein in Drosophila spermatogenesis.

    PubMed Central

    Haynes, S R; Cooper, M T; Pype, S; Stolow, D T

    1997-01-01

    RNA binding proteins mediate posttranscriptional regulation of gene expression via their roles in nuclear and cytoplasmic mRNA metabolism. Many of the proteins involved in these processes have a common RNA binding domain, the RNA recognition motif (RRM). We have characterized the Testis-specific RRM protein gene (Tsr), which plays an important role in spermatogenesis in Drosophila melanogaster. Disruption of Tsr led to a dramatic reduction in male fertility due to the production of spermatids with abnormalities in mitochondrial morphogenesis. Tsr is located on the third chromosome at 87F, adjacent to the nuclear pre-mRNA binding protein gene Hrb87F. A 1.7-kb Tsr transcript was expressed exclusively in the male germ line. It encoded a protein containing two RRMs similar to those found in HRB87F as well as a unique C-terminal domain. TSR protein was located in the cytoplasm of spermatocytes and young spermatids but was absent from mature sperm. The cellular proteins expressed in premeiotic primary spermatocytes from Tsr mutant and wild-type males were assessed by two-dimensional gel electrophoresis. Lack of TSR resulted in the premature expression of a few proteins prior to meiosis; this was abolished by a transgenic copy of Tsr. These data demonstrate that TSR negatively regulated the expression of some testis proteins and, in combination with its expression pattern and subcellular localization, suggest that TSR regulates the stability or translatability of some mRNAs during spermatogenesis. PMID:9111341

  18. 3DSwap: curated knowledgebase of proteins involved in 3D domain swapping.

    PubMed

    Shameer, Khader; Shingate, Prashant N; Manjunath, S C P; Karthika, M; Pugalenthi, Ganesan; Sowdhamini, Ramanathan

    2011-01-01

    Three-dimensional domain swapping is a unique protein structural phenomenon where two or more protein chains in a protein oligomer share a common structural segment between individual chains. This phenomenon is observed in an array of protein structures in oligomeric conformation. Protein structures in swapped conformations perform diverse functional roles and are also associated with deposition diseases in humans. We have performed in-depth literature curation and structural bioinformatics analyses to develop an integrated knowledgebase of proteins involved in 3D domain swapping. The hallmark of 3D domain swapping is the presence of distinct structural segments such as the hinge and swapped regions. We have curated the literature to delineate the boundaries of these regions. In addition, we have defined several new concepts like 'secondary major interface' to represent the interface properties arising as a result of 3D domain swapping, and a new quantitative measure for the 'extent of swapping' in structures. The catalog of proteins reported in 3DSwap knowledgebase has been generated using an integrated structural bioinformatics workflow of database searches, literature curation, by structure visualization and sequence-structure-function analyses. The current version of the 3DSwap knowledgebase reports 293 protein structures, the analysis of such a compendium of protein structures will further the understanding molecular factors driving 3D domain swapping.

  19. Functional dissection of protein complexes involved in yeast chromosome biology using a genetic interaction map.

    PubMed

    Collins, Sean R; Miller, Kyle M; Maas, Nancy L; Roguev, Assen; Fillingham, Jeffrey; Chu, Clement S; Schuldiner, Maya; Gebbia, Marinella; Recht, Judith; Shales, Michael; Ding, Huiming; Xu, Hong; Han, Junhong; Ingvarsdottir, Kristin; Cheng, Benjamin; Andrews, Brenda; Boone, Charles; Berger, Shelley L; Hieter, Phil; Zhang, Zhiguo; Brown, Grant W; Ingles, C James; Emili, Andrew; Allis, C David; Toczyski, David P; Weissman, Jonathan S; Greenblatt, Jack F; Krogan, Nevan J

    2007-04-12

    Defining the functional relationships between proteins is critical for understanding virtually all aspects of cell biology. Large-scale identification of protein complexes has provided one important step towards this goal; however, even knowledge of the stoichiometry, affinity and lifetime of every protein-protein interaction would not reveal the functional relationships between and within such complexes. Genetic interactions can provide functional information that is largely invisible to protein-protein interaction data sets. Here we present an epistatic miniarray profile (E-MAP) consisting of quantitative pairwise measurements of the genetic interactions between 743 Saccharomyces cerevisiae genes involved in various aspects of chromosome biology (including DNA replication/repair, chromatid segregation and transcriptional regulation). This E-MAP reveals that physical interactions fall into two well-represented classes distinguished by whether or not the individual proteins act coherently to carry out a common function. Thus, genetic interaction data make it possible to dissect functionally multi-protein complexes, including Mediator, and to organize distinct protein complexes into pathways. In one pathway defined here, we show that Rtt109 is the founding member of a novel class of histone acetyltransferases responsible for Asf1-dependent acetylation of histone H3 on lysine 56. This modification, in turn, enables a ubiquitin ligase complex containing the cullin Rtt101 to ensure genomic integrity during DNA replication.

  20. Aberrant expression of DNA damage response proteins is associated with breast cancer subtype and clinical features

    PubMed Central

    Guler, Gulnur; Himmetoglu, Cigdem; Jimenez, Rafael E.; Geyer, Susan M.; Wang, Wenle P.; Costinean, Stefan; Pilarski, Robert T.; Morrison, Carl; Suren, Dinc; Liu, Jianhua; Chen, Jingchun; Kamal, Jyoti; Shapiro, Charles L.

    2013-01-01

    Landmark studies of the status of DNA damage checkpoints and associated repair functions in preneoplastic and neoplastic cells has focused attention on importance of these pathways in cancer development, and inhibitors of repair pathways are in clinical trials for treatment of triple negative breast cancer. Cancer heterogeneity suggests that specific cancer subtypes will have distinct mechanisms of DNA damage survival, dependent on biological context. In this study, status of DNA damage response (DDR)-associated proteins was examined in breast cancer subtypes in association with clinical features; 479 breast cancers were examined for expression of DDR proteins γH2AX, BRCA1, pChk2, and p53, DNA damage-sensitive tumor suppressors Fhit and Wwox, and Wwox-interacting proteins Ap2α, Ap2γ, ErbB4, and correlations among proteins, tumor subtypes, and clinical features were assessed. In a multivariable model, triple negative cancers showed significantly reduced Fhit and Wwox, increased p53 and Ap2γ protein expression, and were significantly more likely than other subtype tumors to exhibit aberrant expression of two or more DDR-associated proteins. Disease-free survival was associated with subtype, Fhit and membrane ErbB4 expression level and aberrant expression of multiple DDR-associated proteins. These results suggest that definition of specific DNA repair and checkpoint defects in subgroups of triple negative cancer might identify new treatment targets. Expression of Wwox and its interactor, ErbB4, was highly significantly reduced in metastatic tissues vs. matched primary tissues, suggesting that Wwox signal pathway loss contributes to lymph node metastasis, perhaps by allowing survival of tumor cells that have detached from basement membranes, as proposed for the role of Wwox in ovarian cancer spread. PMID:21069451

  1. Fusions involving protein kinase C and membrane-associated proteins in benign fibrous histiocytoma.

    PubMed

    Płaszczyca, Anna; Nilsson, Jenny; Magnusson, Linda; Brosjö, Otte; Larsson, Olle; Vult von Steyern, Fredrik; Domanski, Henryk A; Lilljebjörn, Henrik; Fioretos, Thoas; Tayebwa, Johnbosco; Mandahl, Nils; Nord, Karolin H; Mertens, Fredrik

    2014-08-01

    Benign fibrous histiocytoma (BFH) is a mesenchymal tumor that most often occurs in the skin (so-called dermatofibroma), but may also appear in soft tissues (so-called deep BFH) and in the skeleton (so-called non-ossifying fibroma). The origin of BFH is unknown, and it has been questioned whether it is a true neoplasm. Chromosome banding, fluorescence in situ hybridization, single nucleotide polymorphism arrays, RNA sequencing, RT-PCR and quantitative real-time PCR were used to search for recurrent somatic mutations in a series of BFH. BFHs were found to harbor recurrent fusions of genes encoding membrane-associated proteins (podoplanin, CD63 and LAMTOR1) with genes encoding protein kinase C (PKC) isoforms PRKCB and PRKCD. PKCs are serine-threonine kinases that through their many phosphorylation targets are implicated in a variety of cellular processes, as well as tumor development. When inactive, the amino-terminal, regulatory domain of PKCs suppresses the activity of their catalytic domain. Upon activation, which requires several steps, they typically translocate to cell membranes, where they interact with different signaling pathways. The detected PDPN-PRKCB, CD63-PRKCD and LAMTOR1-PRKCD gene fusions are all predicted to result in chimeric proteins consisting of the membrane-binding part of PDPN, CD63 or LAMTOR1 and the entire catalytic domain of the PKC. This novel pathogenetic mechanism should result in constitutive kinase activity at an ectopic location. The results show that BFH indeed is a true neoplasm, and that distorted PKC activity is essential for tumorigenesis. The findings also provide means to differentiate BFH from other skin and soft tissue tumors. This article is part of a Directed Issue entitled: Rare cancers.

  2. Expression of proteins involved in host plant defense against greenbug infestation

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The greenbug, Schizaphis graminum (Rondani), has been recognized as a major pest of small grains, including sorghum and wheat. To understand the molecular mechanisms involved in host plant defense against greenbug aphids, a proteomic analysis of greenbug-induced proteins in the seedlings of sorghum...

  3. Spermidine-Induced Improvement of Reconsolidation of Memory Involves Calcium-Dependent Protein Kinase in Rats

    ERIC Educational Resources Information Center

    Girardi, Bruna Amanda; Ribeiro, Daniela Aymone; Signor, Cristiane; Muller, Michele; Gais, Mayara Ana; Mello, Carlos Fernando; Rubin, Maribel Antonello

    2016-01-01

    In this study, we determined whether the calcium-dependent protein kinase (PKC) signaling pathway is involved in the improvement of fear memory reconsolidation induced by the intrahippocampal administration of spermidine in rats. Male Wistar rats were trained in a fear conditioning apparatus using a 0.4-mA footshock as an unconditioned stimulus.…

  4. Protein-protein interactions involving voltage-gated sodium channels: Post-translational regulation, intracellular trafficking and functional expression.

    PubMed

    Shao, Dongmin; Okuse, Kenji; Djamgoz, Mustafa B A

    2009-07-01

    Voltage-gated sodium channels (VGSCs), classically known to play a central role in excitability and signalling in nerves and muscles, have also been found to be expressed in a range of 'non-excitable' cells, including lymphocytes, fibroblasts and endothelia. VGSC abnormalities are associated with various diseases including epilepsy, long-QT syndrome 3, Brugada syndrome, sudden infant death syndrome and, more recently, various human cancers. Given their pivotal role in a wide range of physiological and pathophysiological processes, regulation of functional VGSC expression has been the subject of intense study. An emerging theme is post-translational regulation and macro-molecular complexing by protein-protein interactions and intracellular trafficking, leading to changes in functional VGSC expression in plasma membrane. This partially involves endoplasmic reticulum associated degradation and ubiquitin-proteasome system. Several proteins have been shown to associate with VGSCs. Here, we review the interactions involving VGSCs and the following proteins: p11, ankyrin, syntrophin, beta-subunit of VGSC, papin, ERM and Nedd4 proteins. Protein kinases A and C, as well as Ca(2+)-calmodulin dependent kinase II that have also been shown to regulate intracellular trafficking of VGSCs by changing the balance of externalization vs. internalization, and an effort is made to separate these effects from the short-term phosphorylation of mature proteins in plasma membrane. Two further modulatory mechanisms are reciprocal interactions with the cytoskeleton and, late-stage, activity-dependent regulation. Thus, the review gives an updated account of the range of post-translational molecular mechanisms regulating functional VGSC expression. However, many details of VGSC subtype-specific regulation and pathophysiological aspects remain unknown and these are highlighted throughout for completeness.

  5. Functional features and protein network of human sperm-egg interaction.

    PubMed

    Sabetian, Soudabeh; Shamsir, Mohd Shahir; Abu Naser, Mohammed

    2014-12-01

    Elucidation of the sperm-egg interaction at the molecular level is one of the unresolved problems in sexual reproduction, and understanding the molecular mechanism is crucial in solving problems in infertility and failed in vitro fertilization (IVF). Many molecular interactions in the form of protein-protein interactions (PPIs) mediate the sperm-egg membrane interaction. Due to the complexity of the problem such as difficulties in analyzing in vivo membrane PPIs, many efforts have failed to comprehensively elucidate the fusion mechanism and the molecular interactions that mediate sperm-egg membrane fusion. The main purpose of this study was to reveal possible protein interactions and associated molecular function during sperm-egg interaction using a protein interaction network approach. Different databases have been used to construct the human sperm-egg interaction network. The constructed network revealed new interactions. These included CD151 and CD9 in human oocyte that interact with CD49 in sperm, and CD49 and ITGA4 in sperm that interact with CD63 and CD81, respectively, in the oocyte. These results showed that the different integrins in sperm may be involved in human sperm-egg interaction. It was also suggested that sperm ADAM2 plays a role as a protein candidate involved in sperm-egg membrane interaction by interacting with CD9 in the oocyte. Interleukin-4 receptor activity, receptor signaling protein tyrosine kinase activity, and manganese ion transmembrane transport activity are the major molecular functions in sperm-egg interaction protein network. The disease association analysis indicated that sperm-egg interaction defects are also reflected in other disease networks such as cardiovascular, hematological, and breast cancer diseases. By analyzing the network, we identified the major molecular functions and disease association genes in sperm-egg interaction protein. Further experimental studies will be required to confirm the significance of these new

  6. Differential impact of REM sleep deprivation on cytoskeletal proteins of brain regions involved in sleep regulation.

    PubMed

    Rodríguez-Vázquez, Jennifer; Camacho-Arroyo, Ignacio; Velázquez-Moctezuma, Javier

    2012-01-01

    Rapid eye movement (REM) sleep is involved in memory consolidation, which implies synaptic plasticity. This process requires protein synthesis and the reorganization of the neural cytoskeleton. REM sleep deprivation (REMSD) has an impact on some neuronal proteins involved in synaptic plasticity, such as glutamate receptors and postsynaptic density protein 95, but its effects on cytoskeletal proteins is unknown. In this study, the effects of REMSD on the content of the cytoskeletal proteins MAP2 and TAU were analyzed. Adult female rats were submitted to selective REMSD by using the multiple platform technique. After 24, 48 or 72 h of REMSD, rats were decapitated and the following brain areas were dissected: pons, preoptic area, hippocampus and frontal cortex. Protein extraction and Western blot were performed. Results showed an increase in TAU content in the pons, preoptic area and hippocampus after 24 h of REMSD, while in the frontal cortex a significant increase in TAU content was observed after 72 h of REMSD. A TAU content decrease was observed in the hippocampus after 48 h of REMSD. Interestingly, a marked increase in TAU content was observed after 72 h of REMSD. MAP2 content only increased in the preoptic area at 24 h, and in the frontal cortex after 24 and 72 h of REMSD, without significant changes in the pons and hippocampus. These results support the idea that REM sleep plays an important role in the organization of neural cytoskeleton, and that this effect is tissue-specific.

  7. Disseminated Cytomegalovirus Infection and Protein Losing Enteropathy as Presenting Feature of Pediatric Patient with Crohn's Disease

    PubMed Central

    Ersoz, Safak; Akbulut, Ulas Emre

    2015-01-01

    We report a pediatric patient admitted with abdominal pain, diffuse lower extremity edema and watery diarrhea for two months. Laboratory findings including complete blood count, serum albumin, lipid and immunoglobulin levels were compatible with protein losing enteropathy. Colonoscopic examination revealed diffuse ulcers with smooth raised edge (like "punched out holes") in the colon and terminal ileum. Histopathological examination showed active colitis, ulcerations and inclusion bodies. Immunostaining for cytomegalovirus was positive. Despite supportive management, antiviral therapy, the clinical condition of the patient worsened and developed disseminated cytomegalovirus infection and the patient died. Protein losing enteropathy and disseminated cytomegalovirus infection a presenting of feature in steroid-naive patient with inflammatory bowel disease is very rare. Hypogammaglobulinemia associated with protein losing enteropathy in Crohn's disease may predispose the cytomegalovirus infection in previously healthy children. PMID:25866735

  8. Semi-isometric registration of line features for flexible fitting of protein structures

    PubMed Central

    Abeysinghe, S.; Baker, M. L.; Chiu, W.; Ju, T.

    2010-01-01

    In this paper, we study a registration problem that is motivated by a practical biology problem - fitting protein structures to low-resolution density maps. We consider registration between two sets of lines features (e.g., helices in the proteins) that have undergone not a single, but multiple isometric transformations (e.g., hinge-motions). The problem is further complicated by the presence of symmetry in each set. We formulate the problem as a clique-finding problem in a product graph, and propose a heuristic solution that includes a fast clique-finding algorithm unique to the structure of this graph. When tested on a suite of real protein structures, the algorithm achieved high accuracy even for very large inputs containing hundreds of helices. PMID:21124809

  9. Features of whey protein concentrate supplementation in children with rapidly progressive HIV infection.

    PubMed

    Moreno, Y F; Sgarbieri, V C; da Silva, M N; Toro, A A D C; Vilela, M M S

    2006-02-01

    HIV infection is associated with subnormal GSH levels. An increase in glutathione levels has been observed in HIV-infected adults under oral whey protein supplementation. We studied the features associated with a whey protein concentrate supplementation in children with rapidly progressive AIDS. A prospective double-blind clinical trial was carried out for 4 months with 18 vertically HIV-infected children (1.98-6.37 years), under antiretroviral therapy, who had received whey protein, maltodextrin (placebo) or none. Erythrocyte glutathione concentration, T lymphocyte counts (CD4+ and CD8+) and occurrence of associated co-infections were evaluated. Wilcoxon's and Fischer's Exact tests were used to assess differences between whey protein-supplemented and control (placebo and non-supplemented) groups. A significant median increase of 16.14 mg/dl (p = 0.018) in erythrocyte glutathione levels was observed in the whey protein-supplemented group; the TCD4/CD8 lymphocyte ratio showed a non significant increase and lower occurrence of associated co-infections was also observed. In conclusion, whey protein concentrate supplementation can stimulate glutathione synthesis and, possibly, decrease the occurrence of associated co-infections.

  10. ProFold: Protein Fold Classification with Additional Structural Features and a Novel Ensemble Classifier

    PubMed Central

    2016-01-01

    Protein fold classification plays an important role in both protein functional analysis and drug design. The number of proteins in PDB is very large, but only a very small part is categorized and stored in the SCOPe database. Therefore, it is necessary to develop an efficient method for protein fold classification. In recent years, a variety of classification methods have been used in many protein fold classification studies. In this study, we propose a novel classification method called proFold. We import protein tertiary structure in the period of feature extraction and employ a novel ensemble strategy in the period of classifier training. Compared with existing similar ensemble classifiers using the same widely used dataset (DD-dataset), proFold achieves 76.2% overall accuracy. Another two commonly used datasets, EDD-dataset and TG-dataset, are also tested, of which the accuracies are 93.2% and 94.3%, higher than the existing methods. ProFold is available to the public as a web-server. PMID:27660761

  11. Structural Features and Chaperone Activity of the NudC Protein Family

    SciTech Connect

    Zheng, Meiying; Cierpicki, Tomasz; Burdette, Alexander J.; Utepbergenov, Darkhan; Janczyk, Pawe; #322; #321; .; Derewenda, Urszula; Stukenberg, P. Todd; Caldwell, Kim A.; Derewenda, Zygmunt S.

    2012-05-25

    The NudC family consists of four conserved proteins with representatives in all eukaryotes. The archetypal nudC gene from Aspergillus nidulans is a member of the nud gene family that is involved in the maintenance of nuclear migration. This family also includes nudF, whose human orthologue, Lis1, codes for a protein essential for brain cortex development. Three paralogues of NudC are known in vertebrates: NudC, NudC-like (NudCL), and NudC-like 2 (NudCL2). The fourth distantly related member of the family, CML66, contains a NudC-like domain. The three principal NudC proteins have no catalytic activity but appear to play as yet poorly defined roles in proliferating and dividing cells. We present crystallographic and NMR studies of the human NudC protein and discuss the results in the context of structures recently deposited by structural genomics centers (i.e., NudCL and mouse NudCL2). All proteins share the same core CS domain characteristic of proteins acting either as cochaperones of Hsp90 or as independent small heat shock proteins. However, while NudC and NudCL dimerize via an N-terminally located coiled coil, the smaller NudCL2 lacks this motif and instead dimerizes as a result of unique domain swapping. We show that NudC and NudCL, but not NudCL2, inhibit the aggregation of several target proteins, consistent with an Hsp90-independent heat shock protein function. Importantly, and in contrast to several previous reports, none of the three proteins is able to form binary complexes with Lis1. The availability of structural information will be of help in further studies on the cellular functions of the NudC family.

  12. Nuclear substructure reorganization during late stageerythropoiesis is selective and does not involve caspase cleavage ofmajor nuclear substructural proteins

    SciTech Connect

    Krauss, Sharon Wald; Lo, Annie J.; Short, Sarah A.; Koury, MarkJ.; Mohandas, Narla; Chasis, Joel Anne

    2005-04-06

    Enucleation, a rare feature of mammalian differentiation, occurs in three cell types: erythroblasts, lens epithelium and keratinocytes. Previous investigations suggest that caspase activation functions in lens epithelial and keratinocyte enucleation, as well as in early erythropoiesis encompassing BFU-E differentiation to proerythroblast. To determine whether caspase activation contributes to later erythropoiesis and whether nuclear substructures other than chromatin reorganize, we analyzed distributions of nuclear subcompartment proteins and assayed for caspase-induced cleavage of subcompartmental target proteins in mouse erythroblasts. We found that patterns of lamin B in the filamentous network interacting with both the nuclear envelope and DNA, nuclear matrix protein NuMA, and splicing factors Sm and SC35 persisted during nuclear condensation, consistent with effective transcription of genes expressed late in differentiation. Thus nuclear reorganization prior to enucleation is selective, allowing maintenance of critical transcriptional processes independent of extensive chromosomal reorganization. Consistent with these data, we found no evidence for caspase-induced cleavage of major nuclear subcompartment proteins during late erythropoiesis, in contrast to what has been observed in early erythropoiesis and in lens epithelial and keratinocyte differentiation. These findings imply that nuclear condensation and extrusion during terminal erythroid differentiation involve novel mechanisms that do not entail major activation of apoptotic machinery.

  13. Structure of astrotactin-2: a conserved vertebrate-specific and perforin-like membrane protein involved in neuronal development

    PubMed Central

    Ni, Tao; Harlos, Karl; Gilbert, Robert

    2016-01-01

    The vertebrate-specific proteins astrotactin-1 and 2 (ASTN-1 and ASTN-2) are integral membrane perforin-like proteins known to play critical roles in neurodevelopment, while ASTN-2 has been linked to the planar cell polarity pathway in hair cells. Genetic variations associated with them are linked to a variety of neurodevelopmental disorders and other neurological pathologies, including an advanced onset of Alzheimer's disease. Here we present the structure of the majority endosomal region of ASTN-2, showing it to consist of a unique combination of polypeptide folds: a perforin-like domain, a minimal epidermal growth factor-like module, a unique form of fibronectin type III domain and an annexin-like domain. The perforin-like domain differs from that of other members of the membrane attack complex-perforin (MACPF) protein family in ways that suggest ASTN-2 does not form pores. Structural and biophysical data show that ASTN-2 (but not ASTN-1) binds inositol triphosphates, suggesting a mechanism for membrane recognition or secondary messenger regulation of its activity. The annexin-like domain is closest in fold to repeat three of human annexin V and similarly binds calcium, and yet shares no sequence homology with it. Overall, our structure provides the first atomic-resolution description of a MACPF protein involved in development, while highlighting distinctive features of ASTN-2 responsible for its activity. PMID:27249642

  14. Proteomic Analysis Reveals Proteins Involved in Seed Imbibition under Salt Stress in Rice

    PubMed Central

    Xu, Enshun; Chen, Mingming; He, Hui; Zhan, Chengfang; Cheng, Yanhao; Zhang, Hongsheng; Wang, Zhoufei

    2017-01-01

    Enhancement of salinity tolerance during seed germination is very important for direct seeding in rice. In this study, the salt-tolerant japonica landrace Jiucaiqing was used to determine the regulators that are involved in seed imbibition under salt stress. Briefly, the comparative proteomic analysis was conducted between dry (0 h) and imbibed (24 h) seeds with 150 mM NaCl. Under salt stress, the uptake of water increased rapidly before 24 h imbibition (Phase I), followed by a plateau of seed imbibition from 24 to 96 h imbibition (Phase II). We identified 14 proteins involved in seed imbibition, in which the majority of these proteins were involved in energy supply and storage protein. The early imbibition process was mediated by protein catabolism; the most of proteins were down-regulated after 24 h imbibition. Eleven genes in salt stress treated seeds were expressed early during the seed imbibition in comparison to control seeds. By comparison, 2,3-bisphosphoglycerate-independent phosphoglycerate mutase (BPM), glutelin (GLU2.2 and GLU2.3), glucose-1-phosphate adenylyltransferase large subunit (GAS8), and cupin domain containing protein (CDP3.1 and CDP3.2) were near the regions of quantitative trait loci (QTLs) for seed dormancy, seed reserve utilization, and seed germination in Jiucaiqing. In particular, CDP3.1 was co-located in the region of qIR-3 for imbibition rate, and qGP-3 for germination percentage. The role of CDP3.1 was verified in enhancing seed germination under salt stress using T-DNA mutant. The identified proteins might be applicable for the improvement of seed germination under salt stress in rice. PMID:28105039

  15. Paraoxonase 1 and dietary hyperhomocysteinemia modulate the expression of mouse proteins involved in liver homeostasis.

    PubMed

    Suszyńska-Zajczyk, Joanna; Jakubowski, Hieronim

    2014-01-01

    Homocysteine (Hcy), a product of methionine metabolism, is elevated by the consumption of a high-methionine diet that can cause fatty liver disease. Paraoxonase 1 (Pon1), a hydrolase expressed mainly in the liver and carried in the circulation on high-density lipoprotein, participates in Hcy metabolism. Low Pon1 activity is linked to fatty liver disease. We hypothesize that hyperhomocysteinemia and low Pon1 induce changes in gene expression that could impair liver homeostasis. To test this hypothesis, we analyzed the liver proteome of Pon1(-/-) and Pon1(+/+) mice fed a high methionine diet (1% methionine in the drinking water) for 8 weeks using 2D IEF/SDS-PAGE gel electrophoresis and MALDI-TOF mass spectrometry. We identified seven liver proteins whose expression was significantly altered in Pon1(-/-) mice. In animals fed with a control diet, the expression of three liver proteins involved in lipoprotein metabolism (ApoE), iron metabolism (Ftl), and regulation of nitric oxide generation (Ddah1) was up-regulated by the Pon1(-/-) genotype. In mice fed with a high-methionine diet, expression of four liver proteins was up-regulated and of three proteins was down-regulated by the Pon1(-/-) genotype. The up-regulated proteins are involved in lipoprotein metabolism (ApoE), energy metabolism (Atp5h), oxidative stress response (Prdx2), and nitric oxide regulation (Ddah1). The down-regulated proteins are involved in energy metabolism (Gamt), iron metabolism (Ftl), and catechol metabolism (Comt). Expression of one protein (Ftl) was up-regulated both by the Pon1(-/-) genotype and a high-methionine diet. Our findings suggest that Pon1 interacts with diverse cellular processes - from lipoprotein metabolism, nitric oxide regulation, and energy metabolism to iron transport and antioxidant defenses - that are essential for normal liver homeostasis and modulation of these interactions by a high-methionine diet may contribute to fatty liver disease.

  16. Proteomic Analysis Reveals Proteins Involved in Seed Imbibition under Salt Stress in Rice.

    PubMed

    Xu, Enshun; Chen, Mingming; He, Hui; Zhan, Chengfang; Cheng, Yanhao; Zhang, Hongsheng; Wang, Zhoufei

    2016-01-01

    Enhancement of salinity tolerance during seed germination is very important for direct seeding in rice. In this study, the salt-tolerant japonica landrace Jiucaiqing was used to determine the regulators that are involved in seed imbibition under salt stress. Briefly, the comparative proteomic analysis was conducted between dry (0 h) and imbibed (24 h) seeds with 150 mM NaCl. Under salt stress, the uptake of water increased rapidly before 24 h imbibition (Phase I), followed by a plateau of seed imbibition from 24 to 96 h imbibition (Phase II). We identified 14 proteins involved in seed imbibition, in which the majority of these proteins were involved in energy supply and storage protein. The early imbibition process was mediated by protein catabolism; the most of proteins were down-regulated after 24 h imbibition. Eleven genes in salt stress treated seeds were expressed early during the seed imbibition in comparison to control seeds. By comparison, 2,3-bisphosphoglycerate-independent phosphoglycerate mutase (BPM), glutelin (GLU2.2 and GLU2.3), glucose-1-phosphate adenylyltransferase large subunit (GAS8), and cupin domain containing protein (CDP3.1 and CDP3.2) were near the regions of quantitative trait loci (QTLs) for seed dormancy, seed reserve utilization, and seed germination in Jiucaiqing. In particular, CDP3.1 was co-located in the region of qIR-3 for imbibition rate, and qGP-3 for germination percentage. The role of CDP3.1 was verified in enhancing seed germination under salt stress using T-DNA mutant. The identified proteins might be applicable for the improvement of seed germination under salt stress in rice.

  17. Silkmapin of Hyriopsis cumingii, a novel silk-like shell matrix protein involved in nacre formation.

    PubMed

    Liu, Xiaojun; Dong, Shaojian; Jin, Can; Bai, Zhiyi; Wang, Guiling; Li, Jiale

    2015-01-25

    Understanding the role of matrix proteins in nacre formation and biomineralization in mollusks is important for the pearl industry. In this study, the gene encoding the novel Hyriopsis cumingii shell matrix protein silkmapin was characterized. The gene encodes a protein of 30.89kDa in which Gly accounts for 34.41% of the amino acid content, and the C-terminal region binds Ca(2+). Secondary structure prediction indicated a predominantly β-fold and a structure typical of filamentous proteins. Real-time quantitative PCR and in situ hybridization showed that silkmapin was expressed in epithelial cells at the edge and pallial of mantle tissue, indicated that silkmapin play roles in the shell nacreous and prismatic layer formation. Further real-time PCR results indicated an involvement in pearl formation via nucleation of calcium carbonate prior to formation of the nacre.

  18. Protein Folding Activity of the Ribosome is involved in Yeast Prion Propagation

    PubMed Central

    Blondel, Marc; Soubigou, Flavie; Evrard, Justine; Nguyen, Phu hai; Hasin, Naushaba; Chédin, Stéphane; Gillet, Reynald; Contesse, Marie-Astrid; Friocourt, Gaëlle; Stahl, Guillaume; Jones, Gary W.; Voisset, Cécile

    2016-01-01

    6AP and GA are potent inhibitors of yeast and mammalian prions and also specific inhibitors of PFAR, the protein-folding activity borne by domain V of the large rRNA of the large subunit of the ribosome. We therefore explored the link between PFAR and yeast prion [PSI+] using both PFAR-enriched mutants and site-directed methylation. We demonstrate that PFAR is involved in propagation and de novo formation of [PSI+]. PFAR and the yeast heat-shock protein Hsp104 partially compensate each other for [PSI+] propagation. Our data also provide insight into new functions for the ribosome in basal thermotolerance and heat-shocked protein refolding. PFAR is thus an evolutionarily conserved cell component implicated in the prion life cycle, and we propose that it could be a potential therapeutic target for human protein misfolding diseases. PMID:27633137

  19. Identification and Characterization of Proteins Involved in Rice Urea and Arginine Catabolism1[W

    PubMed Central

    Cao, Feng-Qiu; Werner, Andrea K.; Dahncke, Kathleen; Romeis, Tina; Liu, Lai-Hua; Witte, Claus-Peter

    2010-01-01

    Rice (Oryza sativa) production relies strongly on nitrogen (N) fertilization with urea, but the proteins involved in rice urea metabolism have not yet been characterized. Coding sequences for rice arginase, urease, and the urease accessory proteins D (UreD), F (UreF), and G (UreG) involved in urease activation were identified and cloned. The functionality of urease and the urease accessory proteins was demonstrated by complementing corresponding Arabidopsis (Arabidopsis thaliana) mutants and by multiple transient coexpression of the rice proteins in Nicotiana benthamiana. Secondary structure models of rice (plant) UreD and UreF proteins revealed a possible functional conservation to bacterial orthologs, especially for UreF. Using amino-terminally StrepII-tagged urease accessory proteins, an interaction between rice UreD and urease could be shown. Prokaryotic and eukaryotic urease activation complexes seem conserved despite limited protein sequence conservation for UreF and UreD. In plant metabolism, urea is generated by the arginase reaction. Rice arginase was transiently expressed as a carboxyl-terminally StrepII-tagged fusion protein in N. benthamiana, purified, and biochemically characterized (Km = 67 mm, kcat = 490 s−1). The activity depended on the presence of manganese (Kd = 1.3 μm). In physiological experiments, urease and arginase activities were not influenced by the external N source, but sole urea nutrition imbalanced the plant amino acid profile, leading to the accumulation of asparagine and glutamine in the roots. Our data indicate that reduced plant performance with urea as N source is not a direct result of insufficient urea metabolism but may in part be caused by an imbalance of N distribution. PMID:20631318

  20. Identification and characterization of proteins involved in rice urea and arginine catabolism.

    PubMed

    Cao, Feng-Qiu; Werner, Andrea K; Dahncke, Kathleen; Romeis, Tina; Liu, Lai-Hua; Witte, Claus-Peter

    2010-09-01

    Rice (Oryza sativa) production relies strongly on nitrogen (N) fertilization with urea, but the proteins involved in rice urea metabolism have not yet been characterized. Coding sequences for rice arginase, urease, and the urease accessory proteins D (UreD), F (UreF), and G (UreG) involved in urease activation were identified and cloned. The functionality of urease and the urease accessory proteins was demonstrated by complementing corresponding Arabidopsis (Arabidopsis thaliana) mutants and by multiple transient coexpression of the rice proteins in Nicotiana benthamiana. Secondary structure models of rice (plant) UreD and UreF proteins revealed a possible functional conservation to bacterial orthologs, especially for UreF. Using amino-terminally StrepII-tagged urease accessory proteins, an interaction between rice UreD and urease could be shown. Prokaryotic and eukaryotic urease activation complexes seem conserved despite limited protein sequence conservation for UreF and UreD. In plant metabolism, urea is generated by the arginase reaction. Rice arginase was transiently expressed as a carboxyl-terminally StrepII-tagged fusion protein in N. benthamiana, purified, and biochemically characterized (K(m) = 67 mm, k(cat) = 490 s(-1)). The activity depended on the presence of manganese (K(d) = 1.3 microm). In physiological experiments, urease and arginase activities were not influenced by the external N source, but sole urea nutrition imbalanced the plant amino acid profile, leading to the accumulation of asparagine and glutamine in the roots. Our data indicate that reduced plant performance with urea as N source is not a direct result of insufficient urea metabolism but may in part be caused by an imbalance of N distribution.

  1. Staphylococcus aureus surface proteins involved in adaptation to oxacillin identified using a novel cell shaving approach.

    PubMed

    Solis, Nestor; Parker, Benjamin L; Kwong, Stephen M; Robinson, Gareth; Firth, Neville; Cordwell, Stuart J

    2014-06-06

    Staphylococcus aureus is a Gram-positive pathogen responsible for a variety of infections, and some strains are resistant to virtually all classes of antibiotics. Cell shaving proteomics using a novel probability scoring algorithm to compare the surfaceomes of the methicillin-resistant, laboratory-adapted S. aureus COL strain with a COL strain in vitro adapted to high levels of oxacillin (APT). APT displayed altered cell morphology compared with COL and increased aggregation in biofilm assays. Increased resistance to β-lactam antibiotics was observed, but adaptation to oxacillin did not confer multidrug resistance. Analysis of the S. aureus COL and APT surfaceomes identified 150 proteins at a threshold determined by the scoring algorithm. Proteins unique to APT included the LytR-CpsA-Psr (LCP) domain-containing MsrR and SACOL2302. Quantitative RT-PCR showed increased expression of sacol2302 in APT grown with oxacillin (>6-fold compared with COL). Overexpression of sacol2302 in COL to levels consistent with APT (+ oxacillin) did not influence biofilm formation or β-lactam resistance. Proteomics using iTRAQ and LC-MS/MS identified 1323 proteins (∼50% of the theoretical S. aureus proteome), and cluster analysis demonstrated elevated APT abundances of LCP proteins, capsule and peptidoglycan biosynthesis proteins, and proteins involved in wall remodelling. Adaptation to oxacillin also induced urease proteins, which maintained culture pH compared to COL. These results show that S. aureus modifies surface architecture in response to antibiotic adaptation.

  2. Evidence against the involvement of ionically bound cell wall proteins in pea epicotyl growth

    NASA Technical Reports Server (NTRS)

    Melan, M. A.; Cosgrove, D. J.

    1988-01-01

    Ionically bound cell wall proteins were extracted from 7 day old etiolated pea (Pisum sativum L. cv Alaska) epicotyls with 3 molar LiCl. Polyclonal antiserum was raised in rabbits against the cell wall proteins. Growth assays showed that treatment of growing region segments (5-7 millimeters) of peas with either dialyzed serum, serum globulin fraction, affinity purified immunoglobulin, or papain-cleaved antibody fragments had no effect on growth. Immunofluorescence microscopy confirmed antibody binding to cell walls and penetration of the antibodies into the tissues. Western blot analysis, immunoassay results, and affinity chromatography utilizing Sepharose-bound antibodies confirmed recognition of the protein preparation by the antibodies. Experiments employing in vitro extension as a screening measure indicated no effect upon extension by antibodies, by 50 millimolar LiCl perfusion of the apoplast or by 3 molar LiCl extraction. Addition of cell wall protein to protease pretreated segments did not restore extension nor did addition of cell wall protein to untreated segments increase extension. It is concluded that, although evidence suggests that protein is responsible for the process of extension, the class(es) of proteins which are extracted from pea cell walls with 3 molar LiCl are probably not involved in this process.

  3. Identification of glandular (preputial and clitoral) proteins in house rat (Rattus rattus) involved in pheromonal communication.

    PubMed

    Archunan, G; Kamalakkannan, S; Achiraman, S; Rajkumar, R

    2004-10-01

    Proteins (18-20 kDa) belonging to lipocalin family have been reported to act as carriers for ligands binding to pheromones in mouse urine, pig saliva, hamster vaginal fluid and human sweat, that are involved in pheromonal communication. As the preputial gland is a major pheromonal source, the present study was aimed to detect the specific protein bands (around 18-20 kDa) in the preputial and clitoral glands of the house rat, R. rattus. The amount of protein was higher in preputial gland of the male than that of female (clitoral) gland. A 20 kDa protein was noted in male and female glands; however, the intensity of the band was much higher in male than in female. In addition, 70, 60, 35 kDa bands, identified in male preputial gland, were absent in females. The presence of higher concentration of glandular proteins in the male preputial gland suggests that male rats may depend more on these glandular proteins for the maintenance of reproductive and dominance behaviours. The results further suggest that these glandular proteins (20 kDa) may act as a carrier for ligand binding.

  4. LanCL proteins are not Involved in Lanthionine Synthesis in Mammals.

    PubMed

    He, Chang; Zeng, Min; Dutta, Debapriya; Koh, Tong Hee; Chen, Jie; van der Donk, Wilfred A

    2017-01-20

    LanC-like (LanCL) proteins are mammalian homologs of bacterial LanC enzymes, which catalyze the addition of the thiol of Cys to dehydrated Ser residues during the biosynthesis of lanthipeptides, a class of natural products formed by post-translational modification of precursor peptides. The functions of LanCL proteins are currently unclear. A recent proposal suggested that LanCL1 catalyzes the addition of the Cys of glutathione to protein- or peptide-bound dehydroalanine (Dha) to form lanthionine, analogous to the reaction catalyzed by LanC in bacteria. Lanthionine has been detected in human brain as the downstream metabolite lanthionine ketimine (LK), which has been shown to have neuroprotective effects. In this study, we tested the proposal that LanCL1 is involved in lanthionine biosynthesis by constructing LanCL1 knock-out mice and measuring LK concentrations in their brains using a mass spectrometric detection method developed for this purpose. To investigate whether other LanCL proteins (LanCL2/3) may confer a compensatory effect, triple knock-out (TKO) mice were also generated and tested. Very similar concentrations of LK (0.5-2.5 nmol/g tissue) were found in LanCL1 knock-out, TKO and wild type (WT) mouse brains, suggesting that LanCL proteins are not involved in lanthionine biosynthesis.

  5. LanCL proteins are not Involved in Lanthionine Synthesis in Mammals

    PubMed Central

    He, Chang; Zeng, Min; Dutta, Debapriya; Koh, Tong Hee; Chen, Jie; van der Donk, Wilfred A.

    2017-01-01

    LanC-like (LanCL) proteins are mammalian homologs of bacterial LanC enzymes, which catalyze the addition of the thiol of Cys to dehydrated Ser residues during the biosynthesis of lanthipeptides, a class of natural products formed by post-translational modification of precursor peptides. The functions of LanCL proteins are currently unclear. A recent proposal suggested that LanCL1 catalyzes the addition of the Cys of glutathione to protein- or peptide-bound dehydroalanine (Dha) to form lanthionine, analogous to the reaction catalyzed by LanC in bacteria. Lanthionine has been detected in human brain as the downstream metabolite lanthionine ketimine (LK), which has been shown to have neuroprotective effects. In this study, we tested the proposal that LanCL1 is involved in lanthionine biosynthesis by constructing LanCL1 knock-out mice and measuring LK concentrations in their brains using a mass spectrometric detection method developed for this purpose. To investigate whether other LanCL proteins (LanCL2/3) may confer a compensatory effect, triple knock-out (TKO) mice were also generated and tested. Very similar concentrations of LK (0.5–2.5 nmol/g tissue) were found in LanCL1 knock-out, TKO and wild type (WT) mouse brains, suggesting that LanCL proteins are not involved in lanthionine biosynthesis. PMID:28106097

  6. Endogenous basic fibroblast growth factor isoforms involved in different intracellular protein complexes.

    PubMed Central

    Patry, V; Bugler, B; Maret, A; Potier, M; Prats, H

    1997-01-01

    Four forms of basic fibroblast growth factor (bFGF or FGF-2) result from an alternative initiation of translation involving one AUG (155-amino acid form) and three CUGs (210-, 201- and 196-amino acid forms). These different forms of bFGF show different intracellular biological activities. To identify their intracellular targets, the 210- and 155-amino acid forms of bFGF were independently transfected into CHO cells and their correct subcellular localizations were verified, the 155-amino acid bFGF form being essentially cytoplasmic whereas the 210-amino acid protein was nuclear. The radiation fragmentation method was used to determine the target size of the different bFGF isoforms in the transfected CHO cells and to show that the 210- and 155-amino acids bFGF isoforms were included in protein complexes of 320 and 130 kDa respectively. Similar results were obtained using the SK-Hep1 cell line, which naturally expressed all forms of bFGF. Co-immunoprecipitation assays using different chimaeric bFGF-chloramphenicol acetyltransferase proteins showed that different cellular proteins are associated with different parts of the bFGF molecule. We conclude that bFGF isoforms are involved in different molecular complexes in the cytosol and nucleus, which would reflect different functions for these proteins. PMID:9337877

  7. Robust feature generation for protein subchloroplast location prediction with a weighted GO transfer model.

    PubMed

    Li, Xiaomei; Wu, Xindong; Wu, Gongqing

    2014-04-21

    Chloroplasts are crucial organelles of green plants and eukaryotic algae since they conduct photosynthesis. Predicting the subchloroplast location of a protein can provide important insights for understanding its biological functions. The performance of subchloroplast location prediction algorithms often depends on deriving predictive and succinct features from genomic and proteomic data. In this work, a novel weighted Gene Ontology (GO) transfer model is proposed to generate discriminating features from sequence data and GO Categories. This model contains two components. First, we transfer the GO terms of the homologous protein, and then assign the bit-score as weights to GO features. Second, we employ term-selection methods to determine weights for GO terms. This model is capable of improving prediction accuracy due to the tolerance of the noise derived from homolog knowledge transfer. The proposed weighted GO transfer method based on bit-score and a logarithmic transformation of CHI-square (WS-LCHI) performs better than the baseline models, and also outperforms the four off-the-shelf subchloroplast prediction methods.

  8. Proteins involved in pRb and p53 pathways are differentially expressed in thin and thick superficial spreading melanomas.

    PubMed

    de Sá, Bianca Costa Soares; Fugimori, Melissa Lissae; Ribeiro, Karina de Cássia Braga; Duprat Neto, João Pedreira; Neves, Rogério Izar; Landman, Gilles

    2009-06-01

    Cutaneous melanoma is one of the leading causes of cancer-related death. Malignant transformation of epidermal melanocytes is a multifactorial process involving cell cycle and death control pathways. The purpose of this study was to analyze the immunohistochemical expression of cell-cycle-related and apoptosis-related proteins in cutaneous superficial spreading melanomas using the tissue microarray technique to further understand tumor development. A total of 20 samples of in-situ melanomas and 44 melanomas proteins: p16INK4 (p16), cyclin D1, cyclin-dependent kinase 4 (Cdk4), retinoblastoma protein, tumor suppressor protein p53, and p21 cell cycle regulator (p21) using a streptavidine-biotin-peroxidase technique for immunohistochemistry. Thick melanomas (>1.0 mm) and metastases lost p16 expression in 100% of the cases and in-situ and thin melanomas (protein p53, and p21. Primary tumors, when compared with metastases, had higher cytoplasmatic Cdk4 expression. None of the studied proteins influenced overall or disease-free survival. Our results suggest that loss of p16 expression was a constant feature in primary and metastatic melanomas. Cyclin D1 expression seems to be related to initial phases of melanoma development. An increase in p21 expression could represent a cell cycle control in proliferating cells with reduced p16 and/or increased nuclear Cdk4 expression.

  9. Protein subcellular localization prediction based on compartment-specific features and structure conservation

    PubMed Central

    Su, Emily Chia-Yu; Chiu, Hua-Sheng; Lo, Allan; Hwang, Jenn-Kang; Sung, Ting-Yi; Hsu, Wen-Lian

    2007-01-01

    Background Protein subcellular localization is crucial for genome annotation, protein function prediction, and drug discovery. Determination of subcellular localization using experimental approaches is time-consuming; thus, computational approaches become highly desirable. Extensive studies of localization prediction have led to the development of several methods including composition-based and homology-based methods. However, their performance might be significantly degraded if homologous sequences are not detected. Moreover, methods that integrate various features could suffer from the problem of low coverage in high-throughput proteomic analyses due to the lack of information to characterize unknown proteins. Results We propose a hybrid prediction method for Gram-negative bacteria that combines a one-versus-one support vector machines (SVM) model and a structural homology approach. The SVM model comprises a number of binary classifiers, in which biological features derived from Gram-negative bacteria translocation pathways are incorporated. In the structural homology approach, we employ secondary structure alignment for structural similarity comparison and assign the known localization of the top-ranked protein as the predicted localization of a query protein. The hybrid method achieves overall accuracy of 93.7% and 93.2% using ten-fold cross-validation on the benchmark data sets. In the assessment of the evaluation data sets, our method also attains accurate prediction accuracy of 84.0%, especially when testing on sequences with a low level of homology to the training data. A three-way data split procedure is also incorporated to prevent overestimation of the predictive performance. In addition, we show that the prediction accuracy should be approximately 85% for non-redundant data sets of sequence identity less than 30%. Conclusion Our results demonstrate that biological features derived from Gram-negative bacteria translocation pathways yield a significant

  10. Yeast prions and human prion-like proteins: sequence features and prediction methods.

    PubMed

    Cascarina, Sean M; Ross, Eric D

    2014-06-01

    Prions are self-propagating infectious protein isoforms. A growing number of prions have been identified in yeast, each resulting from the conversion of soluble proteins into an insoluble amyloid form. These yeast prions have served as a powerful model system for studying the causes and consequences of prion aggregation. Remarkably, a number of human proteins containing prion-like domains, defined as domains with compositional similarity to yeast prion domains, have recently been linked to various human degenerative diseases, including amyotrophic lateral sclerosis. This suggests that the lessons learned from yeast prions may help in understanding these human diseases. In this review, we examine what has been learned about the amino acid sequence basis for prion aggregation in yeast, and how this information has been used to develop methods to predict aggregation propensity. We then discuss how this information is being applied to understand human disease, and the challenges involved in applying yeast prediction methods to higher organisms.

  11. Conserved Features in the Structure, Mechanism, and Biogenesis of the Inverse Autotransporter Protein Family

    PubMed Central

    Heinz, Eva; Stubenrauch, Christopher J.; Grinter, Rhys; Croft, Nathan P.; Purcell, Anthony W.; Strugnell, Richard A.; Dougan, Gordon; Lithgow, Trevor

    2016-01-01

    The bacterial cell surface proteins intimin and invasin are virulence factors that share a common domain structure and bind selectively to host cell receptors in the course of bacterial pathogenesis. The β-barrel domains of intimin and invasin show significant sequence and structural similarities. Conversely, a variety of proteins with sometimes limited sequence similarity have also been annotated as “intimin-like” and “invasin” in genome datasets, while other recent work on apparently unrelated virulence-associated proteins ultimately revealed similarities to intimin and invasin. Here we characterize the sequence and structural relationships across this complex protein family. Surprisingly, intimins and invasins represent a very small minority of the sequence diversity in what has been previously the “intimin/invasin protein family”. Analysis of the assembly pathway for expression of the classic intimin, EaeA, and a characteristic example of the most prevalent members of the group, FdeC, revealed a dependence on the translocation and assembly module as a common feature for both these proteins. While the majority of the sequences in the grouping are most similar to FdeC, a further and widespread group is two-partner secretion systems that use the β-barrel domain as the delivery device for secretion of a variety of virulence factors. This comprehensive analysis supports the adoption of the “inverse autotransporter protein family” as the most accurate nomenclature for the family and, in turn, has important consequences for our overall understanding of the Type V secretion systems of bacterial pathogens. PMID:27190006

  12. Protein Kinase C-{delta} mediates down-regulation of heterogeneous nuclear ribonucleoprotein K protein: involvement in apoptosis induction

    SciTech Connect

    Gao, Feng-Hou; Wu, Ying-Li; Zhao, Meng; Chen, Guo-Qiang

    2009-11-15

    We reported previously that NSC606985, a camptothecin analogue, induces apoptosis of acute myeloid leukemia (AML) cells through proteolytic activation of protein kinase C delta ({Delta}PKC-{delta}). By subcellular proteome analysis, heterogeneous nuclear ribonucleoprotein K (hnRNP K) was identified as being significantly down-regulated in NSC606985-treated leukemic NB4 cells. HnRNP K, a docking protein for DNA, RNA, and transcriptional or translational molecules, is implicated in a host of processes involving the regulation of gene expression. However, the molecular mechanisms of hnRNP K reduction and its roles during apoptosis are still not understood. In the present study, we found that, following the appearance of the {Delta}PKC-{delta}, hnRNP K protein was significantly down-regulated in NSC606985, doxorubicin, arsenic trioxide and ultraviolet-induced apoptosis. We further provided evidence that {Delta}PKC-{delta} mediated the down-regulation of hnRNP K protein during apoptosis: PKC-{delta} inhibitor could rescue the reduction of hnRNP K; hnRNP K failed to be decreased in PKC-{delta}-deficient apoptotic KG1a cells; conditional induction of {Delta}PKC-{delta} in U937T cells directly down-regulated hnRNP K protein. Moreover, the proteasome inhibitor also inhibited the down-regulation of hnRNP K protein by apoptosis inducer and the conditional expression of {Delta}PKC-{delta}. More intriguingly, the suppression of hnRNP K with siRNA transfection significantly induced apoptosis. To our knowledge, this is the first demonstration that proteolytically activated PKC-{delta} down-regulates hnRNP K protein in a proteasome-dependent manner, which plays an important role in apoptosis induction.

  13. Nanometer polymer surface features: the influence on surface energy, protein adsorption and endothelial cell adhesion

    NASA Astrophysics Data System (ADS)

    Carpenter, Joseph; Khang, Dongwoo; Webster, Thomas J.

    2008-12-01

    Current small diameter (<5 mm) synthetic vascular graft materials exhibit poor long-term patency due to thrombosis and intimal hyperplasia. Tissue engineered solutions have yielded functional vascular tissue, but some require an eight-week in vitro culture period prior to implantation—too long for immediate clinical bedside applications. Previous in vitro studies have shown that nanostructured poly(lactic-co-glycolic acid) (PLGA) surfaces elevated endothelial cell adhesion, proliferation, and extracellular matrix synthesis when compared to nanosmooth surfaces. Nonetheless, these studies failed to address the importance of lateral and vertical surface feature dimensionality coupled with surface free energy; nor did such studies elicit an optimum specific surface feature size for promoting endothelial cell adhesion. In this study, a series of highly ordered nanometer to submicron structured PLGA surfaces of identical chemistry were created using a technique employing polystyrene nanobeads and poly(dimethylsiloxane) (PDMS) molds. Results demonstrated increased endothelial cell adhesion on PLGA surfaces with vertical surface features of size less than 18.87 nm but greater than 0 nm due to increased surface energy and subsequently protein (fibronectin and collagen type IV) adsorption. Furthermore, this study provided evidence that the vertical dimension of nanometer surface features, rather than the lateral dimension, is largely responsible for these increases. In this manner, this study provides key design parameters that may promote vascular graft efficacy.

  14. Absence of aquaporin-4 in skeletal muscle alters proteins involved in bioenergetic pathways and calcium handling.

    PubMed

    Basco, Davide; Nicchia, Grazia Paola; D'Alessandro, Angelo; Zolla, Lello; Svelto, Maria; Frigeri, Antonio

    2011-04-28

    Aquaporin-4 (AQP4) is a water channel expressed at the sarcolemma of fast-twitch skeletal muscle fibers, whose expression is altered in several forms of muscular dystrophies. However, little is known concerning the physiological role of AQP4 in skeletal muscle and its functional and structural interaction with skeletal muscle proteome. Using AQP4-null mice, we analyzed the effect of the absence of AQP4 on the morphology and protein composition of sarcolemma as well as on the whole skeletal muscle proteome. Immunofluorescence analysis showed that the absence of AQP4 did not perturb the expression and cellular localization of the dystrophin-glycoprotein complex proteins, aside from those belonging to the extracellular matrix, and no alteration was found in sarcolemma integrity by dye extravasation assay. With the use of a 2DE-approach (BN/SDS-PAGE), protein maps revealed that in quadriceps, out of 300 Coomassie-blue detected and matched spots, 19 proteins exhibited changed expression in AQP4(-/-) compared to WT mice. In particular, comparison of the protein profiles revealed 12 up- and 7 down-regulated protein spots in AQP4-/- muscle. Protein identification by MS revealed that the perturbed expression pattern belongs to proteins involved in energy metabolism (i.e. GAPDH, creatine kinase), as well as in Ca(2+) handling (i.e. parvalbumin, SERCA1). Western blot analysis, performed on some significantly changed proteins, validated the 2D results. Together these findings suggest AQP4 as a novel determinant in the regulation of skeletal muscle metabolism and better define the role of this water channel in skeletal muscle physiology.

  15. Absence of Aquaporin-4 in Skeletal Muscle Alters Proteins Involved in Bioenergetic Pathways and Calcium Handling

    PubMed Central

    Basco, Davide; Nicchia, Grazia Paola; D'Alessandro, Angelo; Zolla, Lello; Svelto, Maria; Frigeri, Antonio

    2011-01-01

    Aquaporin-4 (AQP4) is a water channel expressed at the sarcolemma of fast-twitch skeletal muscle fibers, whose expression is altered in several forms of muscular dystrophies. However, little is known concerning the physiological role of AQP4 in skeletal muscle and its functional and structural interaction with skeletal muscle proteome. Using AQP4-null mice, we analyzed the effect of the absence of AQP4 on the morphology and protein composition of sarcolemma as well as on the whole skeletal muscle proteome. Immunofluorescence analysis showed that the absence of AQP4 did not perturb the expression and cellular localization of the dystrophin-glycoprotein complex proteins, aside from those belonging to the extracellular matrix, and no alteration was found in sarcolemma integrity by dye extravasation assay. With the use of a 2DE-approach (BN/SDS-PAGE), protein maps revealed that in quadriceps, out of 300 Coomassie-blue detected and matched spots, 19 proteins exhibited changed expression in AQP4−/− compared to WT mice. In particular, comparison of the protein profiles revealed 12 up- and 7 down-regulated protein spots in AQP4−/− muscle. Protein identification by MS revealed that the perturbed expression pattern belongs to proteins involved in energy metabolism (i.e. GAPDH, creatine kinase), as well as in Ca2+ handling (i.e. parvalbumin, SERCA1). Western blot analysis, performed on some significantly changed proteins, validated the 2D results. Together these findings suggest AQP4 as a novel determinant in the regulation of skeletal muscle metabolism and better define the role of this water channel in skeletal muscle physiology. PMID:21552523

  16. SEORious business: structural proteins in sieve tubes and their involvement in sieve element occlusion.

    PubMed

    Knoblauch, Michael; Froelich, Daniel R; Pickard, William F; Peters, Winfried S

    2014-04-01

    The phloem provides a network of sieve tubes for long-distance translocation of photosynthates. For over a century, structural proteins in sieve tubes have presented a conundrum since they presumably increase the hydraulic resistance of the tubes while no potential function other than sieve tube or wound sealing in the case of injury has been suggested. Here we summarize and critically evaluate current speculations regarding the roles of these proteins. Our understanding suffers from the suggestive power of images; what looks like a sieve tube plug on micrographs may not actually impede translocation very much. Recent reports of an involvement of SEOR (sieve element occlusion-related) proteins, a class of P-proteins, in the sealing of injured sieve tubes are inconclusive; various lines of evidence suggest that, in neither intact nor injured plants, are SEORs determinative of translocation stoppage. Similarly, the popular notion that P-proteins serve in the defence against phloem sap-feeding insects is unsupported by empirical facts; it is conceivable that in functional sieve tubes, aphids actually could benefit from inducing a plug. The idea that rising cytosolic Ca(2+) generally triggers sieve tube blockage by P-proteins appears widely accepted, despite lacking experimental support. Even in forisomes, P-protein assemblages restricted to one single plant family and the only Ca(2+)-responsive P-proteins known, the available evidence does not unequivocally suggest that plug formation is the cause rather than a consequence of translocation stoppage. We conclude that the physiological roles of structural P-proteins remain elusive, and that in vivo studies of their dynamics in continuous sieve tube networks combined with flow velocity measurements will be required to (hopefully) resolve this scientific roadblock.

  17. Lacrimal gland PKC isoforms are differentially involved in agonist-induced protein secretion.

    PubMed

    Zoukhri, D; Hodges, R R; Sergheraert, C; Toker, A; Dartt, D A

    1997-01-01

    In the present study, we have synthesized and N-myristoylated peptides derived from the pseudosubstrate sequences of protein kinase C (PKC)-alpha, -delta, and -epsilon [Myr-PKC-alpha-(15-28), Myr-PKC-delta-(142-153), and Myr-PKC-epsilon-(149-164)], three isoforms present in rat lacrimal gland, and a peptide derived from the sequence of the endogenous inhibitor of protein kinase A [Myr-PKI-(17-25)]. Lacrimal gland acini were preincubated for 60 min with the myristoylated peptides (10(-10) to 3 x 10(-7) M), then protein secretion was stimulated with a phorbol ester, phorbol 12,13-dibutyrate (10(-6) M); vasoactive intestinal peptide (10(-8) M); a cholinergic agonist, carbachol (10(-5) M); or an alpha 1-adrenergic agonist, phenylephrine (10(-4) M), for 20 min. In intact lacrimal gland acini, Myr-PKC-alpha-(15-28) inhibited phorbol 12,13-dibutyrate-induced protein secretion. This effect was not reproduced by the acetylated peptide or by the myristoylated PKI, which inhibited vasoactive intestinal peptide-induced protein secretion, a response mediated by protein kinase A. Carbachol-induced protein secretion was inhibited by all three peptides. In contrast, phenylephrine-induced protein secretion was inhibited only by Myr-PKC-epsilon-(149-164), whereas Myr-PKC-alpha-(15-28) and Myr-PKC-delta-(142-153) had a stimulatory effect. None of these myristoylated peptides affected the calcium increase evoked by cholinergic or alpha 1-adrenergic agonists. We concluded that phorbol ester- and receptor-induced protein secretion involve different PKC isoforms in lacrimal gland.

  18. DNABP: Identification of DNA-Binding Proteins Based on Feature Selection Using a Random Forest and Predicting Binding Residues

    PubMed Central

    Guo, Jing; Sun, Xiao

    2016-01-01

    DNA-binding proteins are fundamentally important in cellular processes. Several computational-based methods have been developed to improve the prediction of DNA-binding proteins in previous years. However, insufficient work has been done on the prediction of DNA-binding proteins from protein sequence information. In this paper, a novel predictor, DNABP (DNA-binding proteins), was designed to predict DNA-binding proteins using the random forest (RF) classifier with a hybrid feature. The hybrid feature contains two types of novel sequence features, which reflect information about the conservation of physicochemical properties of the amino acids, and the binding propensity of DNA-binding residues and non-binding propensities of non-binding residues. The comparisons with each feature demonstrated that these two novel features contributed most to the improvement in predictive ability. Furthermore, to improve the prediction performance of the DNABP model, feature selection using the minimum redundancy maximum relevance (mRMR) method combined with incremental feature selection (IFS) was carried out during the model construction. The results showed that the DNABP model could achieve 86.90% accuracy, 83.76% sensitivity, 90.03% specificity and a Matthews correlation coefficient of 0.727. High prediction accuracy and performance comparisons with previous research suggested that DNABP could be a useful approach to identify DNA-binding proteins from sequence information. The DNABP web server system is freely available at http://www.cbi.seu.edu.cn/DNABP/. PMID:27907159

  19. A retroviral-derived peptide phosphorylates protein kinase D/protein kinase Cmu involving phospholipase C and protein kinase C.

    PubMed

    Luangwedchakarn, Voravich; Day, Noorbibi K; Hitchcock, Remi; Brown, Pam G; Lerner, Danica L; Rucker, Rajivi P; Cianciolo, George J; Good, Robert A; Haraguchi, Soichi

    2003-05-01

    CKS-17, a synthetic peptide representing a unique amino acid motif which is highly conserved in retroviral transmembrane proteins and other immunoregulatory proteins, induces selective immunomodulatory functions, both in vitro and in vivo, and activates intracellular signaling molecules such as cAMP and extracellular signal-regulated kinases. In the present study, using Jurkat T-cells, we report that CKS-17 phosphorylates protein kinase D (PKD)/protein kinase C (PKC) mu. Total cell extracts from CKS-17-stimulated Jurkat cells were immunoblotted with an anti-phospho-PKCmu antibody. The results show that CKS-17 significantly phosphorylates PKD/PKCmu in a dose- and time-dependent manner. Treatment of cells with the PKC inhibitors GF 109203X and Ro 31-8220, which do not act directly on PKD/PKCmu, attenuates CKS-17-induced phosphorylation of PKD/PKCmu. In contrast, the selective protein kinase A inhibitor H-89 does not reverse the action of CKS-17. Furthermore, a phospholipase C (PLC) selective inhibitor, U-73122, completely blocks the phosphorylation of PKD/PKCmu by CKS-17 while a negative control U-73343 does not. In addition, substitution of lysine for arginine residues in the CKS-17 sequence completely abrogates the ability of CKS-17 to phosphorylate PKD/PKCmu. These results clearly indicate that CKS-17 phosphorylates PKD/PKCmu through a PLC- and PKC-dependent mechanism and that arginine residues play an essential role in this activity of CKS-17, presenting a novel modality of the retroviral peptide CKS-17 and molecular interaction of this compound with target cells.

  20. Protein receptor for activated C kinase 1 is involved in morphine reward in mice.

    PubMed

    Wan, L; Su, L; Xie, Y; Liu, Y; Wang, Y; Wang, Z

    2009-07-07

    Opiate addiction is associated with upregulation of cAMP signaling in the brain. cAMP-responsive element binding protein (CREB), a nuclear transcription factor, is a downstream component of the extracellular signal-regulated protein kinase (ERK) pathway, which has been shown to regulate different physiological and psychological responses of drug addiction. RACK1, the protein receptor for activated C kinase 1, is a multifunctional scaffolding protein known to be a key regulator of various signaling cascades in the CNS. RACK1 functions specifically in integrin mediated activation of ERK cascade and targets active ERK. We examined if RACK1 is involved in the mechanism of drug addiction by regulating CREB in mouse hippocampus and prefrontal cortex. Several expressions were observed. Chronic administration of morphine made the expression of RACK1 and CREB mRNA increase in hippocampus and prefrontal cortex. The expression of RACK1 and CREB protein was strongly positive in CA1, CA3 and dentate gyrus (DG) of the hippocampus of morphine-treated mice brain, especially the pyramidal neurons in the DG of the hippocampus. Using the small interfering RNA technology, we determined that the expression of CREB mRNA was decreased in hippocampus and prefrontal cortex of morphine-treated mice. The expression of RACK1 and CREB protein was negative in CA1, CA3 and DG of hippocampus. These findings suggest that morphine reward can influence the expression of RACK1 in mouse hippocampus and prefrontal cortex through regulating CREB transcription.

  1. Proteomic analysis of male 4C germ cell proteins involved in mouse meiosis.

    PubMed

    Guo, Xuejiang; Zhang, Ping; Qi, Yujuan; Chen, Wen; Chen, Xiangxiang; Zhou, Zuomin; Sha, Jiahao

    2011-01-01

    Male meiosis is a specialized type of cell division that gives rise to sperm. Errors in this process can result in the generation of aneuploid gametes, which are associated with birth defects and infertility in humans. Until now, there has been a lack of a large-scale identification of proteins involved in male meiosis in mammals. In this study, we report the high-confidence identification of 3625 proteins in mouse male germ cells with 4C DNA content undergoing meiosis I. Of these, 397 were found to be testis specific. Bioinformatics analysis of the proteome led to the identification of 28 proteins known to be essential for male meiosis in mice. We also found 172 proteins that had yeast orthologs known to be essential for meiosis. Chromosome distribution analysis of the proteome showed underrepresentation of the identified proteins on the X chromosome, which may be due to meiotic sex chromosome inactivation. Characterization of the proteome of 4C germ cells from mouse testis provides an inventory of proteins, which is useful for understanding meiosis and the mechanisms of male infertility.

  2. Protein Sequence Comparison Based on Physicochemical Properties and the Position-Feature Energy Matrix

    PubMed Central

    Yu, Lulu; Zhang, Yusen; Gutman, Ivan; Shi, Yongtang; Dehmer, Matthias

    2017-01-01

    We develop a novel position-feature-based model for protein sequences by employing physicochemical properties of 20 amino acids and the measure of graph energy. The method puts the emphasis on sequence order information and describes local dynamic distributions of sequences, from which one can get a characteristic B-vector. Afterwards, we apply the relative entropy to the sequences representing B-vectors to measure their similarity/dissimilarity. The numerical results obtained in this study show that the proposed methods leads to meaningful results compared with competitors such as Clustal W. PMID:28393857

  3. The abi proteins and their involvement in bacteriocin self-immunity.

    PubMed

    Kjos, Morten; Snipen, Lars; Salehian, Zhian; Nes, Ingolf F; Diep, Dzung B

    2010-04-01

    The Abi protein family consists of putative membrane-bound metalloproteases. While they are involved in membrane anchoring of proteins in eukaryotes, little is known about their function in prokaryotes. In some known bacteriocin loci, Abi genes have been found downstream of bacteriocin structural genes (e.g., pln locus from Lactobacillus plantarum and sag locus from Streptococcus pyogenes), where they probably are involved in self-immunity. By modifying the profile hidden Markov model used to select Abi proteins in the Pfam protein family database, we show that this family is larger than presently recognized. Using bacteriocin-associated Abi genes as a means to search for novel bacteriocins in sequenced genomes, seven new bacteriocin-like loci were identified in Gram-positive bacteria. One such locus, from Lactobacillus sakei 23K, was selected for further experimental study, and it was confirmed that the bacteriocin-like genes (skkAB) exhibited antimicrobial activity when expressed in a heterologous host and that the associated Abi gene (skkI) conferred immunity against the cognate bacteriocin. Similar investigation of the Abi gene plnI and the Abi-like gene plnL from L. plantarum also confirmed their involvement in immunity to their cognate bacteriocins (PlnEF and PlnJK, respectively). Interestingly, the immunity genes from these three systems conferred a high degree of cross-immunity against each other's bacteriocins, suggesting the recognition of a common receptor. Site-directed mutagenesis demonstrated that the conserved motifs constituting the putative proteolytic active site of the Abi proteins are essential for the immunity function of SkkI, and to our knowledge, this represents a new concept in self-immunity.

  4. Hyperhomocysteinemia and bleomycin hydrolase modulate the expression of mouse brain proteins involved in neurodegeneration.

    PubMed

    Suszyńska-Zajczyk, Joanna; Luczak, Magdalena; Marczak, Lukasz; Jakubowski, Hieronim

    2014-01-01

    Homocysteine (Hcy) is a risk factor for Alzheimer's disease (AD). Bleomycin hydrolase (BLMH) participates in Hcy metabolism and is also linked to AD. The inactivation of the Blmh gene in mice causes accumulation of Hcy-thiolactone in the brain and increases susceptibility to Hcy-thiolactone-induced seizures. To gain insight into brain-related Blmh function, we used two-dimensional IEF/SDS-PAGE gel electrophoresis and MALDI-TOF/TOF mass spectrometry to examine brain proteomes of Blmh-/- mice and their Blmh+/+ littermates fed with a hyperhomocysteinemic high-Met or a control diet. We found that: (1) proteins involved in brain-specific function (Ncald, Nrgn, Stmn1, Stmn2), antioxidant defenses (Aop1), cell cycle (RhoGDI1, Ran), and cytoskeleton assembly (Tbcb, CapZa2) were differentially expressed in brains of Blmh-null mice; (2) hyperhomocysteinemia amplified effects of the Blmh-/- genotype on brain protein expression; (3) proteins involved in brain-specific function (Pebp1), antioxidant defenses (Sod1, Prdx2, DJ-1), energy metabolism (Atp5d, Ak1, Pgam-B), and iron metabolism (Fth) showed differential expression in Blmh-null brains only in hyperhomocysteinemic animals; (4) most proteins regulated by the Blmh-/- genotype were also regulated by high-Met diet, albeit in the opposite direction; and (5) the differentially expressed proteins play important roles in neural development, learning, plasticity, and aging and are linked to neurodegenerative diseases, including AD. Taken together, our findings suggest that Blmh interacts with diverse cellular processes from energy metabolism and anti-oxidative defenses to cell cycle, cytoskeleton dynamics, and synaptic plasticity essential for normal brain homeostasis and that modulation of these interactions by hyperhomocysteinemia underlies the involvement of Hcy in AD.

  5. Identification of proteins involved in Hg-Se antagonism in water hyacinth (Eichhornia crassipes).

    PubMed

    Pacheco, Pablo; Hanley, Traci; Figueroa, Julio A Landero

    2014-03-01

    Different studies have established the presence of a proteinaceus complex involved in Hg-Se agonism/antagonism in plants. In order to identify proteins involved in this mechanism, water hyacinth plants were divided into groups and supplemented with Hg, Se and a Hg-Se mixture. Proteins involved were identified through a screening separation by SEC-ICPMS followed by SAX-ICPMS and then peptide mapping of selected fractions by nanoLC-ESI-ITMS(2). Determination of total metal concentration showed that Se inhibits Hg translocation from roots to aerial compartments of the plant and that Se and Hg are antagonists to each other in terms of plant toxicity. In roots, stems and leaves Se was distributed mainly in two molecular mass fractions <670 kDa and ∼40 kDa, however, the proportion between these two fractions was inverted when Hg was co-administered. Hg throughout the plant was distributed in high and medium molecular mass compounds. Hg associated with molecules, ranging from <1.5 kDa to 15 kDa, was found in the root extract of Hg(ii) supplemented plants, but was absent in the root extract of Se(iv) and Hg(ii) supplemented plants. SAX showed that Hg and Se were mostly not associated with the same entity, since the complete overlapping of Hg and Se signals in all the peaks of SEC chromatograms was not observed. Changes in Se and Hg levels in water hyacinth were more evident in leaves in contrast to other compartments. Several proteins, possibly associated with either Se or Hg, were identified in roots, stems and leaves. Most of the identified proteins were associated with Hg and located in leaves, and these are associated specifically with chloroplast and mitochondria proteins, related to essential mechanisms in plants such as photosynthesis, carbon fixation and the electron transport chain.

  6. The SERRATE protein is involved in alternative splicing in Arabidopsis thaliana

    PubMed Central

    Raczynska, Katarzyna Dorota; Stepien, Agata; Kierzkowski, Daniel; Kalak, Malgorzata; Bajczyk, Mateusz; McNicol, Jim; Simpson, Craig G.; Szweykowska-Kulinska, Zofia; Brown, John W. S.; Jarmolowski, Artur

    2014-01-01

    How alternative splicing (AS) is regulated in plants has not yet been elucidated. Previously, we have shown that the nuclear cap-binding protein complex (AtCBC) is involved in AS in Arabidopsis thaliana. Here we show that both subunits of AtCBC (AtCBP20 and AtCBP80) interact with SERRATE (AtSE), a protein involved in the microRNA biogenesis pathway. Moreover, using a high-resolution reverse transcriptase-polymerase chain reaction AS system we have found that AtSE influences AS in a similar way to the cap-binding complex (CBC), preferentially affecting selection of 5′ splice site of first introns. The AtSE protein acts in cooperation with AtCBC: many changes observed in the mutant lacking the correct SERRATE activity were common to those observed in the cbp mutants. Interestingly, significant changes in AS of some genes were also observed in other mutants of plant microRNA biogenesis pathway, hyl1-2 and dcl1-7, but a majority of them did not correspond to the changes observed in the se-1 mutant. Thus, the role of SERRATE in AS regulation is distinct from that of HYL1 and DCL1, and is similar to the regulation of AS in which CBC is involved. PMID:24137006

  7. Diverse C-terminal sequences involved in Flavobacterium johnsoniae protein secretion.

    PubMed

    Kulkarni, Surashree S; Zhu, Yongtao; Brendel, Colton J; McBride, Mark J

    2017-04-10

    Flavobacterium johnsoniae and many related bacteria secrete proteins across the outer membrane using the type IX secretion system (T9SS). Proteins secreted by T9SSs have amino-terminal signal peptides for export across the cytoplasmic membrane by the Sec system and carboxy-terminal domains (CTDs) targeting them for secretion across the outer membrane by the T9SS. Most but not all T9SS CTDs belong to family TIGR04183 (type-A CTDs). We functionally characterized diverse CTDs for secretion by the F. johnsoniae T9SS. Attachment of the CTDs from F. johnsoniae RemA, AmyB, and ChiA to the foreign protein sfGFP that had a signal peptide at the amino terminus resulted in secretion across the outer membrane. In each case approximately 80 to 100 amino acids from the extreme carboxy-termini was needed for efficient secretion. Several type-A CTDs from distantly related members of the phylum Bacteroidetes functioned in F. johnsoniae, supporting secretion of sfGFP by the F. johnsoniae T9SS. F. johnsoniae SprB requires the T9SS for secretion but lacks a type-A CTD. It has a conserved C-terminal domain belonging to family TIGR04131, which we refer to as a type-B CTD. The CTD of SprB was required for its secretion, but attachment of C-terminal regions of SprB of up to 1182 amino acids to sfGFP failed to result in secretion. Additional features outside of the C-terminal region of SprB may be required for its secretion.Importance Type IX protein secretion systems (T9SSs) are common in but limited to members of the phylum Bacteroidetes Most proteins that are secreted by T9SSs have conserved carboxy-terminal domains that belong to either protein domain family TIGR04183 (type-A CTDs) or TIGR04131 (type-B CTDs). Here we identify features of T9SS CTDs of F. johnsoniae that are required for protein secretion and demonstrate that type-A CTDs from distantly related members of the phylum function with the F. johnsoniae T9SS to secrete the foreign protein sfGFP. In contrast, type-B CTDs failed

  8. Two Neuronal G Proteins Are Involved in Chemosensation of the Caenorhabditis Elegans Dauer-Inducing Pheromone

    PubMed Central

    Zwaal, R. R.; Mendel, J. E.; Sternberg, P. W.; Plasterk, RHA.

    1997-01-01

    Caenorhabditis elegans uses chemosensation to determine its course of development. Young larvae can arrest as dauer larvae in response to increasing population density, which they measure by a nematode-excreted pheromone, and decreasing food supply. Dauer larvae can resume development in response to a decrease in pheromone and increase in food concentration. We show here that two novel G protein alpha subunits (GPA-2 and GPA-3) show promoter activity in subsets of chemosensory neurons and are involved in the decision to form dauer larvae primarily through the response to dauer pheromone. Dominant activating mutations in these G proteins result in constitutive, pheromone-independent dauer formation, whereas inactivation results in reduced sensitivity to pheromone, and, under certain conditions, an alteration in the response to food. Interactions between gpa-2, gpa-3 and other genes controlling dauer formation suggest that these G proteins may act in parallel to regulate the neuronal decision making that precedes dauer formation. PMID:9055081

  9. The ESCRT-II proteins are involved in shaping the sarcoplasmic reticulum in C. elegans.

    PubMed

    Lefebvre, Christophe; Largeau, Céline; Michelet, Xavier; Fourrage, Cécile; Maniere, Xavier; Matic, Ivan; Legouis, Renaud; Culetto, Emmanuel

    2016-04-01

    The sarcoplasmic reticulum is a network of tubules and cisternae localized in close association with the contractile apparatus, and regulates Ca(2+)dynamics within striated muscle cell. The sarcoplasmic reticulum maintains its shape and organization despite repeated muscle cell contractions, through mechanisms which are still under investigation. The ESCRT complexes are essential to organize membrane subdomains and modify membrane topology in multiple cellular processes. Here, we report for the first time that ESCRT-II proteins play a role in the maintenance of sarcoplasmic reticulum integrity inC. elegans ESCRT-II proteins colocalize with the sarcoplasmic reticulum marker ryanodine receptor UNC-68. The localization at the sarcoplasmic reticulum of ESCRT-II and UNC-68 are mutually dependent. Furthermore, the characterization of ESCRT-II mutants revealed a fragmentation of the sarcoplasmic reticulum network, associated with an alteration of Ca(2+)dynamics. Our data provide evidence that ESCRT-II proteins are involved in sarcoplasmic reticulum shaping.

  10. Involvement of dietary bioactive proteins and peptides in autism spectrum disorders.

    PubMed

    Siniscalco, Dario; Antonucci, Nicola

    2013-12-01

    Autism and autism spectrum disorders (ASDs) are heterogeneous, severe neurodevelopmental pathologies. These enigmatic conditions have their origins in the interaction of multiple genes and environmental factors. Dysfunctions in social interactions and communication skills, restricted interests, repetitive and stereotypic verbal and non-verbal behaviours are the main core symptoms. Several biochemical processes are associated with ASDs: oxidative stress; endoplasmic reticulum stress; decreased methylation capacity; limited production of glutathione; mitochondrial dysfunction; intestinal impaired permeability and dysbiosis; increased toxic metal burden; immune dysregulation. Current available treatments for ASDs can be divided into behavioural, nutritional and medical approaches, although no defined standard approach exists. Dietary bioactive proteins and peptides show potential for application as health-promoting agents. Nowadays, increasing studies highlight a key role of bioactive proteins and peptides in ASDs. This review will focus on the state-of-the-art regarding the involvement of dietary bioactive proteins and peptides in ASDs. Identification of novel therapeutic targets for ASD management will be also discussed.

  11. Involvement of F-BOX proteins in progression and development of human malignancies.

    PubMed

    Uddin, Shahab; Bhat, Ajaz A; Krishnankutty, Roopesh; Mir, Fayaz; Kulinski, Michal; Mohammad, Ramzi M

    2016-02-01

    The Ubiquitin Proteasome System (UPS) is a core regulator with various protein components (ubiquitin-activating E1 enzymes, ubiquitin-conjugating E2 enzymes, ubiquitin-protein E3 ligases, and the 26S proteasome) which work together in a coordinated fashion to ensure the appropriate and efficient proteolysis of target substrates. E3 ubiquitin ligases are essential components of the UPS machinery, working with E1 and E2 enzymes to bind substrates and assist the transport of ubiquitin molecules onto the target protein. As the UPS controls the degradation of several oncogenes and tumor suppressors, dysregulation of this pathway leads to several human malignancies. A major category of E3 Ub ligases, the SCF (Skp-Cullin-F-box) complex, is composed of four principal components: Skp1, Cul1/Cdc53, Roc1/Rbx1/Hrt1, and an F-box protein (FBP). FBPs are the substrate recognition components of SCF complexes and function as adaptors that bring substrates into physical proximity with the rest of the SCF. Besides acting as a component of SCF complexes, FBPs are involved in DNA replication, transcription, cell differentiation and cell death. This review will highlight the recent literature on three well characterized FBPs SKP2, Fbw7, and beta-TRCP. In particular, we will focus on the involvement of these deregulated FBPs in the progression and development of various human cancers. We will also highlight some novel substrates recently identified for these FBPs.

  12. Involvement of regucalcin as a suppressor protein in human carcinogenesis: insight into the gene therapy.

    PubMed

    Yamaguchi, Masayoshi

    2015-08-01

    Regucalcin, which its gene is located on the X chromosome, plays a multifunctional role as a suppressor protein in cell signal transduction in various types of cells and tissues. The suppression of regucalcin gene expression has been shown to involve in carcinogenesis. Regucalcin gene expression was uniquely downregulated in carcinogenesis of rat liver in vivo, although the expression of other many genes was upregulated, indicating that endogenous regucalcin plays a suppressive role in the development of hepatocarcinogenesis. Overexpression of endogenous regucalcin was found to suppress proliferation of rat cloned hepatoma cells in vitro. Moreover, the regucalcin gene and its protein levels were demonstrated specifically to downregulate in human hepatocellular carcinoma by analysis with multiple gene expression profiles and proteomics. Regucalcin gene expression was also found to suppress in human tumor tissues including kidney, lung, brain, breast and prostate, suggesting that repressed regucalcin gene expression leads to the development of carcinogenesis in various tissues. Regucalcin may play a role as a suppressor protein in carcinogenesis. Overexpression of endogenous regucalcin is suggested to reveal preventive and therapeutic effects on carcinogenesis. Delivery of the regucalcin gene may be a novel useful tool in the gene therapy of carcinogenesis. This review will discuss regarding to an involvement of regucalcin as a suppressor protein in human carcinogenesis in insight into the gene therapy.

  13. NAP-1, Nucleosome assembly protein 1, a histone chaperone involved in Drosophila telomeres.

    PubMed

    López-Panadès, Elisenda; Casacuberta, Elena

    2016-03-01

    Telomere elongation is a function that all eukaryote cells must accomplish in order to guarantee, first, the stability of the end of the chromosomes and second, to protect the genetic information from the inevitable terminal erosion. The targeted transposition of the telomere transposons HeT-A, TART and TAHRE perform this function in Drosophila, while the telomerase mechanism elongates the telomeres in most eukaryotes. In order to integrate telomere maintenance together with cell cycle and metabolism, different components of the cell interact, regulate, and control the proteins involved in telomere elongation. Different partners of the telomerase mechanism have already been described, but in contrast, very few proteins have been related with assisting the telomere transposons of Drosophila. Here, we describe for the first time, the implication of NAP-1 (Nucleosome assembly protein 1), a histone chaperone that has been involved in nuclear transport, transcription regulation, and chromatin remodeling, in telomere biology. We find that Nap-1 and HeT-A Gag, one of the major components of the Drosophila telomeres, are part of the same protein complex. We also demonstrate that their close interaction is necessary to guarantee telomere stability in dividing cells. We further show that NAP-1 regulates the transcription of the HeT-A retrotransposon, pointing to a positive regulatory role of NAP-1 in telomere expression. All these results facilitate the understanding of the transposon telomere maintenance mechanism, as well as the integration of telomere biology with the rest of the cell metabolism.

  14. CUP-1 Is a Novel Protein Involved in Dietary Cholesterol Uptake in Caenorhabditis elegans

    PubMed Central

    Valdes, Victor J.; Athie, Alejandro; Salinas, Laura S.; Navarro, Rosa E.; Vaca, Luis

    2012-01-01

    Sterols transport and distribution are essential processes in all multicellular organisms. Survival of the nematode Caenorhabditis elegans depends on dietary absorption of sterols present in the environment. However the general mechanisms associated to sterol uptake in nematodes are poorly understood. In the present work we provide evidence showing that a previously uncharacterized transmembrane protein, designated Cholesterol Uptake Protein-1 (CUP-1), is involved in dietary cholesterol uptake in C. elegans. Animals lacking CUP-1 showed hypersensitivity to cholesterol limitation and were unable to uptake cholesterol. A CUP-1-GFP fusion protein colocalized with cholesterol-rich vesicles, endosomes and lysosomes as well as the plasma membrane. Additionally, by FRET imaging, a direct interaction was found between the cholesterol analog DHE and the transmembrane “cholesterol recognition/interaction amino acid consensus” (CRAC) motif present in C. elegans CUP-1. In-silico analysis identified two mammalian homologues of CUP-1. Most interestingly, CRAC motifs are conserved in mammalian CUP-1 homologous. Our results suggest a role of CUP-1 in cholesterol uptake in C. elegans and open up the possibility for the existence of a new class of proteins involved in sterol absorption in mammals. PMID:22479487

  15. Enhanced detection of lipid transfer inhibitor protein activity by an assay involving only low density lipoprotein.

    PubMed

    Morton, R E; Greene, D J

    1994-11-01

    Lipid transfer inhibitor protein (LTIP) activity has been typically quantitated by its ability to suppress lipid transfer protein-mediated lipid movement between low density lipoprotein (LDL) and high density lipoprotein (HDL). In an attempt to establish an LTIP activity assay that is more sensitive, we have exploited the reported preference of the inhibitor protein to interact with LDL. A lipid transfer assay was established that involves LDL as both the donor and the acceptor; LDL in one of these two pools was biotinylated to facilitate its removal with immobilized avidin. Compared to the standard LDL to HDL assay, LTIP inhibited lipid transfer from radiolabeled LDL to biotin-LDL 7-fold more. In the absence of LTIP, lipid transfer activity was the same in both assays. An added benefit of this assay was the near linearity (up to 85%) of the inhibitory response, in contrast to the highly curvilinear response of LTIP in LDL to HDL transfer assays. The high sensitivity of the LDL to biotin-LDL transfer assay in measuring LTIP activity could not be duplicated by other transfer assays including assays containing only HDL (HDL to biotin-HDL), assays between liposomes and LDL, or assays between LDL and HDL where the concentration of lipoproteins was reduced 10-fold. Thus, LTIP activity is most effectively measured in homologous lipid transfer assays involving only LDL (and its biotin derivative). This increased sensitivity to LTIP suggests that the inhibitor binds more avidly to the LDL surface than does lipid transfer protein.

  16. Mild copper deficiency alters gene expression of proteins involved in iron metabolism.

    PubMed

    Auclair, Sylvain; Feillet-Coudray, Christine; Coudray, Charles; Schneider, Susanne; Muckenthaler, Martina U; Mazur, Andrzej

    2006-01-01

    Iron and copper homeostasis share common proteins and are therefore closely linked to each other. For example, copper-containing proteins like ceruloplasmin and hephaestin oxidize Fe(2+) during cellular export processes for transport in the circulation bound to transferrin. Indeed, copper deficiency provokes iron metabolism disorders leading to anemia and liver iron accumulation. The aim of the present work was to understand the cross-talk between copper status and iron metabolism. For this purpose we have established dietary copper deficiency in C57BL6 male mice during twelve weeks. Hematological parameters, copper and iron status were evaluated. cDNA microarray studies were performed to investigate gene expression profiles of proteins involved in iron metabolism in the liver, duodenum and spleen. Our results showed that copper deficiency induces microcytic and hypochromic anemia as well as liver iron overload. Gene expression profiles, however, indicate that hepatic and intestinal mRNA expression neither compensates for hepatic iron overload nor the anemia observed in this mouse model. Instead, major modifications of gene expression occurred in the spleen. We observed increased mRNA levels of the transferrin receptors 1 and 2 and of several proteins involved in the heme biosynthesis pathway (ferrochelatase, UroD, UroS,...). These results suggest that copper-deficient mice respond to the deficiency induced anemia by an adaptation leading to an increase in erythrocyte synthesis.

  17. Short-Time Glassy-like Dynamics Observed in Viscous Protein Solutions with Competing Potential Features

    NASA Astrophysics Data System (ADS)

    Wagner, Norman; Godfrin, Doug; Liu, Yun

    Structures in concentrated protein solutions caused by the combination of short-range attraction (SA) and long-range repulsion (LR) have been extensively studied due to their importance in understanding therapeutic protein formulations and the phase behavior in general. Despite extensive studies of kinetically arrested states in colloidal systems with short-range attraction, less is understood for the effect of an additional longer-range repulsion on model colloidal systems with a SA interaction. Highly purified lysozyme is used a model experimental system due to its stable globular structure and SALR interactions at low ionic strength that can be quantitatively modeled. The fluid microstructure and protein short time self diffusion are measured across a broad range of conditions by small angle neutron scattering (SANS) and neutron spin echo (NSE), respectively. Newtonian liquid behavior is observed at all concentrations, even with an increase of zero shear viscosity by almost four orders of magnitude with increasing concentration. However, dynamic measurements demonstrate a sub-diffusive regime at relatively short time scales for concentrated samples at low temperature. The formation of a heterogeneous density distribution is shown to produce localized regions of high density that reduce protein motion, giving it a glassy-like behavior at the short time scale. This heterogeneity occurs at the length scale associated with the intermediate range order driven by the competing potential features, distinguishable from heterogeneous colloidal gels.

  18. The endocytic adaptor protein ARH associates with motor and centrosomal proteins and is involved in centrosome assembly and cytokinesis.

    PubMed

    Lehtonen, Sanna; Shah, Mehul; Nielsen, Rikke; Iino, Noriaki; Ryan, Jennifer J; Zhou, Huilin; Farquhar, Marilyn G

    2008-07-01

    Numerous proteins involved in endocytosis at the plasma membrane have been shown to be present at novel intracellular locations and to have previously unrecognized functions. ARH (autosomal recessive hypercholesterolemia) is an endocytic clathrin-associated adaptor protein that sorts members of the LDL receptor superfamily (LDLR, megalin, LRP). We report here that ARH also associates with centrosomes in several cell types. ARH interacts with centrosomal (gamma-tubulin and GPC2 and GPC3) and motor (dynein heavy and intermediate chains) proteins. ARH cofractionates with gamma-tubulin on isolated centrosomes, and gamma-tubulin and ARH interact on isolated membrane vesicles. During mitosis, ARH sequentially localizes to the nuclear membrane, kinetochores, spindle poles and the midbody. Arh(-/-) embryonic fibroblasts (MEFs) show smaller or absent centrosomes suggesting ARH plays a role in centrosome assembly. Rat-1 fibroblasts depleted of ARH by siRNA and Arh(-/-) MEFs exhibit a slower rate of growth and prolonged cytokinesis. Taken together the data suggest that the defects in centrosome assembly in ARH depleted cells may give rise to cell cycle and mitotic/cytokinesis defects. We propose that ARH participates in centrosomal and mitotic dynamics by interacting with centrosomal proteins. Whether the centrosomal and mitotic functions of ARH are related to its endocytic role remains to be established.

  19. Protein phosphatase and kinase activities possibly involved in exocytosis regulation in Paramecium tetraurelia.

    PubMed

    Kissmehl, R; Treptau, T; Hofer, H W; Plattner, H

    1996-07-01

    In Paramecium tetraurelia cells synchronous exocytosis induced by aminoethyldextran (AED) is accompanied by an equally rapid dephosphorylation of a 63 kDa phosphoprotein (PP63) within 80 ms. In vivo, rephosphorylation occurs within a few seconds after AED triggering. In homogenates (P)P63 can be solubilized in all three phosphorylation states (phosphorylated, dephosphorylated and rephosphorylated) and thus tested in vitro. By using chelators of different divalent cations, de- and rephosphorylation of PP63 and P63 respectively can be achieved by an endogenous protein phosphatase/kinase system. Dephosphorylation occurs in the presence of EDTA, whereas in the presence of EGTA this was concealed by phosphorylation by endogenous kinase(s), thus indicating that phosphorylation of P63 is calcium-independent. Results obtained with protein phosphatase inhibitors (okadaic acid, calyculin A) allowed us to exclude a protein serine/threonine phosphatase of type I (with selective sensitivity in Paramecium). Protein phosphatase 2C is also less likely to be a candidate because of its requirement for high Mg2+ concentrations. According to previous evidence a protein serine/threonine phosphatase of type 2B (calcineurin; CaN) is possibly involved. We have now found that bovine brain CaN dephosphorylates PP63 in vitro. Taking into account the specific requirements of this phosphatase in vitro, with p-nitrophenyl phosphate as a substrate, we have isolated a cytosolic phosphatase of similar characteristics by combined preparative gel electrophoresis and affinity-column chromatography. In Paramecium this phosphatase also dephosphorylates PP63 in vitro (after 32P labelling in vivo). Using various combinations of ion exchange, affinity and hydrophobic interaction chromatography we have also isolated three different protein kinases from the soluble fraction, i.e. a cAMP-dependent protein kinase (PKA), a cGMP-dependent protein kinase (PKG) and a casein kinase. Among the kinases tested, PKA

  20. Origin and spread of photosynthesis based upon conserved sequence features in key bacteriochlorophyll biosynthesis proteins.

    PubMed

    Gupta, Radhey S

    2012-11-01

    The origin of photosynthesis and how this capability has spread to other bacterial phyla remain important unresolved questions. I describe here a number of conserved signature indels (CSIs) in key proteins involved in bacteriochlorophyll (Bchl) biosynthesis that provide important insights in these regards. The proteins BchL and BchX, which are essential for Bchl biosynthesis, are derived by gene duplication in a common ancestor of all phototrophs. More ancient gene duplication gave rise to the BchX-BchL proteins and the NifH protein of the nitrogenase complex. The sequence alignment of NifH-BchX-BchL proteins contain two CSIs that are uniquely shared by all NifH and BchX homologs, but not by any BchL homologs. These CSIs and phylogenetic analysis of NifH-BchX-BchL protein sequences strongly suggest that the BchX homologs are ancestral to BchL and that the Bchl-based anoxygenic photosynthesis originated prior to the chlorophyll (Chl)-based photosynthesis in cyanobacteria. Another CSI in the BchX-BchL sequence alignment that is uniquely shared by all BchX homologs and the BchL sequences from Heliobacteriaceae, but absent in all other BchL homologs, suggests that the BchL homologs from Heliobacteriaceae are primitive in comparison to all other photosynthetic lineages. Several other identified CSIs in the BchN homologs are commonly shared by all proteobacterial homologs and a clade consisting of the marine unicellular Cyanobacteria (Clade C). These CSIs in conjunction with the results of phylogenetic analyses and pair-wise sequence similarity on the BchL, BchN, and BchB proteins, where the homologs from Clade C Cyanobacteria and Proteobacteria exhibited close relationship, provide strong evidence that these two groups have incurred lateral gene transfers. Additionally, phylogenetic analyses and several CSIs in the BchL-N-B proteins that are uniquely shared by all Chlorobi and Chloroflexi homologs provide evidence that the genes for these proteins have also been

  1. Identification and characterization of plastid-type proteins from sequence-attributed features using machine learning

    PubMed Central

    2013-01-01

    Background Plastids are an important component of plant cells, being the site of manufacture and storage of chemical compounds used by the cell, and contain pigments such as those used in photosynthesis, starch synthesis/storage, cell color etc. They are essential organelles of the plant cell, also present in algae. Recent advances in genomic technology and sequencing efforts is generating a huge amount of DNA sequence data every day. The predicted proteome of these genomes needs annotation at a faster pace. In view of this, one such annotation need is to develop an automated system that can distinguish between plastid and non-plastid proteins accurately, and further classify plastid-types based on their functionality. We compared the amino acid compositions of plastid proteins with those of non-plastid ones and found significant differences, which were used as a basis to develop various feature-based prediction models using similarity-search and machine learning. Results In this study, we developed separate Support Vector Machine (SVM) trained classifiers for characterizing the plastids in two steps: first distinguishing the plastid vs. non-plastid proteins, and then classifying the identified plastids into their various types based on their function (chloroplast, chromoplast, etioplast, and amyloplast). Five diverse protein features: amino acid composition, dipeptide composition, the pseudo amino acid composition, Nterminal-Center-Cterminal composition and the protein physicochemical properties are used to develop SVM models. Overall, the dipeptide composition-based module shows the best performance with an accuracy of 86.80% and Matthews Correlation Coefficient (MCC) of 0.74 in phase-I and 78.60% with a MCC of 0.44 in phase-II. On independent test data, this model also performs better with an overall accuracy of 76.58% and 74.97% in phase-I and phase-II, respectively. The similarity-based PSI-BLAST module shows very low performance with about 50% prediction

  2. TargetCrys: protein crystallization prediction by fusing multi-view features with two-layered SVM.

    PubMed

    Hu, Jun; Han, Ke; Li, Yang; Yang, Jing-Yu; Shen, Hong-Bin; Yu, Dong-Jun

    2016-11-01

    The accurate prediction of whether a protein will crystallize plays a crucial role in improving the success rate of protein crystallization projects. A common critical problem in the development of machine-learning-based protein crystallization predictors is how to effectively utilize protein features extracted from different views. In this study, we aimed to improve the efficiency of fusing multi-view protein features by proposing a new two-layered SVM (2L-SVM) which switches the feature-level fusion problem to a decision-level fusion problem: the SVMs in the 1st layer of the 2L-SVM are trained on each of the multi-view feature sets; then, the outputs of the 1st layer SVMs, which are the "intermediate" decisions made based on the respective feature sets, are further ensembled by a 2nd layer SVM. Based on the proposed 2L-SVM, we implemented a sequence-based protein crystallization predictor called TargetCrys. Experimental results on several benchmark datasets demonstrated the efficacy of the proposed 2L-SVM for fusing multi-view features. We also compared TargetCrys with existing sequence-based protein crystallization predictors and demonstrated that the proposed TargetCrys outperformed most of the existing predictors and is competitive with the state-of-the-art predictors. The TargetCrys webserver and datasets used in this study are freely available for academic use at: http://csbio.njust.edu.cn/bioinf/TargetCrys .

  3. Quantitative Description of a Protein Fitness Landscape Based on Molecular Features

    PubMed Central

    Meini, María-Rocío; Tomatis, Pablo E.; Weinreich, Daniel M.; Vila, Alejandro J.

    2015-01-01

    Understanding the driving forces behind protein evolution requires the ability to correlate the molecular impact of mutations with organismal fitness. To address this issue, we employ here metallo-β-lactamases as a model system, which are Zn(II) dependent enzymes that mediate antibiotic resistance. We present a study of all the possible evolutionary pathways leading to a metallo-β-lactamase variant optimized by directed evolution. By studying the activity, stability and Zn(II) binding capabilities of all mutants in the preferred evolutionary pathways, we show that this local fitness landscape is strongly conditioned by epistatic interactions arising from the pleiotropic effect of mutations in the different molecular features of the enzyme. Activity and stability assays in purified enzymes do not provide explanatory power. Instead, measurement of these molecular features in an environment resembling the native one provides an accurate description of the observed antibiotic resistance profile. We report that optimization of Zn(II) binding abilities of metallo-β-lactamases during evolution is more critical than stabilization of the protein to enhance fitness. A global analysis of these parameters allows us to connect genotype with fitness based on quantitative biochemical and biophysical parameters. PMID:25767204

  4. Quantitative Description of a Protein Fitness Landscape Based on Molecular Features.

    PubMed

    Meini, María-Rocío; Tomatis, Pablo E; Weinreich, Daniel M; Vila, Alejandro J

    2015-07-01

    Understanding the driving forces behind protein evolution requires the ability to correlate the molecular impact of mutations with organismal fitness. To address this issue, we employ here metallo-β-lactamases as a model system, which are Zn(II) dependent enzymes that mediate antibiotic resistance. We present a study of all the possible evolutionary pathways leading to a metallo-β-lactamase variant optimized by directed evolution. By studying the activity, stability and Zn(II) binding capabilities of all mutants in the preferred evolutionary pathways, we show that this local fitness landscape is strongly conditioned by epistatic interactions arising from the pleiotropic effect of mutations in the different molecular features of the enzyme. Activity and stability assays in purified enzymes do not provide explanatory power. Instead, measurement of these molecular features in an environment resembling the native one provides an accurate description of the observed antibiotic resistance profile. We report that optimization of Zn(II) binding abilities of metallo-β-lactamases during evolution is more critical than stabilization of the protein to enhance fitness. A global analysis of these parameters allows us to connect genotype with fitness based on quantitative biochemical and biophysical parameters.

  5. A Novel RNA-Binding Protein Involves ABA Signaling by Post-transcriptionally Repressing ABI2

    PubMed Central

    Xu, Jianwen; Chen, Yihan; Qian, Luofeng; Mu, Rong; Yuan, Xi; Fang, Huimin; Huang, Xi; Xu, Enshun; Zhang, Hongsheng; Huang, Ji

    2017-01-01

    The Stress Associated RNA-binding protein 1 (SRP1) repressed by ABA, salt and cold encodes a C2C2-type zinc finger protein in Arabidopsis. The knock-out mutation in srp1 reduced the sensitivity of seed to ABA and salt stress during germination and post-germinative growth stages. In contrast, SRP1-overexpressing seedlings were more sensitive to ABA and salt compared to wild type plants. In the presence of ABA, the transcript levels of ABA signaling and germination-related genes including ABI3. ABI5. EM1 and EM6 were less induced in srp1 compared to WT. Interestingly, expression of ABI2 encoding a protein phosphatase 2C protein were significantly up-regulated in srp1 mutants. By in vitro analysis, SRP1 was identified as a novel RNA-binding protein directly binding to 3′UTR of ABI2 mRNA. Moreover, transient expression assay proved the function of SRP1 in reducing the activity of luciferase whose coding sequence was fused with the ABI2 3’UTR. Together, it is suggested that SRP1 is involved in the ABA signaling by post-transcriptionally repressing ABI2 expression in Arabidopsis. PMID:28174577

  6. New proteins involved in sulfur trafficking in the cytoplasm of Allochromatium vinosum.

    PubMed

    Stockdreher, Yvonne; Sturm, Marga; Josten, Michaele; Sahl, Hans-Georg; Dobler, Nadine; Zigann, Renate; Dahl, Christiane

    2014-05-02

    The formation of periplasmic sulfur globules is an intermediate step during the oxidation of reduced sulfur compounds in various sulfur-oxidizing microorganisms. The mechanism of how this sulfur is activated and crosses the cytoplasmic membrane for further oxidation to sulfite by the dissimilatory reductase DsrAB is incompletely understood, but it has been well documented that the pathway involves sulfur trafficking mediated by sulfur-carrying proteins. So far sulfur transfer from DsrEFH to DsrC has been established. Persulfurated DsrC very probably serves as a direct substrate for DsrAB. Here, we introduce further important players in oxidative sulfur metabolism; the proteins Rhd_2599, TusA, and DsrE2 are strictly conserved in the Chromatiaceae, Chlorobiaceae, and Acidithiobacillaceae families of sulfur-oxidizing bacteria and are linked to genes encoding complexes involved in sulfur oxidation (Dsr or Hdr) in the latter two. Here we show via relative quantitative real-time PCR and microarray analysis an increase of mRNA levels under sulfur-oxidizing conditions for rhd_2599, tusA, and dsrE2 in Allochromatium vinosum. Transcriptomic patterns for the three genes match those of major genes for the sulfur-oxidizing machinery rather than those involved in biosynthesis of sulfur-containing biomolecules. TusA appears to be one of the major proteins in A. vinosum. A rhd_2599-tusA-dsrE2-deficient mutant strain, although not viable in liquid culture, was clearly sulfur oxidation negative upon growth on solid media containing sulfide. Rhd_2599, TusA, and DsrE2 bind sulfur atoms via conserved cysteine residues, and experimental evidence is provided for the transfer of sulfur between these proteins as well as to DsrEFH and DsrC.

  7. Protein-ligand binding region prediction (PLB-SAVE) based on geometric features and CUDA acceleration

    PubMed Central

    2013-01-01

    Background Protein-ligand interactions are key processes in triggering and controlling biological functions within cells. Prediction of protein binding regions on the protein surface assists in understanding the mechanisms and principles of molecular recognition. In silico geometrical shape analysis plays a primary step in analyzing the spatial characteristics of protein binding regions and facilitates applications of bioinformatics in drug discovery and design. Here, we describe the novel software, PLB-SAVE, which uses parallel processing technology and is ideally suited to extract the geometrical construct of solid angles from surface atoms. Representative clusters and corresponding anchors were identified from all surface elements and were assigned according to the ranking of their solid angles. In addition, cavity depth indicators were obtained by proportional transformation of solid angles and cavity volumes were calculated by scanning multiple directional vectors within each selected cavity. Both depth and volume characteristics were combined with various weighting coefficients to rank predicted potential binding regions. Results Two test datasets from LigASite, each containing 388 bound and unbound structures, were used to predict binding regions using PLB-SAVE and two well-known prediction systems, SiteHound and MetaPocket2.0 (MPK2). PLB-SAVE outperformed the other programs with accuracy rates of 94.3% for unbound proteins and 95.5% for bound proteins via a tenfold cross-validation process. Additionally, because the parallel processing architecture was designed to enhance the computational efficiency, we obtained an average of 160-fold increase in computational time. Conclusions In silico binding region prediction is considered the initial stage in structure-based drug design. To improve the efficacy of biological experiments for drug development, we developed PLB-SAVE, which uses only geometrical features of proteins and achieves a good overall performance

  8. A deep learning framework for modeling structural features of RNA-binding protein targets

    PubMed Central

    Zhang, Sai; Zhou, Jingtian; Hu, Hailin; Gong, Haipeng; Chen, Ligong; Cheng, Chao; Zeng, Jianyang

    2016-01-01

    RNA-binding proteins (RBPs) play important roles in the post-transcriptional control of RNAs. Identifying RBP binding sites and characterizing RBP binding preferences are key steps toward understanding the basic mechanisms of the post-transcriptional gene regulation. Though numerous computational methods have been developed for modeling RBP binding preferences, discovering a complete structural representation of the RBP targets by integrating their available structural features in all three dimensions is still a challenging task. In this paper, we develop a general and flexible deep learning framework for modeling structural binding preferences and predicting binding sites of RBPs, which takes (predicted) RNA tertiary structural information into account for the first time. Our framework constructs a unified representation that characterizes the structural specificities of RBP targets in all three dimensions, which can be further used to predict novel candidate binding sites and discover potential binding motifs. Through testing on the real CLIP-seq datasets, we have demonstrated that our deep learning framework can automatically extract effective hidden structural features from the encoded raw sequence and structural profiles, and predict accurate RBP binding sites. In addition, we have conducted the first study to show that integrating the additional RNA tertiary structural features can improve the model performance in predicting RBP binding sites, especially for the polypyrimidine tract-binding protein (PTB), which also provides a new evidence to support the view that RBPs may own specific tertiary structural binding preferences. In particular, the tests on the internal ribosome entry site (IRES) segments yield satisfiable results with experimental support from the literature and further demonstrate the necessity of incorporating RNA tertiary structural information into the prediction model. The source code of our approach can be found in https

  9. A deep learning framework for modeling structural features of RNA-binding protein targets.

    PubMed

    Zhang, Sai; Zhou, Jingtian; Hu, Hailin; Gong, Haipeng; Chen, Ligong; Cheng, Chao; Zeng, Jianyang

    2016-02-29

    RNA-binding proteins (RBPs) play important roles in the post-transcriptional control of RNAs. Identifying RBP binding sites and characterizing RBP binding preferences are key steps toward understanding the basic mechanisms of the post-transcriptional gene regulation. Though numerous computational methods have been developed for modeling RBP binding preferences, discovering a complete structural representation of the RBP targets by integrating their available structural features in all three dimensions is still a challenging task. In this paper, we develop a general and flexible deep learning framework for modeling structural binding preferences and predicting binding sites of RBPs, which takes (predicted) RNA tertiary structural information into account for the first time. Our framework constructs a unified representation that characterizes the structural specificities of RBP targets in all three dimensions, which can be further used to predict novel candidate binding sites and discover potential binding motifs. Through testing on the real CLIP-seq datasets, we have demonstrated that our deep learning framework can automatically extract effective hidden structural features from the encoded raw sequence and structural profiles, and predict accurate RBP binding sites. In addition, we have conducted the first study to show that integrating the additional RNA tertiary structural features can improve the model performance in predicting RBP binding sites, especially for the polypyrimidine tract-binding protein (PTB), which also provides a new evidence to support the view that RBPs may own specific tertiary structural binding preferences. In particular, the tests on the internal ribosome entry site (IRES) segments yield satisfiable results with experimental support from the literature and further demonstrate the necessity of incorporating RNA tertiary structural information into the prediction model. The source code of our approach can be found in https://github.com/thucombio/deepnet-rbp.

  10. Sequence-Based Prediction of RNA-Binding Proteins Using Random Forest with Minimum Redundancy Maximum Relevance Feature Selection

    PubMed Central

    Ma, Xin; Guo, Jing; Sun, Xiao

    2015-01-01

    The prediction of RNA-binding proteins is one of the most challenging problems in computation biology. Although some studies have investigated this problem, the accuracy of prediction is still not sufficient. In this study, a highly accurate method was developed to predict RNA-binding proteins from amino acid sequences using random forests with the minimum redundancy maximum relevance (mRMR) method, followed by incremental feature selection (IFS). We incorporated features of conjoint triad features and three novel features: binding propensity (BP), nonbinding propensity (NBP), and evolutionary information combined with physicochemical properties (EIPP). The results showed that these novel features have important roles in improving the performance of the predictor. Using the mRMR-IFS method, our predictor achieved the best performance (86.62% accuracy and 0.737 Matthews correlation coefficient). High prediction accuracy and successful prediction performance suggested that our method can be a useful approach to identify RNA-binding proteins from sequence information. PMID:26543860

  11. The molecular features of uncoupling protein 1 support a conventional mitochondrial carrier-like mechanism.

    PubMed

    Crichton, Paul G; Lee, Yang; Kunji, Edmund R S

    2017-03-01

    Uncoupling protein 1 (UCP1) is an integral membrane protein found in the mitochondrial inner membrane of brown adipose tissue, and facilitates the process of non-shivering thermogenesis in mammals. Its activation by fatty acids, which overcomes its inhibition by purine nucleotides, leads to an increase in the proton conductance of the inner mitochondrial membrane, short-circuiting the mitochondrion to produce heat rather than ATP. Despite 40 years of intense research, the underlying molecular mechanism of UCP1 is still under debate. The protein belongs to the mitochondrial carrier family of transporters, which have recently been shown to utilise a domain-based alternating-access mechanism, cycling between a cytoplasmic and matrix state to transport metabolites across the inner membrane. Here, we review the protein properties of UCP1 and compare them to those of mitochondrial carriers. UCP1 has the same structural fold as other mitochondrial carriers and, in contrast to past claims, is a monomer, binding one purine nucleotide and three cardiolipin molecules tightly. The protein has a single substrate binding site, which is similar to those of the dicarboxylate and oxoglutarate carriers, but also contains a proton binding site and several hydrophobic residues. As found in other mitochondrial carriers, UCP1 has two conserved salt bridge networks on either side of the central cavity, which regulate access to the substrate binding site in an alternating way. The conserved domain structures and mobile inter-domain interfaces are consistent with an alternating access mechanism too. In conclusion, UCP1 has retained all of the key features of mitochondrial carriers, indicating that it operates by a conventional carrier-like mechanism.

  12. Homeodomain Protein Scr Regulates the Transcription of Genes Involved in Juvenile Hormone Biosynthesis in the Silkworm.

    PubMed

    Meng, Meng; Liu, Chun; Peng, Jian; Qian, Wenliang; Qian, Heying; Tian, Ling; Li, Jiarui; Dai, Dandan; Xu, Anying; Li, Sheng; Xia, Qingyou; Cheng, Daojun

    2015-11-02

    The silkworm Dominant trimolting (Moltinism, M³) mutant undergoes three larval molts and exhibits precocious metamorphosis. In this study, we found that compared with the wild-type (WT) that undergoes four larval molts, both the juvenile hormone (JH) concentration and the expression of the JH-responsive gene Krüppel homolog 1 (Kr-h1) began to be greater in the second instar of the M³ mutant. A positional cloning analysis revealed that only the homeodomain transcription factor gene Sex combs reduced (Scr) is located in the genomic region that is tightly linked to the M³ locus. The expression level of the Scr gene in the brain-corpora cardiaca-corpora allata (Br-CC-CA) complex, which controls the synthesis of JH, was very low in the final larval instar of both the M³ and WT larvae, and exhibited a positive correlation with JH titer changes. Importantly, luciferase reporter analysis and electrophoretic mobility shift assay (EMSA) demonstrated that the Scr protein could promote the transcription of genes involved in JH biosynthesis by directly binding to the cis-regulatory elements (CREs) of homeodomain protein on their promoters. These results conclude that the homeodomain protein Scr is transcriptionally involved in the regulation of JH biosynthesis in the silkworm.

  13. Interferon-inducible GTPase: a novel viral response protein involved in rabies virus infection.

    PubMed

    Li, Ling; Wang, Hualei; Jin, Hongli; Cao, Zengguo; Feng, Na; Zhao, Yongkun; Zheng, Xuexing; Wang, Jianzhong; Li, Qian; Zhao, Guoxing; Yan, Feihu; Wang, Lina; Wang, Tiecheng; Gao, Yuwei; Tu, Changchun; Yang, Songtao; Xia, Xianzhu

    2016-05-01

    Rabies virus infection is a major public health concern because of its wide host-interference spectrum and nearly 100 % lethality. However, the interactions between host and virus remain unclear. To decipher the authentic response in the central nervous system after rabies virus infection, a dynamic analysis of brain proteome alteration was performed. In this study, 104 significantly differentially expressed proteins were identified, and intermediate filament, interferon-inducible GTPases, and leucine-rich repeat-containing protein 16C were the three outstanding groups among these proteins. Interferon-inducible GTPases were prominent because of their strong upregulation. Moreover, quantitative real-time PCR showed distinct upregulation of interferon-inducible GTPases at the level of transcription. Several studies have shown that interferon-inducible GTPases are involved in many biological processes, such as viral infection, endoplasmic reticulum stress response, and autophagy. These findings indicate that interferon-inducible GTPases are likely to be a potential target involved in rabies pathogenesis or the antiviral process.

  14. Homeodomain Protein Scr Regulates the Transcription of Genes Involved in Juvenile Hormone Biosynthesis in the Silkworm

    PubMed Central

    Meng, Meng; Liu, Chun; Peng, Jian; Qian, Wenliang; Qian, Heying; Tian, Ling; Li, Jiarui; Dai, Dandan; Xu, Anying; Li, Sheng; Xia, Qingyou; Cheng, Daojun

    2015-01-01

    The silkworm Dominant trimolting (Moltinism, M3) mutant undergoes three larval molts and exhibits precocious metamorphosis. In this study, we found that compared with the wild-type (WT) that undergoes four larval molts, both the juvenile hormone (JH) concentration and the expression of the JH-responsive gene Krüppel homolog 1 (Kr-h1) began to be greater in the second instar of the M3 mutant. A positional cloning analysis revealed that only the homeodomain transcription factor gene Sex combs reduced (Scr) is located in the genomic region that is tightly linked to the M3 locus. The expression level of the Scr gene in the brain-corpora cardiaca-corpora allata (Br-CC-CA) complex, which controls the synthesis of JH, was very low in the final larval instar of both the M3 and WT larvae, and exhibited a positive correlation with JH titer changes. Importantly, luciferase reporter analysis and electrophoretic mobility shift assay (EMSA) demonstrated that the Scr protein could promote the transcription of genes involved in JH biosynthesis by directly binding to the cis-regulatory elements (CREs) of homeodomain protein on their promoters. These results conclude that the homeodomain protein Scr is transcriptionally involved in the regulation of JH biosynthesis in the silkworm. PMID:26540044

  15. Identification of ICIS-1, a new protein involved in cilia stability.

    PubMed

    Ponsard, Cecile; Skowron-Zwarg, Marie; Seltzer, Virginie; Perret, Eric; Gallinger, Julia; Fisch, Cathy; Dupuis-Williams, Pascale; Caruso, Nathalie; Middendorp, Sandrine; Tournier, Frederic

    2007-01-01

    Cilia are specialized organelles that exert critical functions in numerous organisms, including that of cell motility, fluid transport and protozoan locomotion. Ciliary architecture and function strictly depend on basal body formation, migration and axoneme elongation. Numerous ultrastructural studies have been undertaken in different species to elucidate the process of ciliogenesis. Recent analyses have led to identification of genes specifically expressed in ciliated organisms, but most proteins involved in ciliogenesis remain uncharacterized. Using human nasal epithelial cells capable of ciliary differentiation in vitro, differential display was carried out to identify new proteins associated with ciliogenesis. We isolated a new gene, ICIS-1 (Involved in CIlia Stability-1), upregulated during mucociliary differentiation. This gene is localized within the TGF-beta1 promoter and is ubiquitously expressed in human tissues. Functional analyses of gene expression inhibition by RNA interference in Paramecium tetraurelia indicated that the ICIS-1 homologue interfered with cilia stability or formation. These findings demonstrate that ICIS-1 is a new protein associated with ciliated cells and potentially related to cilia stability.

  16. Distinct Features of Cap Binding by eIF4E1b Proteins

    PubMed Central

    Kubacka, Dorota; Miguel, Ricardo Núñez; Minshall, Nicola; Darzynkiewicz, Edward; Standart, Nancy; Zuberek, Joanna

    2015-01-01

    eIF4E1b, closely related to the canonical translation initiation factor 4E (eIF4E1a), cap-binding protein is highly expressed in mouse, Xenopus and zebrafish oocytes. We have previously characterized eIF4E1b as a component of the CPEB mRNP translation repressor complex along with the eIF4E-binding protein 4E-Transporter, the Xp54/DDX6 RNA helicase and additional RNA-binding proteins. eIF4E1b exhibited only very weak interactions with m7GTP-Sepharose and, rather than binding eIF4G, interacted with 4E-T. Here we undertook a detailed examination of both Xenopus and human eIF4E1b interactions with cap analogues using fluorescence titration and homology modeling. The predicted structure of eIF4E1b maintains the α + β fold characteristic of eIF4E proteins and its cap-binding pocket is similarly arranged by critical amino acids: Trp56, Trp102, Glu103, Trp166, Arg112, Arg157 and Lys162 and residues of the C-terminal loop. However, we demonstrate that eIF4E1b is 3-fold less well able to bind the cap than eIF4E1a, both proteins being highly stimulated by methylation at N7 of guanine. Moreover, eIF4E1b proteins are distinguishable from eIF4E1a by a set of conserved amino acid substitutions, several of which are located near to cap-binding residues. Indeed, eIF4E1b possesses several distinct features, namely, enhancement of cap binding by a benzyl group at N7 position of guanine, a reduced response to increasing length of the phosphate chain and increased binding to a cap separated by a linker from Sepharose, suggesting differences in the arrangement of the protein's core. In agreement, mutagenesis of the amino acids differentiating eIF4E1b from eIF4E1a reduces cap binding by eIF4E1a 2-fold, demonstrating their role in modulating cap binding. PMID:25463438

  17. Involvement of lipid peroxidation-derived aldehyde-protein adducts in autoimmunity mediated by trichloroethene.

    PubMed

    Wang, Gangduo; Ansari, G A S; Khan, M Firoze

    2007-12-01

    Lipid peroxidation, a major contributor to cellular damage, is also implicated in the pathogenesis of autoimmune diseases (AD). The focus of this study was to elucidate the role of lipid peroxidation-derived aldehydes in autoimmunity induced and/or exacerbated by chemical exposure. Previous studies showed that trichloroethene (TCE) is capable of inducing/accelerating autoimmunity. To test whether TCE-induced lipid peroxidation might be involved in the induction/exacerbation of autoimmune responses, groups of autoimmune-prone female MRL +/+ mice were treated with TCE (10 mmol/kg, i.p., every 4th day) for 6 or 12 wk. Significant increases of the formation of malondialdehyde (MDA)- and 4-hydroxynonenal (HNE)-protein adducts were found in the livers of TCE-treated mice at both 6 and 12 wk, but the response was greater at 12 wk. Further characterization of these adducts in liver microsomes showed increased formation of MDA-protein adducts with molecular masses of 86, 65, 56, 44, and 32 kD, and of HNE-protein adducts with molecular masses of 87, 79, 46, and 17 kD in TCE-treated mice. In addition, significant induction of anti-MDA- and anti-HNE-protein adduct-specific antibodies was observed in the sera of TCE-treated mice, and showed a pattern similar to MDA- or HNE-protein adducts. The increases in anti-MDA- and anti-HNE-protein adduct antibodies were associated with significant elevation in serum anti-nuclear-, anti-ssDNA- and anti-dsDNA-antibodies at 6 wk and, to a greater extent, at 12 wk. These studies suggest that TCE-induced lipid peroxidation is associated with induction/exacerbation of autoimmune response in MRL+/+ mice, and thus may play an important role in disease pathogenesis. Further interventional studies are needed to establish a causal relationship between lipid peroxidation and TCE-induced autoimmune response.

  18. The Arabidopsis PLAT domain protein1 is critically involved in abiotic stress tolerance.

    PubMed

    Hyun, Tae Kyung; van der Graaff, Eric; Albacete, Alfonso; Eom, Seung Hee; Großkinsky, Dominik K; Böhm, Hannah; Janschek, Ursula; Rim, Yeonggil; Ali, Walid Wahid; Kim, Soo Young; Roitsch, Thomas

    2014-01-01

    Despite the completion of the Arabidopsis genome sequence, for only a relatively low percentage of the encoded proteins experimental evidence concerning their function is available. Plant proteins that harbour a single PLAT (Polycystin, Lipoxygenase, Alpha-toxin and Triacylglycerol lipase) domain and belong to the PLAT-plant-stress protein family are ubiquitously present in monocot and dicots. However, the function of PLAT-plant-stress proteins is still poorly understood. Therefore, we have assessed the function of the uncharacterised Arabidopsis PLAT-plant-stress family members through a combination of functional genetic and physiological approaches. PLAT1 overexpression conferred increased abiotic stress tolerance, including cold, drought and salt stress, while loss-of-function resulted in opposite effects on abiotic stress tolerance. Strikingly, PLAT1 promoted growth under non-stressed conditions. Abiotic stress treatments induced PLAT1 expression and caused expansion of its expression domain. The ABF/ABRE transcription factors, which are positive mediators of abscisic acid signalling, activate PLAT1 promoter activity in transactivation assays and directly bind to the ABRE elements located in this promoter in electrophoretic mobility shift assays. This suggests that PLAT1 represents a novel downstream target of the abscisic acid signalling pathway. Thus, we showed that PLAT1 critically functions as positive regulator of abiotic stress tolerance, but also is involved in regulating plant growth, and thereby assigned a function to this previously uncharacterised PLAT domain protein. The functional data obtained for PLAT1 support that PLAT-plant-stress proteins in general could be promising targets for improving abiotic stress tolerance without yield penalty.

  19. Evolution and diversity of periplasmic proteins involved in copper homeostasis in gamma proteobacteria

    PubMed Central

    2012-01-01

    Background Different systems contributing to copper homeostasis in bacteria have been described in recent years involving periplasmic and transport proteins that provide resistance via metal efflux to the extracellular media (CopA/Cue, Cus, Cut, and Pco). The participation of these proteins in the assembly of membrane, periplasmic and secreted cuproproteins has also been postulated. The integration and interrelation of these systems and their apparent redundancies are less clear since they have been studied in alternative systems. Based on the idea that cellular copper is not free but rather it is transferred via protein-protein interactions, we hypothesized that systems would coevolve and be constituted by set numbers of essential components. Results By the use of a phylogenomic approach we identified the distribution of 14 proteins previously characterized as members of homeostasis systems in the genomes of 268 gamma proteobacteria. Only 3% of the genomes presented the complete systems and 5% of them, all intracellular parasites, lacked the 14 genes. Surprisingly, copper homeostatic pathways did not behave as evolutionary units with particular species assembling different combinations of basic functions. The most frequent functions, and probably because of its distribution the most vital, were copper extrusion from the cytoplasm to the periplasm performed by CopA and copper export from the cytoplasm to the extracellular space performed by CusC, which along with the remaining 12 proteins, assemble in nine different functional repertoires. Conclusions These observations suggest complex evolutionary dynamics and still unexplored interactions to achieve copper homeostasis, challenging some of the molecular transport mechanism proposed for these systems. PMID:23122209

  20. Proteins involved in flor yeast carbon metabolism under biofilm formation conditions.

    PubMed

    Moreno-García, Jaime; García-Martínez, Teresa; Moreno, Juan; Mauricio, Juan Carlos

    2015-04-01

    A lack of sugars during the production of biologically aged wines after fermentation of grape must causes flor yeasts to metabolize other carbon molecules formed during fermentation (ethanol and glycerol, mainly). In this work, a proteome analysis involving OFFGEL fractionation prior to LC/MS detection was used to elucidate the carbon metabolism of a flor yeast strain under biofilm formation conditions (BFC). The results were compared with those obtained under non-biofilm formation conditions (NBFC). Proteins associated to processes such as non-fermentable carbon uptake, the glyoxylate and TCA cycles, cellular respiration and inositol metabolism were detected at higher concentrations under BFC than under the reference conditions (NBFC). This study constitutes the first attempt at identifying the flor yeast proteins responsible for the peculiar sensory profile of biologically aged wines. A better metabolic knowledge of flor yeasts might facilitate the development of effective strategies for improved production of these special wines.

  1. Involvement of Protein Kinase C-δ in Vascular Permeability in Acute Lung Injury.

    PubMed

    Ahn, Jong J; Jung, Jong P; Park, Soon E; Lee, Minhyun; Kwon, Byungsuk; Cho, Hong R

    2015-08-01

    Pulmonary edema is a major cause of mortality due to acute lung injury (ALI). The involvement of protein kinase C-δ (PKC-δ) in ALI has been a controversial topic. Here we investigated PKC-δ function in ALI using PKC-δ knockout (KO) mice and PKC inhibitors. Our results indicated that although the ability to produce proinflammatory mediators in response to LPS injury in PKC-δ KO mice was similar to that of control mice, they showed enhanced recruitment of neutrophils to the lung and more severe pulmonary edema. PKC-δ inhibition promoted barrier dysfunction in an endothelial cell layer in vitro, and administration of a PKC-δ-specific inhibitor significantly increased steady state vascular permeability. A neutrophil transmigration assay indicated that the PKC-δ inhibition increased neutrophil transmigration through an endothelial monolayer. This suggests that PKC-δ inhibition induces structural changes in endothelial cells, allowing extravasation of proteins and neutrophils.

  2. The H-NS protein is involved in the biogenesis of flagella in Escherichia coli.

    PubMed Central

    Bertin, P; Terao, E; Lee, E H; Lejeune, P; Colson, C; Danchin, A; Collatz, E

    1994-01-01

    The function of the flagellum-chemotaxis regulon requires the expression of many genes and is positively regulated by the cyclic AMP-catabolite activator protein (cAMP-CAP) complex. In this paper, we show that motile behavior was affected in Escherichia coli hns mutants. The loss of motility resulted from a complete lack of flagella. A decrease in the level of transcription of the flhD and fliA genes, which are both required for the synthesis of flagella, was observed in the presence of an hns mutation. Furthermore, the Fla- phenotype was not reversed to the wild type in the presence of a cfs mutation which renders the flagellum synthesis independent of the cAMP-CAP complex. These results suggest that the H-NS protein acts as a positive regulator of genes involved in the biogenesis of flagella by a mechanism independent of the cAMP-CAP pathway. Images PMID:8071234

  3. The Arabidopsis CLASP gene encodes a microtubule-associated protein involved in cell expansion and division.

    PubMed

    Ambrose, J Christian; Shoji, Tsubasa; Kotzer, Amanda M; Pighin, Jamie A; Wasteneys, Geoffrey O

    2007-09-01

    Controlling microtubule dynamics and spatial organization is a fundamental requirement of eukaryotic cell function. Members of the ORBIT/MAST/CLASP family of microtubule-associated proteins associate with the plus ends of microtubules, where they promote the addition of tubulin subunits into attached kinetochore fibers during mitosis and stabilize microtubules in the vicinity of the plasma membrane during interphase. To date, nothing is known about their function in plants. Here, we show that the Arabidopsis thaliana CLASP protein is a microtubule-associated protein that is involved in both cell division and cell expansion. Green fluorescent protein-CLASP localizes along the full length of microtubules and shows enrichment at growing plus ends. Our analysis suggests that CLASP promotes microtubule stability. clasp-1 T-DNA insertion mutants are hypersensitive to microtubule-destabilizing drugs and exhibit more sparsely populated, yet well ordered, root cortical microtubule arrays. Overexpression of CLASP promotes microtubule bundles that are resistant to depolymerization with oryzalin. Furthermore, clasp-1 mutants have aberrant microtubule preprophase bands, mitotic spindles, and phragmoplasts, indicating a role for At CLASP in stabilizing mitotic arrays. clasp-1 plants are dwarf, have significantly reduced cell numbers in the root division zone, and have defects in directional cell expansion. We discuss possible mechanisms of CLASP function in higher plants.

  4. Tyrosine phosphorylation and protein degradation control the transcriptional activity of WRKY involved in benzylisoquinoline alkaloid biosynthesis

    PubMed Central

    Yamada, Yasuyuki; Sato, Fumihiko

    2016-01-01

    Benzylisoquinoline alkaloids (BIQ) are among the most structurally diverse and pharmaceutically valuable secondary metabolites. A plant-specific WRKY-type transcription factor, CjWRKY1, was isolated from Coptis japonica and identified as a transcriptional activator of BIQ biosynthesis. However, the expression of CjWRKY1 gene alone was not sufficient for the activation of genes encoding biosynthetic enzymes. Here, we report the importance of post-translational regulation of CjWRKY1 in BIQ biosynthesis. First, we detected the differential accumulation of CjWRKY1 protein in two cell lines with similar CjWRKY1 gene expression but different levels of accumulated alkaloids. Further investigation of the WRKY protein identified the phosphorylation of the WRKYGQK core domain at Y115. The CjWRKYY115E phosphorylation-mimic mutant showed loss of nuclear localization, DNA-binding activity, and transactivation activity compared to wild-type CjWRKY1. Rapid degradation of the CjWRKY1 protein was also confirmed following treatment with inhibitors of the 26S proteasome and protease inhibitors. The existence of two independent degradation pathways as well as protein phosphorylation suggests the fine-tuning of CjWRKY1 activities is involved in the regulation of biosynthesis of BIQs. PMID:27552928

  5. Vps41, a protein involved in lysosomal trafficking, interacts with caspase-8.

    PubMed

    Wang, Lu; Pan, Xiao; He, Liangqiang; Zhang, Rong; Chen, Wei; Zhang, Jing; Lu, Min; Hua, Zi-Chun

    2013-01-01

    Caspase-8 is a member of the cysteine-aspartic acid protease (caspase) family which plays a central role in apoptosis and development. We screened caspase-8 interacting proteins from mouse T-cell lymphoma and 7.5-day embryo cDNA libraries by yeast two-hybrid system and obtained eleven positive clones, including Vacuolar protein sorting 41 (Vps41), a protein involved in trafficking of proteins from the late Golgi to the vacuole. The interaction of Vps41 with caspase-8 was confirmed by co-immunoprecipitation (co-IP) and co-localization studies in HEK293T cells. Co-IP experiments also showed that Vps41 binds to the p18 subunit of caspase-8 through its WD40 region and RING-finger motif. Furthermore, we found that overexpression of Vps41 promotes Fas-induced apoptosis in A549 human lung adenocarcinoma cells. The cleavage of caspase-3, a caspase-8 downstream effector, was increased when cells were transfected with Vps41-overexpressing plasmid. Together, these results suggest a novel interaction of caspase-8 with Vps41 and provide a potential role of Vps41 beyond lysosomal trafficking.

  6. Water-soluble chlorophyll protein is involved in herbivore resistance activation during greening of Arabidopsis thaliana

    PubMed Central

    Boex-Fontvieille, Edouard; Rustgi, Sachin; von Wettstein, Diter; Reinbothe, Steffen; Reinbothe, Christiane

    2015-01-01

    Water-soluble chlorophyll proteins (WSCPs) constitute a small family of unusual chlorophyll (Chl)-binding proteins that possess a Kunitz-type protease inhibitor domain. In Arabidopsis thaliana, a WSCP has been identified, named AtWSCP, that forms complexes with Chl and the Chl precursor chlorophyllide (Chlide) in vitro. AtWSCP exhibits a quite unexpected expression pattern for a Chl binding protein and accumulated to high levels in the apical hook of etiolated plants. AtWSCP expression was negatively light-regulated. Transgenic expression of AtWSCP fused to green fluorescent protein (GFP) revealed that AtWSCP is localized to cell walls/apoplastic spaces. Biochemical assays identified AtWSCP as interacting with RD21 (RESPONSIVE TO DESICCATION 21), a granulin domain-containing cysteine protease implicated in stress responses and defense. Reconstitution experiments showed tight interactions between RD21 and WSCP that were relieved upon Chlide binding. Laboratory feeding experiments with two herbivorous isopod crustaceans, Porcellio scaber (woodlouse) and Armadillidium vulgare (pillbug), identified the apical hook as Achilles’ heel of etiolated plants and that this was protected by RD21 during greening. Because Chlide is formed in the apical hook during seedling emergence from the soil, our data suggest an unprecedented mechanism of herbivore resistance activation that is triggered by light and involves AtWSCP. PMID:26016527

  7. Analysis of nitrated proteins in Saccharomyces cerevisiae involved in mating signal transduction.

    PubMed

    Kang, Jeong Won; Lee, Na Young; Cho, Kyung-Cho; Lee, Min Young; Choi, Do-Young; Park, Sang-Hyun; Kim, Kwang Pyo

    2015-01-01

    Protein tyrosine nitration (PTN) is a PTM that regulates signal transduction and inflammatory responses, and is related to neurodegenerative and cardiovascular diseases. The cellular function of PTN remains unclear because the low stoichiometry of PTN limits the identification and quantification of nitrated peptides. Effective enrichment is an important aspect of PTN analysis. In this study, we analyzed the in vivo nitroproteome elicited by mating signal transduction in Saccharomyces cerevisiae using a novel chemical enrichment method followed by LC-MS/MS. Nitroproteome profiling successfully identified changes in the nitration states of 14 proteins during mating signal transduction in S. cerevisiae, making this the first reported in vivo nitroproteome in yeast. We investigated the biological functions of these nitroproteins and their relationships to mating signal transduction in S. cerevisiae using a protein-protein interaction network. Our results suggest that PTN and denitration may be involved in nonreactive nitrogen species-mediated signal transduction and can provide clues for understanding the functional roles of PTN in vivo.

  8. Identification of proteins involved in desiccation tolerance in the red seaweed Pyropia orbicularis (Rhodophyta, Bangiales).

    PubMed

    López-Cristoffanini, Camilo; Zapata, Javier; Gaillard, Fanny; Potin, Philippe; Correa, Juan A; Contreras-Porcia, Loretto

    2015-12-01

    Extreme reduction in cellular water content leads to desiccation, which, if persistent, affects the physiology of organisms, mainly through oxidative stress. Some organisms are highly tolerant to desiccation, including resurrection plants and certain intertidal seaweeds. One such species is Pyropia orbicularis, a rhodophycean that colonizes upper intertidal zones along the Chilean coast. Despite long, daily periods of air exposure due to tides, this alga is highly tolerant to desiccation. The present study examined the proteome of P. orbicularis by 2DE and LC-MS/MS analyses to determine the proteins associated with desiccation tolerance (DT). The results showed that, under natural conditions, there were significant changes in the protein profile during low tide as compared to naturally hydrated plants at high tide. These changes were mainly in newly appeared proteins spots such as chaperones, monodehydroascorbate reductase, and manganese superoxide dismutase, among others. Previously undescribed proteins under desiccation conditions included phycobiliproteins, glyoxalase I, and phosphomannomutase. These changes evidenced that several physiological responses involved in DT are activated during low tide, including decreased photosynthetic activity, increased antioxidant capacity, and the preservation of cell physiology by regulating water content, cell wall structure, and cell volume. Similar responses have been observed in resurrection plants and bryophytes exposed to desiccation. Therefore, the coordinated activation of different desiccation tolerance pathways in P. orbicularis could explain the successful biological performance of this seaweed in the upper intertidal rocky zones.

  9. Neuronal and microglial involvement in beta-amyloid protein deposition in Alzheimer's disease.

    PubMed Central

    Cras, P.; Kawai, M.; Siedlak, S.; Mulvihill, P.; Gambetti, P.; Lowery, D.; Gonzalez-DeWhitt, P.; Greenberg, B.; Perry, G.

    1990-01-01

    This study was undertaken to localize amyloid precursor protein (APP) and to determine how APP might be released and proteolyzed to yield the beta-amyloid protein deposits found in senile plaques in the brains of Alzheimer's disease patients. We found that antibodies to recombinantly expressed APP labeled many normal neurons and neurites. In addition, dystrophic neurites in different types of senile plaques and degenerating neurons in the temporal cortex and hippocampus of Alzheimer's disease patients were immunostained. We also detected small clusters of dystrophic APP immunoreactive neurites that were not associated with beta-amyloid protein deposits. Microglia was involved in different types of senile plaques and often were associated closely with APP immunoreactive neurites and neurons. The greatest concurrence of APP immunoreactivity and reactive microglia was seen in the subiculum and area CA1, regions with a high density of congophilic plaques and subject to intense Alzheimer's pathology. Our findings suggest that neuronally derived APP is the source for senile plaque beta-amyloid protein, while microglia may act as processing cells. Images Figure 1 Figure 2 PMID:2117395

  10. Development of neurodevelopmental disorders: a regulatory mechanism involving bromodomain-containing proteins.

    PubMed

    Li, Junlin; Zhao, Guifang; Gao, Xiaocai

    2013-02-20

    Neurodevelopmental disorders are classified as diseases that cause abnormal functions of the brain or central nervous system. Children with neurodevelopmental disorders show impaired language and speech abilities, learning and memory damage, and poor motor skills. However, we still know very little about the molecular etiology of these disorders. Recent evidence implicates the bromodomain-containing proteins (BCPs) in the initiation and development of neurodevelopmental disorders. BCPs have a particular domain, the bromodomain (Brd), which was originally identified as specifically binding acetyl-lysine residues at the N-terminus of histone proteins in vitro and in vivo. Other domains of BCPs are responsible for binding partner proteins to form regulatory complexes. Once these complexes are assembled, BCPs alter chromosomal states and regulate gene expression. Some BCP complexes bind nucleosomes, are involved in basal transcription regulation, and influence the transcription of many genes. However, most BCPs are involved in targeting. For example, some BCPs function as a recruitment platform or scaffold through their Brds-binding targeting sites. Others are recruited to form a complex to bind the targeting sites of their partners. The regulation mediated by these proteins is especially critical during normal and abnormal development. Mutant BCPs or dysfunctional BCP-containing complexes are implicated in the initiation and development of neurodevelopmental disorders. However, the pathogenic molecular mechanisms are not fully understood. In this review, we focus on the roles of regulatory BCPs associated with neurodevelopmental disorders, including mental retardation, Fragile X syndrome (FRX), Williams syndrome (WS), Rett syndrome and Rubinstein-Taybi syndrome (RTS). A better understanding of the molecular pathogenesis, based upon the roles of BCPs, will lead to screening of targets for the treatment of neurodevelopmental disorders.

  11. The involvement of a protein kinase in phototaxis and gravitaxis of Euglena gracilis.

    PubMed

    Daiker, Viktor; Häder, Donat-P; Richter, Peter R; Lebert, Michael

    2011-05-01

    The unicellular flagellate Euglena gracilis shows positive phototaxis at low-light intensities (<10 W/m(2)) and a negative one at higher irradiances (>10 W/m(2)). Phototaxis is based on blue light-activated adenylyl cyclases, which produce cAMP upon irradiation. In the absence of light the cells swim upward in the water column (negative gravitaxis). The results of sounding rocket campaigns and of a large number of ground experiments led to the following model of signal perception and transduction in gravitaxis of E. gracilis: The body of the cell is heavier than the surrounding medium, sediments and thereby exerts a force onto the lower membrane. Upon deviation from a vertical swimming path mechano-sensitive ion channels are activated. Calcium is gated inwards which leads to an increase in the intracellular calcium concentration and causes a change of the membrane potential. After influx, calcium activates one of several calmodulins found in Euglena, which in turn activates an adenylyl cyclase (different from the one involved in phototaxis) to produce cAMP from ATP. One further element in the sensory transduction chain of both phototaxis and gravitaxis is a specific protein kinase A. We found five different protein kinases A in E. gracilis. The blockage of only one of these (PK.4, accession No. EU935859) by means of RNAi inhibited both phototaxis and gravitaxis, while inhibition of the other four affected neither phototaxis nor gravitaxis. It is assumed that cAMP directly activates this protein kinase A which may in turn phosphorylate a protein involved in the flagellar beating mechanism.

  12. Unique features of different members of the human guanylate-binding protein family.

    PubMed

    Tripal, Philipp; Bauer, Michael; Naschberger, Elisabeth; Mörtinger, Thomas; Hohenadl, Christine; Cornali, Emmanuelle; Thurau, Mathias; Stürzl, Michael

    2007-01-01

    Guanylate-binding proteins (GBPs) are the most abundant cellular proteins expressed in response to interferon-gamma (IFN-gamma), with seven highly homologous members in humans, termed HuGBP-1 to HuGBP-7. To date, differential features that may indicate differential functions of these proteins have not been described. Here, we investigated the expression and subcellular localization of the different HuGBPs in endothelial cells (EC). IFN-gamma, tumor necrosis factor-alpha (TNF-alpha), and interleukin-1beta (IL-1beta) induced the expression of HuGBP-1, HuGBP-2, and HuGBP-3 at similar high levels. In contrast, expression of HuGBP-4 and HuGBP-5 was robustly induced only by IFN-gamma and not by TNF-alpha and IL-1beta. Expression of HuGBP-6 and HuGBP-7 was not detected in EC under the various conditions examined. Investigating subcellular localization of the EC-expressed HuGBPs, HuGBP-1, HuGBP-3, and HuGBP-5 were exclusively detected in the cytoplasm, whereas HuGBP-2 and HuGBP-4 displayed a nucleocytoplasmic distribution. Treatment of the cells with IFN-gamma and aluminum fluoride caused rapid enrichment of HuGBP-1 and HuGBP-2 in the Golgi apparatus, as demonstrated by time-lapse microscopy and fluorescence analyses of GFP-tagged HuGBPs. HuGBP-3 and HuGBP-4 were never detected in the Golgi apparatus, whereas HuGBP-5 was constitutively enriched in this cytosolic compartment, irrespective of stimulation. These results assign a characteristic pattern of expression and subcellular localization to each of the HuGBPs, indicating for the first time that these proteins may have different cellular functions.

  13. Accurate Prediction of One-Dimensional Protein Structure Features Using SPINE-X.

    PubMed

    Faraggi, Eshel; Kloczkowski, Andrzej

    2017-01-01

    Accurate prediction of protein secondary structure and other one-dimensional structure features is essential for accurate sequence alignment, three-dimensional structure modeling, and function prediction. SPINE-X is a software package to predict secondary structure as well as accessible surface area and dihedral angles ϕ and ψ. For secondary structure SPINE-X achieves an accuracy of between 81 and 84 % depending on the dataset and choice of tests. The Pearson correlation coefficient for accessible surface area prediction is 0.75 and the mean absolute error from the ϕ and ψ dihedral angles are 20(∘) and 33(∘), respectively. The source code and a Linux executables for SPINE-X are available from Research and Information Systems at http://mamiris.com .

  14. Analytical techniques and computer algorithms combined for the rapid characterization of structural peptide and protein features.

    PubMed

    Caporale, C

    1998-11-01

    The most recent algorithms based on the use of modern analytical techniques for the assessment of structural peptide and protein features have been reviewed. No algorithm devoted to the realization of predictive models or statistical analysis has been discussed, but only methods furnishing information on the real structure of the molecules. In particular, the procedures designed for handling sequence and mass spectrometric data obtained from the analysis of unfractionated digestion mixtures allow the user to get rapid information on the structure of the target polypeptide. Two classes of methods are illustrated: the first regards the determination of the amino acid sequence, whereas the second used its knowledge to supply data on the localization, function and three-dimensional structure of disulphides.

  15. STAT5 proteins are involved in down-regulation of iron regulatory protein 1 gene expression by nitric oxide.

    PubMed

    Starzynski, Rafal Radoslaw; Gonçalves, Ana Sofia; Muzeau, Françoise; Tyrolczyk, Zofia; Smuda, Ewa; Drapier, Jean-Claude; Beaumont, Carole; Lipinski, Pawel

    2006-12-01

    RNA-binding activity of IRP1 (iron regulatory protein 1) is regulated by the insertion/extrusion of a [4Fe-4S] cluster into/from the IRP1 molecule. NO (nitic oxide), whose ability to activate IRP1 by removing its [4Fe-4S] cluster is well known, has also been shown to down-regulate expression of the IRP1 gene. In the present study, we examine whether this regulation occurs at the transcriptional level. Analysis of the mouse IRP1 promoter sequence revealed two conserved putative binding sites for transcription factor(s) regulated by NO and/or changes in intracellular iron level: Sp1 (promoter-selective transcription factor 1) and MTF1 (metal transcription factor 1), plus GAS (interferon-gamma-activated sequence), a binding site for STAT (signal transducer and activator of transcription) proteins. In order to define the functional activity of these sequences, reporter constructs were generated through the insertion of overlapping fragments of the mouse IRP1 promoter upstream of the luciferase gene. Transient expression assays following transfection of HuH7 cells with these plasmids revealed that while both the Sp1 and GAS sequences are involved in basal transcriptional activity of the IRP1 promoter, the role of the latter is predominant. Analysis of protein binding to these sequences in EMSAs (electrophoretic mobility-shift assays) using nuclear extracts from mouse RAW 264.7 macrophages stimulated to synthesize NO showed a significant decrease in the formation of Sp1-DNA and STAT-DNA complexes, compared with controls. We have also demonstrated that the GAS sequence is involved in NO-dependent down-regulation of IRP1 transcription. Further analysis revealed that levels of STAT5a and STAT5b in the nucleus and cytosol of NO-producing macrophages are substantially lower than in control cells. These findings provide evidence that STAT5 proteins play a role in NO-mediated down-regulation of IRP1 gene expression.

  16. Characterization of a Novel Endoplasmic Reticulum Protein Involved in Tubercidin Resistance in Leishmania major

    PubMed Central

    Aoki, Juliana Ide; Coelho, Adriano Cappellazzo; Muxel, Sandra Marcia; Zampieri, Ricardo Andrade; Sanchez, Eduardo Milton Ramos; Nerland, Audun Helge; Floeter-Winter, Lucile Maria; Cotrim, Paulo Cesar

    2016-01-01

    Background Tubercidin (TUB) is a toxic adenosine analog with potential antiparasitic activity against Leishmania, with mechanism of action and resistance that are not completely understood. For understanding the mechanisms of action and identifying the potential metabolic pathways affected by this drug, we employed in this study an overexpression/selection approach using TUB for the identification of potential targets, as well as, drug resistance genes in L. major. Although, TUB is toxic to the mammalian host, these findings can provide evidences for a rational drug design based on purine pathway against leishmaniasis. Methodology/Principal findings After transfection of a cosmid genomic library into L. major Friedlin (LmjF) parasites and application of the overexpression/selection method, we identified two cosmids (cosTUB1 and cosTU2) containing two different loci capable of conferring significant levels of TUB resistance. In the cosTUB1 contained a gene encoding NUPM1-like protein, which has been previously described as associated with TUB resistance in L. amazonensis. In the cosTUB2 we identified and characterized a gene encoding a 63 kDa protein that we denoted as tubercidin-resistance protein (TRP). Functional analysis revealed that the transfectants were less susceptible to TUB than LmjF parasites or those transfected with the control vector. In addition, the trp mRNA and protein levels in cosTUB2 transfectants were higher than LmjF. TRP immunolocalization revealed that it was co-localized to the endoplasmic reticulum (ER), a cellular compartment with many functions. In silico predictions indicated that TRP contains only a hypothetical transmembrane domain. Thus, it is likely that TRP is a lumen protein involved in multidrug efflux transport that may be involved in the purine metabolic pathway. Conclusions/Significance This study demonstrated for the first time that TRP is associated with TUB resistance in Leishmania. The next challenge is to determine how

  17. Automatic segmentation and 3D feature extraction of protein aggregates in Caenorhabditis elegans

    NASA Astrophysics Data System (ADS)

    Rodrigues, Pedro L.; Moreira, António H. J.; Teixeira-Castro, Andreia; Oliveira, João; Dias, Nuno; Rodrigues, Nuno F.; Vilaça, João L.

    2012-03-01

    In the last years, it has become increasingly clear that neurodegenerative diseases involve protein aggregation, a process often used as disease progression readout and to develop therapeutic strategies. This work presents an image processing tool to automatic segment, classify and quantify these aggregates and the whole 3D body of the nematode Caenorhabditis Elegans. A total of 150 data set images, containing different slices, were captured with a confocal microscope from animals of distinct genetic conditions. Because of the animals' transparency, most of the slices pixels appeared dark, hampering their body volume direct reconstruction. Therefore, for each data set, all slices were stacked in one single 2D image in order to determine a volume approximation. The gradient of this image was input to an anisotropic diffusion algorithm that uses the Tukey's biweight as edge-stopping function. The image histogram median of this outcome was used to dynamically determine a thresholding level, which allows the determination of a smoothed exterior contour of the worm and the medial axis of the worm body from thinning its skeleton. Based on this exterior contour diameter and the medial animal axis, random 3D points were then calculated to produce a volume mesh approximation. The protein aggregations were subsequently segmented based on an iso-value and blended with the resulting volume mesh. The results obtained were consistent with qualitative observations in literature, allowing non-biased, reliable and high throughput protein aggregates quantification. This may lead to a significant improvement on neurodegenerative diseases treatment planning and interventions prevention.

  18. Spa2p Interacts with Cell Polarity Proteins and Signaling Components Involved in Yeast Cell Morphogenesis

    PubMed Central

    Sheu, Yi-Jun; Santos, Beatriz; Fortin, Nathalie; Costigan, Christine; Snyder, Michael

    1998-01-01

    The yeast protein Spa2p localizes to growth sites and is important for polarized morphogenesis during budding, mating, and pseudohyphal growth. To better understand the role of Spa2p in polarized growth, we analyzed regions of the protein important for its function and proteins that interact with Spa2p. Spa2p interacts with Pea2p and Bud6p (Aip3p) as determined by the two-hybrid system; all of these proteins exhibit similar localization patterns, and spa2Δ, pea2Δ, and bud6Δ mutants display similar phenotypes, suggesting that these three proteins are involved in the same biological processes. Coimmunoprecipitation experiments demonstrate that Spa2p and Pea2p are tightly associated with each other in vivo. Velocity sedimentation experiments suggest that a significant portion of Spa2p, Pea2p, and Bud6p cosediment, raising the possibility that these proteins form a large, 12S multiprotein complex. Bud6p has been shown previously to interact with actin, suggesting that the 12S complex functions to regulate the actin cytoskeleton. Deletion analysis revealed that multiple regions of Spa2p are involved in its localization to growth sites. One of the regions involved in Spa2p stability and localization interacts with Pea2p; this region contains a conserved domain, SHD-II. Although a portion of Spa2p is sufficient for localization of itself and Pea2p to growth sites, only the full-length protein is capable of complementing spa2 mutant defects, suggesting that other regions are required for Spa2p function. By using the two-hybrid system, Spa2p and Bud6p were also found to interact with components of two mitogen-activated protein kinase (MAPK) pathways important for polarized cell growth. Spa2p interacts with Ste11p (MAPK kinase [MEK] kinase) and Ste7p (MEK) of the mating signaling pathway as well as with the MEKs Mkk1p and Mkk2p of the Slt2p (Mpk1p) MAPK pathway; for both Mkk1p and Ste7p, the Spa2p-interacting region was mapped to the N-terminal putative regulatory domain

  19. Many Local Pattern Texture Features: Which Is Better for Image-Based Multilabel Human Protein Subcellular Localization Classification?

    PubMed Central

    Xu, Ying-Ying; Shen, Hong-Bin

    2014-01-01

    Human protein subcellular location prediction can provide critical knowledge for understanding a protein's function. Since significant progress has been made on digital microscopy, automated image-based protein subcellular location classification is urgently needed. In this paper, we aim to investigate more representative image features that can be effectively used for dealing with the multilabel subcellular image samples. We prepared a large multilabel immunohistochemistry (IHC) image benchmark from the Human Protein Atlas database and tested the performance of different local texture features, including completed local binary pattern, local tetra pattern, and the standard local binary pattern feature. According to our experimental results from binary relevance multilabel machine learning models, the completed local binary pattern, and local tetra pattern are more discriminative for describing IHC images when compared to the traditional local binary pattern descriptor. The combination of these two novel local pattern features and the conventional global texture features is also studied. The enhanced performance of final binary relevance classification model trained on the combined feature space demonstrates that different features are complementary to each other and thus capable of improving the accuracy of classification. PMID:25050396

  20. Shewanella oneidensis MR-1 sensory box protein involved in aerobic and anoxic growth.

    PubMed

    Sundararajan, A; Kurowski, J; Yan, T; Klingeman, D M; Joachimiak, M P; Zhou, J; Naranjo, B; Gralnick, J A; Fields, M W

    2011-07-01

    Although little is known of potential function for conserved signaling proteins, it is hypothesized that such proteins play important roles to coordinate cellular responses to environmental stimuli. In order to elucidate the function of a putative sensory box protein (PAS domains) in Shewanella oneidensis MR-1, the physiological role of SO3389 was characterized. The predicted open reading frame (ORF) encodes a putative sensory box protein that has PAS, GGDEF, and EAL domains, and an in-frame deletion mutant was constructed (ΔSO3389) with approximately 95% of the ORF deleted. Under aerated conditions, wild-type and mutant cultures had similar growth rates, but the mutant culture had a lower growth rate under static, aerobic conditions. Oxygen consumption rates were lower for mutant cultures (1.5-fold), and wild-type cultures also maintained lower dissolved oxygen concentrations under aerated growth conditions. When transferred to anoxic conditions, the mutant did not grow with fumarate, iron(III), or dimethyl sulfoxide (DMSO) as electron acceptors. Biochemical assays demonstrated the expression of different c-type cytochromes as well as decreased fumarate reductase activity in the mutant transferred to anoxic growth conditions. Transcriptomic studies showed the inability of the mutant to up-express and down-express genes, including c-type cytochromes (e.g., SO4047/SO4048, SO3285/SO3286), reductases (e.g., SO0768, SO1427), and potential regulators (e.g., SO1329). The complemented strain was able to grow when transferred from aerobic to anoxic growth conditions with the tested electron acceptors. The modeled structure for the SO3389 PAS domains was highly similar to the crystal structures of FAD-binding PAS domains that are known O2/redox sensors. Based on physiological, genomic, and bioinformatic results, we suggest that the sensory box protein, SO3389, is an O2/redox sensor that is involved in optimization of aerobic growth and transitions to anoxia in S

  1. Mechanosensitive Molecular Networks Involved in Transducing Resistance Exercise-Signals into Muscle Protein Accretion

    PubMed Central

    Rindom, Emil; Vissing, Kristian

    2016-01-01

    Loss of skeletal muscle myofibrillar protein with disease and/or inactivity can severely deteriorate muscle strength and function. Strategies to counteract wasting of muscle myofibrillar protein are therefore desirable and invite for considerations on the potential superiority of specific modes of resistance exercise and/or the adequacy of low load resistance exercise regimens as well as underlying mechanisms. In this regard, delineation of the potentially mechanosensitive molecular mechanisms underlying muscle protein synthesis (MPS), may contribute to an understanding on how differentiated resistance exercise can transduce a mechanical signal into stimulation of muscle accretion. Recent findings suggest specific upstream exercise-induced mechano-sensitive myocellular signaling pathways to converge on mammalian target of rapamycin complex 1 (mTORC1), to influence MPS. This may e.g. implicate mechanical activation of signaling through a diacylglycerol kinase (DGKζ)-phosphatidic acid (PA) axis or implicate integrin deformation to signal through a Focal adhesion kinase (FAK)-Tuberous Sclerosis Complex 2 (TSC2)-Ras homolog enriched in brain (Rheb) axis. Moreover, since initiation of translation is reliant on mRNA, it is also relevant to consider potentially mechanosensitive signaling pathways involved in muscle myofibrillar gene transcription and whether some of these pathways converge with those affecting mTORC1 activation for MPS. In this regard, recent findings suggest how mechanical stress may implicate integrin deformation and/or actin dynamics to signal through a Ras homolog gene family member A protein (RhoA)-striated muscle activator of Rho signaling (STARS) axis or implicate deformation of Notch to affect Bone Morphogenetic Protein (BMP) signaling through a small mother of decapentaplegic (Smad) axis. PMID:27909410

  2. Mechanosensitive Molecular Networks Involved in Transducing Resistance Exercise-Signals into Muscle Protein Accretion.

    PubMed

    Rindom, Emil; Vissing, Kristian

    2016-01-01

    Loss of skeletal muscle myofibrillar protein with disease and/or inactivity can severely deteriorate muscle strength and function. Strategies to counteract wasting of muscle myofibrillar protein are therefore desirable and invite for considerations on the potential superiority of specific modes of resistance exercise and/or the adequacy of low load resistance exercise regimens as well as underlying mechanisms. In this regard, delineation of the potentially mechanosensitive molecular mechanisms underlying muscle protein synthesis (MPS), may contribute to an understanding on how differentiated resistance exercise can transduce a mechanical signal into stimulation of muscle accretion. Recent findings suggest specific upstream exercise-induced mechano-sensitive myocellular signaling pathways to converge on mammalian target of rapamycin complex 1 (mTORC1), to influence MPS. This may e.g. implicate mechanical activation of signaling through a diacylglycerol kinase (DGKζ)-phosphatidic acid (PA) axis or implicate integrin deformation to signal through a Focal adhesion kinase (FAK)-Tuberous Sclerosis Complex 2 (TSC2)-Ras homolog enriched in brain (Rheb) axis. Moreover, since initiation of translation is reliant on mRNA, it is also relevant to consider potentially mechanosensitive signaling pathways involved in muscle myofibrillar gene transcription and whether some of these pathways converge with those affecting mTORC1 activation for MPS. In this regard, recent findings suggest how mechanical stress may implicate integrin deformation and/or actin dynamics to signal through a Ras homolog gene family member A protein (RhoA)-striated muscle activator of Rho signaling (STARS) axis or implicate deformation of Notch to affect Bone Morphogenetic Protein (BMP) signaling through a small mother of decapentaplegic (Smad) axis.

  3. Quantitative LC-MS/MS Analysis of Proteins Involved in Metastasis of Breast Cancer

    PubMed Central

    Goto, Rieko; Nakamura, Yasushi; Takami, Tomonori; Sanke, Tokio; Tozuka, Zenzaburo

    2015-01-01

    The purpose of this study was to develop quantitative liquid chromatography-tandem mass spectrometry (LC-MS/MS) methods for the analysis of proteins involved in metastasis of breast cancer for diagnosis and determining disease prognosis, as well as to further our understand of metastatic mechanisms. We have previously demonstrated that the protein type XIV collagen may be specifically expressed in metastatic tissues by two dimensional LC-MS/MS. In this study, we developed quantitative LC-MS/MS methods for type XIV collagen. Type XIV collagen was quantified by analyzing 2 peptides generated by digesting type XIV collagen using stable isotope-labeled peptides. The individual concentrations were equivalent between 2 different peptides of type XIV collagen by evaluation of imprecise transitions and using the best transition for the peptide concentration. The results indicated that type XIV collagen is highly expressed in metastatic tissues of patients with massive lymph node involvement compared to non-metastatic tissues. These findings were validated by quantitative real-time RT-PCR. Further studies on type XIV collagen are desired to verify its role as a prognostic factor and diagnosis marker for metastasis. PMID:26176947

  4. Identification of a novel thylakoid protein gene involved in cold acclimation in cyanobacteria.

    PubMed

    Li, Weizhi; Gao, Hong; Yin, Chuntao; Xu, Xudong

    2012-09-01

    In cyanobacteria, genes involved in cold acclimation can be upregulated in response to cold stress with or without light. By inactivating 17 such genes in Synechocystis sp. PCC 6803, slr0815 (ccr2) was identified to be a novel gene required for survival at 15 °C. It was upregulated by cold stress in the light. Upon exposure to low temperature, a ccr2-null mutant showed greatly reduced photosynthetic and respiratory activities within 12 h relative to the wild-type. At 48 h, the photosystem (PS)II-mediated electron transport in the mutant was reduced to less than one-third of the wild-type level, and the duration of electron transfer from the Q(B) binding site of PSII to PSI was increased to about eight times the wild-type level, whereas the PSI-mediated electron transport remained unchanged. Using an antibody against GFP, a Ccr2-GFP fusion protein was localized to the thylakoid membrane rather than the cytoplasmic and outer membranes. Homologues to Ccr2 can be found in most cyanobacteria, algae and higher plants with sequenced genomes. Ccr2 is probably representative of a group of novel thylakoid proteins involved in acclimation to cold or other stresses.

  5. Text as data: using text-based features for proteins representation and for computational prediction of their characteristics.

    PubMed

    Shatkay, Hagit; Brady, Scott; Wong, Andrew

    2015-03-01

    The current era of large-scale biology is characterized by a fast-paced growth in the number of sequenced genomes and, consequently, by a multitude of identified proteins whose function has yet to be determined. Simultaneously, any known or postulated information concerning genes and proteins is part of the ever-growing published scientific literature, which is expanding at a rate of over a million new publications per year. Computational tools that attempt to automatically predict and annotate protein characteristics, such as function and localization patterns, are being developed along with systems that aim to support the process via text mining. Most work on protein characterization focuses on features derived directly from protein sequence data. Protein-related work that does aim to utilize the literature typically concentrates on extracting specific facts (e.g., protein interactions) from text. In the past few years we have taken a different route, treating the literature as a source of text-based features, which can be employed just as sequence-based protein-features were used in earlier work, for predicting protein subcellular location and possibly also function. We discuss here in detail the overall approach, along with results from work we have done in this area demonstrating the value of this method and its potential use.

  6. Weak conservation of structural features in the interfaces of homologous transient protein–protein complexes

    PubMed Central

    Sudha, Govindarajan; Singh, Prashant; Swapna, Lakshmipuram S; Srinivasan, Narayanaswamy

    2015-01-01

    Residue types at the interface of protein–protein complexes (PPCs) are known to be reasonably well conserved. However, we show, using a dataset of known 3-D structures of homologous transient PPCs, that the 3-D location of interfacial residues and their interaction patterns are only moderately and poorly conserved, respectively. Another surprising observation is that a residue at the interface that is conserved is not necessarily in the interface in the homolog. Such differences in homologous complexes are manifested by substitution of the residues that are spatially proximal to the conserved residue and structural differences at the interfaces as well as differences in spatial orientations of the interacting proteins. Conservation of interface location and the interaction pattern at the core of the interfaces is higher than at the periphery of the interface patch. Extents of variability of various structural features reported here for homologous transient PPCs are higher than the variation in homologous permanent homomers. Our findings suggest that straightforward extrapolation of interfacial nature and inter-residue interaction patterns from template to target could lead to serious errors in the modeled complex structure. Understanding the evolution of interfaces provides insights to improve comparative modeling of PPC structures. PMID:26311309

  7. A novel pax-like protein involved in transcriptional activation of cyst wall protein genes in Giardia lamblia.

    PubMed

    Wang, Yi-Ting; Pan, Yu-Jiao; Cho, Chao-Cheng; Lin, Bo-Chi; Su, Li-Hsin; Huang, Yu-Chang; Sun, Chin-Hung

    2010-10-15

    Giardia lamblia differentiates into infectious cysts to survive outside of the host. It is of interest to identify factors involved in up-regulation of cyst wall proteins (CWPs) during this differentiation. Pax proteins are important regulators of development and cell differentiation in Drosophila and vertebrates. No member of this gene family has been reported to date in yeast, plants, or protozoan parasites. We have identified a pax-like gene (pax1) encoding a putative paired domain in the G. lamblia genome. Epitope-tagged Pax1 localized to nuclei during both vegetative growth and encystation. Recombinant Pax1 specifically bound to the AT-rich initiator elements of the encystation-induced cwp1 to -3 and myb2 genes. Interestingly, overexpression of Pax1 increased cwp1 to -3 and myb2 gene expression and cyst formation. Deletion of the C-terminal paired domain or mutation of the basic amino acids of the paired domain resulted in a decrease of the transactivation function of Pax1. Our results indicate that the Pax family has been conserved during evolution, and Pax1 could up-regulate the key encystation-induced genes to regulate differentiation of the protozoan eukaryote, G. lamblia.

  8. The cyanobacterial Fluorescence Recovery Protein has two distinct activities: Orange Carotenoid Protein amino acids involved in FRP interaction.

    PubMed

    Thurotte, Adrien; Bourcier de Carbon, Céline; Wilson, Adjélé; Talbot, Léa; Cot, Sandrine; López-Igual, Rocio; Kirilovsky, Diana

    2017-04-01

    To deal with fluctuating light condition, cyanobacteria have developed a photoprotective mechanism which, under high light conditions, decreases the energy arriving at the photochemical centers. It relies on a photoswitch, the Orange Carotenoid Protein (OCP). Once photoactivated, OCP binds to the light harvesting antenna, the phycobilisome (PBS), and triggers the thermal dissipation of the excess energy absorbed. Deactivation of the photoprotective mechanism requires the intervention of a third partner, the Fluorescence Recovery Protein (FRP). FRP by interacting with the photoactivated OCP accelerates its conversion to the non-active form and its detachment from the phycobilisome. We have studied the interaction of FRP with free and phycobilisome-bound OCP. Several OCP variants were constructed and characterized. In this article we show that OCP amino acid F299 is essential and D220 important for OCP deactivation mediated by FRP. Mutations of these amino acids did not affect FRP activity as helper to detach OCP from phycobilisomes. In addition, while mutated R60L FRP is inactive on OCP deactivation, its activity on the detachment of the OCP from the phycobilisomes is not affected. Thus, our results demonstrate that FRP has two distinct activities: it accelerates OCP detachment from phycobilisomes and then it helps deactivation of the OCP. They also suggest that different OCP and FRP amino acids could be involved in these two activities.

  9. Identification and Characterization of Anaplasma phagocytophilum Proteins Involved in Infection of the Tick Vector, Ixodes scapularis.

    PubMed

    Villar, Margarita; Ayllón, Nieves; Kocan, Katherine M; Bonzón-Kulichenko, Elena; Alberdi, Pilar; Blouin, Edmour F; Weisheit, Sabine; Mateos-Hernández, Lourdes; Cabezas-Cruz, Alejandro; Bell-Sakyi, Lesley; Vancová, Marie; Bílý, Tomáš; Meyer, Damien F; Sterba, Jan; Contreras, Marinela; Rudenko, Nataliia; Grubhoffer, Libor; Vázquez, Jesús; de la Fuente, José

    2015-01-01

    Anaplasma phagocytophilum is an emerging zoonotic pathogen transmitted by Ixodes scapularis that causes human granulocytic anaplasmosis. Here, a high throughput quantitative proteomics approach was used to characterize A. phagocytophilum proteome during rickettsial multiplication and identify proteins involved in infection of the tick vector, I. scapularis. The first step in this research was focused on tick cells infected with A. phagocytophilum and sampled at two time points containing 10-15% and 65-71% infected cells, respectively to identify key bacterial proteins over-represented in high percentage infected cells. The second step was focused on adult female tick guts and salivary glands infected with A. phagocytophilum to compare in vitro results with those occurring during bacterial infection in vivo. The results showed differences in the proteome of A. phagocytophilum in infected ticks with higher impact on protein synthesis and processing than on bacterial replication in tick salivary glands. These results correlated well with the developmental cycle of A. phagocytophilum, in which cells convert from an intracellular reticulated, replicative form to the nondividing infectious dense-core form. The analysis of A. phagocytophilum differentially represented proteins identified stress response (GroEL, HSP70) and surface (MSP4) proteins that were over-represented in high percentage infected tick cells and salivary glands when compared to low percentage infected cells and guts, respectively. The results demonstrated that MSP4, GroEL and HSP70 interact and bind to tick cells, thus playing a role in rickettsia-tick interactions. The most important finding of these studies is the increase in the level of certain bacterial stress response and surface proteins in A. phagocytophilum-infected tick cells and salivary glands with functional implication in tick-pathogen interactions. These results gave a new dimension to the role of these stress response and surface

  10. Identification and Characterization of Anaplasma phagocytophilum Proteins Involved in Infection of the Tick Vector, Ixodes scapularis

    PubMed Central

    Kocan, Katherine M.; Bonzón-Kulichenko, Elena; Alberdi, Pilar; Blouin, Edmour F.; Weisheit, Sabine; Mateos-Hernández, Lourdes; Cabezas-Cruz, Alejandro; Bell-Sakyi, Lesley; Vancová, Marie; Bílý, Tomáš; Meyer, Damien F.; Sterba, Jan; Contreras, Marinela; Rudenko, Nataliia; Grubhoffer, Libor; Vázquez, Jesús; de la Fuente, José

    2015-01-01

    Anaplasma phagocytophilum is an emerging zoonotic pathogen transmitted by Ixodes scapularis that causes human granulocytic anaplasmosis. Here, a high throughput quantitative proteomics approach was used to characterize A. phagocytophilum proteome during rickettsial multiplication and identify proteins involved in infection of the tick vector, I. scapularis. The first step in this research was focused on tick cells infected with A. phagocytophilum and sampled at two time points containing 10–15% and 65–71% infected cells, respectively to identify key bacterial proteins over-represented in high percentage infected cells. The second step was focused on adult female tick guts and salivary glands infected with A. phagocytophilum to compare in vitro results with those occurring during bacterial infection in vivo. The results showed differences in the proteome of A. phagocytophilum in infected ticks with higher impact on protein synthesis and processing than on bacterial replication in tick salivary glands. These results correlated well with the developmental cycle of A. phagocytophilum, in which cells convert from an intracellular reticulated, replicative form to the nondividing infectious dense-core form. The analysis of A. phagocytophilum differentially represented proteins identified stress response (GroEL, HSP70) and surface (MSP4) proteins that were over-represented in high percentage infected tick cells and salivary glands when compared to low percentage infected cells and guts, respectively. The results demonstrated that MSP4, GroEL and HSP70 interact and bind to tick cells, thus playing a role in rickettsia-tick interactions. The most important finding of these studies is the increase in the level of certain bacterial stress response and surface proteins in A. phagocytophilum-infected tick cells and salivary glands with functional implication in tick-pathogen interactions. These results gave a new dimension to the role of these stress response and surface

  11. Structural and Functional Features of a Developmentally Regulated Lipopolysaccharide-Binding Protein

    PubMed Central

    Krasity, Benjamin C.; Troll, Joshua V.; Lehnert, Erik M.; Hackett, Kathleen T.; Dillard, Joseph P.; Apicella, Michael A.; Goldman, William E.

    2015-01-01

    ABSTRACT Mammalian lipopolysaccharide (LPS) binding proteins (LBPs) occur mainly in extracellular fluids and promote LPS delivery to specific host cell receptors. The function of LBPs has been studied principally in the context of host defense; the possible role of LBPs in nonpathogenic host-microbe interactions has not been well characterized. Using the Euprymna scolopes-Vibrio fischeri model, we analyzed the structure and function of an LBP family protein, E. scolopes LBP1 (EsLBP1), and provide evidence for its role in triggering a symbiont-induced host developmental program. Previous studies showed that, during initial host colonization, the LPS of V. fischeri synergizes with peptidoglycan (PGN) monomer to induce morphogenesis of epithelial tissues of the host animal. Computationally modeled EsLBP1 shares some but not all structural features of mammalian LBPs that are thought important for LPS binding. Similar to human LBP, recombinant EsLBP1 expressed in insect cells bound V. fischeri LPS and Neisseria meningitidis lipooligosaccharide (LOS) with nanomolar or greater affinity but bound Francisella tularensis LPS only weakly and did not bind PGN monomer. Unlike human LBP, EsLBP1 did not bind N. meningitidis LOS:CD14 complexes. The eslbp1 transcript was upregulated ~22-fold by V. fischeri at 24 h postinoculation. Surprisingly, this upregulation was not induced by exposure to LPS but, rather, to the PGN monomer alone. Hybridization chain reaction-fluorescent in situ hybridization (HCR-FISH) and immunocytochemistry (ICC) localized eslbp1 transcript and protein in crypt epithelia, where V. fischeri induces morphogenesis. The data presented here provide a window into the evolution of LBPs and the scope of their roles in animal symbioses. PMID:26463160

  12. Unconventional N-H…N Hydrogen Bonds Involving Proline Backbone Nitrogen in Protein Structures.

    PubMed

    Deepak, R N V Krishna; Sankararamakrishnan, Ramasubbu

    2016-05-10

    Contrary to DNA double-helical structures, hydrogen bonds (H-bonds) involving nitrogen as the acceptor are not common in protein structures. We systematically searched N-H…N H-bonds in two different sets of protein structures. Data set I consists of neutron diffraction and ultrahigh-resolution x-ray structures (0.9 Å resolution or better) and the hydrogen atom positions in these structures were determined experimentally. Data set II contains structures determined using x-ray diffraction (resolution ≤ 1.8 Å) and the positions of hydrogen atoms were generated using a computational method. We identified 114 and 14,347 potential N-H…N H-bonds from these two data sets, respectively, and 56-66% of these were of the Ni+1-Hi+1…Ni type, with Ni being the proline backbone nitrogen. To further understand the nature of such unusual contacts, we performed quantum chemical calculations on the model compound N-acetyl-L-proline-N-methylamide (Ace-Pro-NMe) with coordinates taken from the experimentally determined structures. A potential energy profile generated by varying the ψ dihedral angle in Ace-Pro-NMe indicates that the conformation with the N-H…N H-bond is the most stable. An analysis of H-bond-forming proline residues reveals that more than 30% of the proline carbonyl groups are also involved in n → π(∗) interactions with the carbonyl carbon of the preceding residue. Natural bond orbital analyses demonstrate that the strength of N-H…N H-bonds is less than half of that observed for a conventional H-bond. This study clearly establishes the H-bonding capability of proline nitrogen and its prevalence in protein structures. We found many proteins with multiple instances of H-bond-forming prolines. With more than 15% of all proline residues participating in N-H…N H-bonds, we suggest a new, to our knowledge, structural role for proline in providing stability to loops and capping regions of secondary structures in proteins.

  13. Identification of a Drug Targeting an Intrinsically Disordered Protein Involved in Pancreatic Adenocarcinoma

    PubMed Central

    Neira, José L.; Bintz, Jennifer; Arruebo, María; Rizzuti, Bruno; Bonacci, Thomas; Vega, Sonia; Lanas, Angel; Velázquez-Campoy, Adrián; Iovanna, Juan L.; Abián, Olga

    2017-01-01

    Intrinsically disordered proteins (IDPs) are prevalent in eukaryotes, performing signaling and regulatory functions. Often associated with human diseases, they constitute drug-development targets. NUPR1 is a multifunctional IDP, over-expressed and involved in pancreatic ductal adenocarcinoma (PDAC) development. By screening 1120 FDA-approved compounds, fifteen candidates were selected, and their interactions with NUPR1 were characterized by experimental and simulation techniques. The protein remained disordered upon binding to all fifteen candidates. These compounds were tested in PDAC-derived cell-based assays, and all induced cell-growth arrest and senescence, reduced cell migration, and decreased chemoresistance, mimicking NUPR1-deficiency. The most effective compound completely arrested tumor development in vivo on xenografted PDAC-derived cells in mice. Besides reporting the discovery of a compound targeting an intact IDP and specifically active against PDAC, our study proves the possibility to target the ‘fuzzy’ interface of a protein that remains disordered upon binding to its natural biological partners or to selected drugs. PMID:28054562

  14. Interatomic Coulombic Decay Effects in Theoretical DNA Recombination Systems Involving Protein Interaction Sites

    NASA Astrophysics Data System (ADS)

    Vargas, E. L.; Rivas, D. A.; Duot, A. C.; Hovey, R. T.; Andrianarijaona, V. M.

    2015-03-01

    DNA replication is the basis for all biological reproduction. A strand of DNA will ``unzip'' and bind with a complimentary strand, creating two identical strands. In this study, we are considering how this process is affected by Interatomic Coulombic Decay (ICD), specifically how ICD affects the individual coding proteins' ability to hold together. ICD mainly deals with how the electron returns to its original state after excitation and how this affects its immediate atomic environment, sometimes affecting the connectivity between interaction sites on proteins involved in the DNA coding process. Biological heredity is fundamentally controlled by DNA and its replication therefore it affects every living thing. The small nature of the proteins (within the range of nanometers) makes it a good candidate for research of this scale. Understanding how ICD affects DNA molecules can give us invaluable insight into the human genetic code and the processes behind cell mutations that can lead to cancer. Authors wish to give special thanks to Pacific Union College Student Senate in Angwin, California, for their financial support.

  15. A VAMP-associated protein, PVA31 is involved in leaf senescence in Arabidopsis

    PubMed Central

    Ichikawa, Mie; Nakai, Yusuke; Arima, Keita; Nishiyama, Sayo; Hirano, Tomoko; Sato, Masa H

    2015-01-01

    VAMP-associated proteins (VAPs) are highly conserved among eukaryotes. Here, we report a functional analysis of one of the VAPs, PVA31, and demonstrate its novel function on leaf senescence in Arabidopsis. The expression of PVA31 is highly induced in senescence leaves, and localizes to the plasma membrane as well as the ARA7-positive endosomes. Yeast two-hybrid analysis demonstrates that PVA31 is interacted with the plasma membrane localized-VAMP proteins, VAMP721/722/724 but not with the endosome-localized VAMPs, VAMP711 and VAMP727, indicating that PVA31 is associated with VAMP721/722/724 on the plasma membrane. Strong constitutive expression of PVA31 under the control of the Cauliflower mosaic virus 35S promoter induces the typical symptom of leaf senescence earlier than WT in normal growth and an artificially induced senescence conditions. In addition, the marker genes for the SA-mediated signaling pathways, PR-1, is promptly expressed with elicitor application. These data indicate that PVA31-overexpressing plants exhibit the early senescence phenotype in their leaves, and suggest that PVA31 is involved in the SA-mediated programmed cell death process during leaf senescence and PR-protein secretion during pathogen infection in Arabidopsis. PMID:25897470

  16. Plasmodium falciparum proteins involved in cytoadherence of infected erythrocytes to chemokine CX3CL1

    PubMed Central

    Hermand, Patricia; Cicéron, Liliane; Pionneau, Cédric; Vaquero, Catherine; Combadière, Christophe; Deterre, Philippe

    2016-01-01

    Malaria caused by Plasmodium falciparum is associated with cytoadherence of infected red blood cells (iRBC) to endothelial cells. Numerous host molecules have been involved in cytoadherence, including the adhesive chemokine CX3CL1. Most of the identified parasite ligands are from the multigenic and hypervariable Plasmodium falciparum Erythrocyte Membrane Protein 1 (PfEMP1) family which makes them poor targets for the development of a broadly protective vaccine. Using proteomics, we have identified two 25-kDa parasite proteins with adhesive properties for CX3CL1, called CBP for CX3CL1 Binding Proteins. CBPs are coded by single-copy genes with little polymorphic variation and no homology with other P. falciparum gene products. Specific antibodies raised against epitopes from the predicted extracellular domains of each CBP efficiently stain the surface of RBC infected with trophozoites or schizonts, which is a strong indication of CBP expression at the surface of iRBC. These anti-CBP antibodies partially neutralize iRBC adherence to CX3CL1. This adherence is similarly inhibited in the presence of peptides from the CBP extracellular domains, while irrelevant peptides had no such effect. CBP1 and CBP2 are new P. falciparum ligands for the human chemokine CX3CL1. The identification of this non-polymorphic P. falciparum factors provides a new avenue for innovative vaccination approaches. PMID:27653778

  17. A novel protein involved in heart development in Ambystoma mexicanum is localized in endoplasmic reticulum.

    PubMed

    Jia, P; Zhang, C; Huang, X P; Poda, M; Akbas, F; Lemanski, S L; Erginel-Unaltuna, N; Lemanski, L F

    2008-11-01

    The discovery of the naturally occurring cardiac non-function (c) animal strain in Ambystoma mexicanum (axolotl) provides a valuable animal model to study cardiomyocyte differentiation. In homozygous mutant animals (c/c), rhythmic contractions of the embryonic heart are absent due to a lack of organized myofibrils. We have previously cloned a partial sequence of a peptide cDNA (N1) from an anterior-endoderm-conditioned-medium RNA library that had been shown to be able to rescue the mutant phenotype. In the current studies we have fully cloned the N1 full length cDNA sequence from the library. N1 protein has been detected in both adult heart and skeletal muscle but not in any other adult tissues. GFP-tagged expression of the N1 protein has revealed localization of the N1 protein in the endoplasmic reticulum (ER). Results from in situ hybridization experiments have confirmed the dramatic decrease of expression of N1 mRNA in mutant (c/c) embryos indicating that the N1 gene is involved in heart development.

  18. Arabidopsis Membrane Steroid Binding Protein 1 Is Involved in Inhibition of Cell ElongationW⃞

    PubMed Central

    Yang, Xiao-Hua; Xu, Zhi-Hong; Xue, Hong-Wei

    2005-01-01

    A putative Membrane Steroid Binding Protein (designated MSBP1) was identified and functionally characterized as a negative regulator of cell elongation in Arabidopsis thaliana. The MSBP1 gene encodes a 220–amino acid protein that can bind to progesterone, 5-dihydrotestosterone, 24-epi-brassinolide (24-eBL), and stigmasterol with different affinities in vitro. Transgenic plants overexpressing MSBP1 showed short hypocotyl phenotype and increased steroid binding capacity in membrane fractions, whereas antisense MSBP1 transgenic plants showed long hypocotyl phenotypes and reduced steroid binding capacity, indicating that MSBP1 negatively regulates hypocotyl elongation. The reduced cell elongation of MSBP1-overexpressing plants was correlated with altered expression of genes involved in cell elongation, such as expansins and extensins, indicating that enhanced MSBP1 affected a regulatory pathway for cell elongation. Suppression or overexpression of MSBP1 resulted in enhanced or reduced sensitivities, respectively, to exogenous progesterone and 24-eBL, suggesting a negative role of MSBP1 in steroid signaling. Expression of MSBP1 in hypocotyls is suppressed by darkness and activated by light, suggesting that MSBP1, as a negative regulator of cell elongation, plays a role in plant photomorphogenesis. This study demonstrates the functional roles of a steroid binding protein in growth regulation in higher plants. PMID:15608331

  19. Abrogation of TNF-mediated cytotoxicity by space flight involves protein kinase C

    NASA Technical Reports Server (NTRS)

    Woods, K. M.; Chapes, S. K.; Spooner, B. S. (Principal Investigator)

    1994-01-01

    Experiments conducted on STS-50 indicated that space flight significantly inhibited tumor necrosis factor (TNF)-mediated killing of LM929 cells compared to ground controls. In ground-based studies, activation of protein kinase C (PKC) with phorbol 12-myristate 13-acetate (PMA) also inhibited TNF-mediated killing of LM929 cells. Therefore, we used PKC inhibitors to determine if the inhibitory effects of spaceflight on TNF-mediated cytotoxicity involved the activation of PKC. In experiments conducted onboard space shuttle mission STS-54, we saw that in the presence of the protein kinase C inhibitors H7 and H8, TNF-mediated cytotoxicity was restored to levels of those observed in the ground controls. Subsequent experiments done during the STS-57 mission tested the dose response of two protein kinase inhibitors, H7 and HA1004. We again saw that killing was restored in a dose-dependent manner, with inhibitor concentrations known to inhibit PKC being most effective. These data suggest that space flight ameliorates the action of TNF by affecting PKC in target cells.

  20. The Saccharomyces cerevisiae actin-related protein Arp2 is involved in the actin cytoskeleton

    PubMed Central

    1996-01-01

    Arp2p is an essential yeast actin-related protein. Disruption of the corresponding ARP2 gene leads to a terminal phenotype characterized by the presence of a single large bud. Thus, Arp2p may be important for a late stage of the cell cycle (Schwob, E., and R.P. Martin, 1992. Nature (Lond.). 355:179-182). We have localized Arp2p by indirect immunofluorescence. Specific peptide antibodies revealed punctate staining under the plasma membrane, which partially colocalizes with actin. Temperature-sensitive arp2 mutations were created by PCR mutagenesis and selected by an ade2/SUP11 sectoring screen. One temperature-sensitive mutant that was characterized, arp2-H330L, was osmosensitive and had an altered actin cytoskeleton at a nonpermissive temperature, suggesting a role of Arp2p in the actin cytoskeleton. Random budding patterns were observed in both haploid and diploid arp2- H330L mutant cells. Endocytosis, as judged by Lucifer yellow uptake, was severely reduced in the mutant, at all temperatures. In addition, genetic interaction was observed between temperature-sensitive alleles arp2-H330L and cdc10-1. CDC10 is a gene encoding a neck filament- associated protein that is necessary for polarized growth and cytokinesis. Overall, the immunolocalization, mutant phenotypes, and genetic interaction suggest that the Arp2 protein is an essential component of the actin cytoskeleton that is involved in membrane growth and polarity, as well as in endocytosis. PMID:8698808

  1. Identification of a Drug Targeting an Intrinsically Disordered Protein Involved in Pancreatic Adenocarcinoma.

    PubMed

    Neira, José L; Bintz, Jennifer; Arruebo, María; Rizzuti, Bruno; Bonacci, Thomas; Vega, Sonia; Lanas, Angel; Velázquez-Campoy, Adrián; Iovanna, Juan L; Abián, Olga

    2017-01-05

    Intrinsically disordered proteins (IDPs) are prevalent in eukaryotes, performing signaling and regulatory functions. Often associated with human diseases, they constitute drug-development targets. NUPR1 is a multifunctional IDP, over-expressed and involved in pancreatic ductal adenocarcinoma (PDAC) development. By screening 1120 FDA-approved compounds, fifteen candidates were selected, and their interactions with NUPR1 were characterized by experimental and simulation techniques. The protein remained disordered upon binding to all fifteen candidates. These compounds were tested in PDAC-derived cell-based assays, and all induced cell-growth arrest and senescence, reduced cell migration, and decreased chemoresistance, mimicking NUPR1-deficiency. The most effective compound completely arrested tumor development in vivo on xenografted PDAC-derived cells in mice. Besides reporting the discovery of a compound targeting an intact IDP and specifically active against PDAC, our study proves the possibility to target the 'fuzzy' interface of a protein that remains disordered upon binding to its natural biological partners or to selected drugs.

  2. HNE-modified proteins in Down syndrome: Involvement in development of Alzheimer disease neuropathology.

    PubMed

    Barone, Eugenio; Head, Elizabeth; Butterfield, D Allan; Perluigi, Marzia

    2016-11-10

    Down syndrome (DS), trisomy of chromosome 21, is the most common genetic form of intellectual disability. The neuropathology of DS involves multiple molecular mechanisms, similar to AD, including the deposition of beta-amyloid (Aβ) into senile plaques and tau hyperphosphorylationg in neurofibrillary tangles. Interestingly, many genes encoded by chromosome 21, in addition to being primarily linked to amyloid-beta peptide (Aβ) pathology, are responsible for increased oxidative stress (OS) conditions that also result as a consequence of reduced antioxidant system efficiency. However, redox homeostasis is disturbed by overproduction of Aβ, which accumulates into plaques across the lifespan in DS as well as in AD, thus generating a vicious cycle that amplifies OS-induced intracellular changes. The present review describes the current literature that demonstrates the accumulation of oxidative damage in DS with a focus on the lipid peroxidation by-product, 4-hydroxy-2-nonenal (HNE). HNE reacts with proteins and can irreversibly impair their functions. We suggest that among different post-translational modifications, HNE-adducts on proteins accumulate in DS brain and play a crucial role in causing the impairment of glucose metabolism, neuronal trafficking, protein quality control and antioxidant response. We hypothesize that dysfunction of these specific pathways contribute to accelerated neurodegeneration associated with AD neuropathology.

  3. Nuclear pore proteins are involved in the biogenesis of functional tRNA.

    PubMed Central

    Simos, G; Tekotte, H; Grosjean, H; Segref, A; Sharma, K; Tollervey, D; Hurt, E C

    1996-01-01

    Los1p and Pus1p, which are involved in tRNA biogenesis, were found in a genetic screen for components interacting with the nuclear pore protein Nsp1p. LOS1, PUS1 and NSP1 interact functionally, since the combination of mutations in the three genes causes synthetic lethality. Pus1p is an intranuclear protein which exhibits a nucleotide-specific and intron-dependent tRNA pseudouridine synthase activity. Los1p was shown previously to be required for efficient pre-tRNA splicing; we report here that Los1p localizes to the nuclear pores and is linked functionally to several components of the tRNA biogenesis machinery including Pus1p and Tfc4p. When the formation of functional tRNA was analyzed by an in vivo assay, the los1(-) pus1(-) double mutant, as well as several thermosensitive nucleoporin mutants including nsp1, nup116, nup133 and nup85, exhibited loss of suppressor tRNA activity even at permissive temperatures. These data suggest that nuclear pore proteins are required for the biogenesis of functional tRNA. Images PMID:8641292

  4. Adaptive expression pattern of different proteins involved in cellular calcium homeostasis in denervated rat vas deferens.

    PubMed

    Quintas, Luis Eduardo M; Cunha, Valéria M N; Scaramello, Christianne B V; da Silva, Cláudia L M; Caricati-Neto, Afonso; Lafayette, Simone S L; Jurkiewicz, Aron; Noël, François

    2005-11-21

    The activity and protein expression of plasma membrane and sarco(endo)plasmic reticulum (Ca2+-Mg2+)ATPases and ryanodine receptors were investigated in surgically denervated rat vas deferens. The function of thapsigargin-sensitive but not thapsigargin-resistant (Ca2+-Mg2+)ATPase (from sarco(endo)plasmic reticulum and plasma membrane, respectively), evidenced by enzyme activity and Ca2+ uptake experiments, was significantly depressed by 30-50% when compared to innervated vas. Western blots showed that such reduction in sarco(endo)plasmic reticulum (Ca2+-Mg2+)ATPase performance was accompanied by a decrement of similar magnitude in sarco(endo)plasmic reticulum (Ca2+-Mg2+)ATPase type 2 protein expression, without any significant change in plasma membrane (Ca2+-Mg2+)ATPase expression. Finally, [3H]ryanodine binding revealed that the density of ryanodine binding sites was reduced by 45% after denervation without modification in affinity. The present findings demonstrate that sarco(endo)plasmic reticulum proteins involved in intracellular calcium homeostasis are clearly down-regulated and brings further evidence of a modified calcium translocation in denervated rat vas deferens.

  5. Looking for prosocial genes: ITRAQ analysis of proteins involved in MDMA-induced sociability in mice.

    PubMed

    Kuteykin-Teplyakov, Konstantin; Maldonado, Rafael

    2014-11-01

    Social behavior plays a fundamental role in life of many animal species, allowing the interaction between individuals and sharing of experiences, needs, and goals across them. In humans, some neuropsychiatric diseases, including anxiety, posttraumatic stress disorder and autism spectrum disorders, are often characterized by impaired sociability. Here we report that N-Methyl-3,4-methylenedioxyamphetamine (MDMA, "Ecstasy") at low dose (3mg/kg) has differential effects on mouse social behavior. In some animals, MDMA promotes sociability without hyperlocomotion, whereas in other mice it elevates locomotor activity without affecting sociability. Both WAY-100635, a selective antagonist of 5-HT1A receptor, and L-368899, a selective oxytocin receptor antagonist, abolish prosocial effects of MDMA. Differential quantitative analysis of brain proteome by isobaric tag for relative and absolute quantification technology (iTRAQ) revealed 21 specific proteins that were highly correlated with sociability, and allowed to distinguish between entactogenic prosocial and hyperlocomotor effects of MDMA on proteome level. Our data suggest particular relevance of neurotransmission mediated by GABA B receptor, as well as proteins involved in energy maintenance for MDMA-induced sociability. Functional association network for differentially expressed proteins in cerebral cortex, hippocampus and amygdala were identified. These results provide new information for understanding the neurobiological substrate of sociability and may help to discover new therapeutic approaches to modulate social behavior in patients suffering from social fear and low sociability.

  6. Xanthorrhizol induced DNA fragmentation in HepG2 cells involving Bcl-2 family proteins

    SciTech Connect

    Tee, Thiam-Tsui; Cheah, Yew-Hoong; Meenakshii, Nallappan; Mohd Sharom, Mohd Yusof; Azimahtol Hawariah, Lope Pihie

    2012-04-20

    Highlights: Black-Right-Pointing-Pointer We isolated xanthorrhizol, a sesquiterpenoid compound from Curcuma xanthorrhiza. Black-Right-Pointing-Pointer Xanthorrhizol induced apoptosis in HepG2 cells as observed using SEM. Black-Right-Pointing-Pointer Apoptosis in xanthorrhizol-treated HepG2 cells involved Bcl-2 family proteins. Black-Right-Pointing-Pointer DNA fragmentation was observed in xanthorrhizol-treated HepG2 cells. Black-Right-Pointing-Pointer DNA fragmentation maybe due to cleavage of PARP and DFF45/ICAD proteins. -- Abstract: Xanthorrhizol is a plant-derived pharmacologically active sesquiterpenoid compound isolated from Curcuma xanthorrhiza. Previously, we have reported that xanthorrhizol inhibited the proliferation of HepG2 human hepatoma cells by inducing apoptotic cell death via caspase activation. Here, we attempt to further elucidate the mode of action of xanthorrhizol. Apoptosis in xanthorrhizol-treated HepG2 cells as observed by scanning electron microscopy was accompanied by truncation of BID; reduction of both anti-apoptotic Bcl-2 and Bcl-X{sub L} expression; cleavage of PARP and DFF45/ICAD proteins and DNA fragmentation. Taken together, these results suggest xanthorrhizol as a potent antiproliferative agent on HepG2 cells by inducing apoptosis via Bcl-2 family members. Hence we proposed that xanthorrhizol could be used as an anti-liver cancer drug for future studies.

  7. Identification of a Drug Targeting an Intrinsically Disordered Protein Involved in Pancreatic Adenocarcinoma

    NASA Astrophysics Data System (ADS)

    Neira, José L.; Bintz, Jennifer; Arruebo, María; Rizzuti, Bruno; Bonacci, Thomas; Vega, Sonia; Lanas, Angel; Velázquez-Campoy, Adrián; Iovanna, Juan L.; Abián, Olga

    2017-01-01

    Intrinsically disordered proteins (IDPs) are prevalent in eukaryotes, performing signaling and regulatory functions. Often associated with human diseases, they constitute drug-development targets. NUPR1 is a multifunctional IDP, over-expressed and involved in pancreatic ductal adenocarcinoma (PDAC) development. By screening 1120 FDA-approved compounds, fifteen candidates were selected, and their interactions with NUPR1 were characterized by experimental and simulation techniques. The protein remained disordered upon binding to all fifteen candidates. These compounds were tested in PDAC-derived cell-based assays, and all induced cell-growth arrest and senescence, reduced cell migration, and decreased chemoresistance, mimicking NUPR1-deficiency. The most effective compound completely arrested tumor development in vivo on xenografted PDAC-derived cells in mice. Besides reporting the discovery of a compound targeting an intact IDP and specifically active against PDAC, our study proves the possibility to target the ‘fuzzy’ interface of a protein that remains disordered upon binding to its natural biological partners or to selected drugs.

  8. Tau pathology involves protein phosphatase 2A in Parkinsonism-dementia of Guam

    PubMed Central

    Arif, Mohammad; Kazim, Syed Faraz; Grundke-Iqbal, Inge; Garruto, Ralph M.; Iqbal, Khalid

    2014-01-01

    Parkinsonism-dementia (PD) of Guam is a neurodegenerative disease with parkinsonism and early-onset Alzheimer-like dementia associated with neurofibrillary tangles composed of hyperphosphorylated microtubule-associated protein, tau. β-N-methylamino-l-alanine (BMAA) has been suspected of being involved in the etiology of PD, but the mechanism by which BMAA leads to tau hyperphosphorylation is not known. We found a decrease in protein phosphatase 2A (PP2A) activity associated with an increase in inhibitory phosphorylation of its catalytic subunit PP2Ac at Tyr307 and abnormal hyperphosphorylation of tau in brains of patients who had Guam PD. To test the possible involvement of BMAA in the etiopathogenesis of PD, we studied the effect of this environmental neurotoxin on PP2A activity and tau hyperphosphorylation in mouse primary neuronal cultures and metabolically active rat brain slices. BMAA treatment significantly decreased PP2A activity, with a concomitant increase in tau kinase activity resulting in elevated tau hyperphosphorylation at PP2A favorable sites. Moreover, we found an increase in the phosphorylation of PP2Ac at Tyr307 in BMAA-treated rat brains. Pretreatment with metabotropic glutamate receptor 5 (mGluR5) and Src antagonists blocked the BMAA-induced inhibition of PP2A and the abnormal hyperphosphorylation of tau, indicating the involvement of an Src-dependent PP2A pathway. Coimmunoprecipitation experiments showed that BMAA treatment dissociated PP2Ac from mGluR5, making it available for phosphorylation at Tyr307. These findings suggest a scenario in which BMAA can lead to tau pathology by inhibiting PP2A through the activation of mGluR5, the consequent release of PP2Ac from the mGluR5–PP2A complex, and its phosphorylation at Tyr307 by Src. PMID:24395787

  9. Effects of Radiation and Dietary Iron on Expression of Genes and Proteins Involved in Drug Metabolism

    NASA Technical Reports Server (NTRS)

    Faust, K. M.; Wotring, V. E.

    2014-01-01

    Liver function, especially the rate of metabolic enzyme activities, determines the concentration of circulating drugs and the duration of their efficacy. Most pharmaceuticals are metabolized by the liver, and clinically-used medication doses are given with normal liver function in mind. A drug overdose can result in the case of a liver that is damaged and removing pharmaceuticals from the circulation at a rate slower than normal. Alternatively, if liver function is elevated and removing drugs from the system more quickly than usual, it would be as if too little drug had been given for effective treatment. Because of the importance of the liver in drug metabolism, we want to understand any effects of spaceflight on the enzymes of the liver. Dietary factors and exposure to radiation are aspects of spaceflight that are potential oxidative stressors and both can be modeled in ground experiments. In this experiment, we examined the effects of high dietary iron and low dose gamma radiation (individually and combined) on the gene expression of enzymes involved in drug metabolism, redox homeostasis, and DNA repair. METHODS All procedures were approved by the JSC Animal Care and Use Committee. Male Sprague-Dawley rats were divided into 4 groups (n=8); control, high Fe diet (650 mg iron/kg), radiation (fractionated 3 Gy exposure from a Cs- 137 source) and combined high Fe diet + radiation exposure. Animals were euthanized 24h after the last treatment of radiation; livers were removed immediately and flash -frozen in liquid nitrogen. Expression of genes thought to be involved in redox homeostasis, drug metabolism and DNA damage repair was measured by RT-qPCR. Where possible, protein expression of the same genes was measured by western blotting. All data are expressed as % change in expression normalized to reference gene expression; comparisons were then made of each treatment group to the sham exposed/ normal diet control group. Data was considered significant at p< 0

  10. Involvement of cyclic nucleotide-dependent protein kinases in cyclic AMP-mediated vasorelaxation

    PubMed Central

    Eckly-Michel, Anita; Martin, Viviane; Lugnier, Claire

    1997-01-01

    The involvement of cyclic AMP-dependent protein kinase (PKA) and cyclic GMP-dependent protein kinase (PKG) in the effects of cyclic AMP-elevating agents on vascular smooth muscle relaxation, cyclic nucleotide dependent-protein kinase activities and ATP-induced calcium signalling ([Ca2+]i) was studied in rat aorta. Cyclic AMP-elevating agents used were a β-adrenoceptor agonist (isoprenaline), a phosphodiesterase 3 (PDE3) inhibitor (SK&F 94120) and a PDE4 inhibitor (rolipram). In rat intact aorta, the relaxant effect induced by isoprenaline (0.01–0.3 μM) was decreased by a specific inhibitor of PKA, H-89, whereas a specific inhibitor of PKG, Rp-8-Br-cyclic GMPS, was without effect. No significant difference in PKA and PKG activity ratios was detected in aortic rings when isoprenaline 10 μM was used. At the same concentration, isoprenaline did not modify ATP-induced changes in [Ca2+]i in smooth muscle cells. Neither H-89 nor Rp-8-Br-cyclic GMPS modified this response. These findings suggest that PKA is only involved in the relaxant effect induced by low concentrations of isoprenaline (0.01–0.3 μM), whereas for higher concentrations, other mechanisms independent of PKA and PKG are involved. The relaxant effects induced by SK&F 94120 and rolipram were inhibited by Rp-8-Br-cyclic GMPS with no significant effect of H-89. Neither SK&F 94120, nor rolipram at 30 μM significantly modified the activity ratios of PKA and PKG. Rolipram inhibited the ATP-induced transient increase in [Ca2+]i. This decrease was abolished by Rp-8-Br-cyclic GMPS whereas H-89 had no significant effect. These results suggest that PKG is involved in the vascular effects induced by the inhibitors of PDE3 and PDE4. Moreover, since it was previously shown that PDE3 and PDE4 inhibitors only increased cyclic AMP levels with no change in cyclic GMP level, these data also suggest a cross-activation of PKG by cyclic AMP in rat aorta. The combination of 5 μM SK&F 94120 with rolipram markedly

  11. Construction of protein interaction network involved in lung adenocarcinomas using a novel algorithm

    PubMed Central

    Chen, Juan; Yang, Hai-Tao; Li, Zhu; Xu, Ning; Yu, Bo; Xu, Jun-Ping; Zhao, Pei-Ge; Wang, Yan; Zhang, Xiu-Juan; Lin, Dian-Jie

    2016-01-01

    Studies that only assess differentially-expressed (DE) genes do not contain the information required to investigate the mechanisms of diseases. A complete knowledge of all the direct and indirect interactions between proteins may act as a significant benchmark in the process of forming a comprehensive description of cellular mechanisms and functions. The results of protein interaction network studies are often inconsistent and are based on various methods. In the present study, a combined network was constructed using selected gene pairs, following the conversion and combination of the scores of gene pairs that were obtained across multiple approaches by a novel algorithm. Samples from patients with and without lung adenocarcinoma were compared, and the RankProd package was used to identify DE genes. The empirical Bayesian (EB) meta-analysis approach, the search tool for the retrieval of interacting genes/proteins database (STRING), the weighted gene coexpression network analysis (WGCNA) package and the differentially-coexpressed genes and links package (DCGL) were used for network construction. A combined network was also constructed with a novel rank-based algorithm using a combined score. The topological features of the 5 networks were analyzed and compared. A total of 941 DE genes were screened. The topological analysis indicated that the gene interaction network constructed using the WGCNA method was more likely to produce a small-world property, which has a small average shortest path length and a large clustering coefficient, whereas the combined network was confirmed to be a scale-free network. Gene pairs that were identified using the novel combined method were mostly enriched in the cell cycle and p53 signaling pathway. The present study provided a novel perspective to the network-based analysis. Each method has advantages and disadvantages. Compared with single methods, the combined algorithm used in the present study may provide a novel method to

  12. TEC protein tyrosine kinase is involved in the Erk signaling pathway induced by HGF

    SciTech Connect

    Li, Feifei; Jiang, Yinan; Zheng, Qiping; Yang, Xiaoming; Wang, Siying

    2011-01-07

    Research highlights: {yields} TEC is rapidly tyrosine-phosphorylated and activated by HGF-stimulation in vivo or after partial hepatectomy in mice. {yields} TEC enhances the activity of Elk and serum response element (SRE) in HGF signaling pathway in hepatocyte. {yields} TEC promotes hepatocyte proliferation through the Erk-MAPK pathway. -- Abstract: Background/aims: TEC, a member of the TEC family of non-receptor type protein tyrosine kinases, has recently been suggested to play a role in hepatocyte proliferation and liver regeneration. This study aims to investigate the putative mechanisms of TEC kinase regulation of hepatocyte differentiation, i.e. to explore which signaling pathway TEC is involved in, and how TEC is activated in hepatocyte after hepatectomy and hepatocyte growth factor (HGF) stimulation. Methods: We performed immunoprecipitation (IP) and immunoblotting (IB) to examine TEC tyrosine phosphorylation after partial hepatectomy in mice and HGF stimulation in WB F-344 hepatic cells. The TEC kinase activity was determined by in vitro kinase assay. Reporter gene assay, antisense oligonucleotide and TEC dominant negative mutant (TEC{sup KM}) were used to examine the possible signaling pathways in which TEC is involved. The cell proliferation rate was evaluated by {sup 3}H-TdR incorporation. Results: TEC phosphorylation and kinase activity were increased in 1 h after hepatectomy or HGF treatment. TEC enhanced the activity of Elk and serum response element (SRE). Inhibition of MEK1 suppressed TEC phosphorylation. Blocking TEC activity dramatically decreased the activation of Erk. Reduced TEC kinase activity also suppressed the proliferation of WB F-344 cells. These results suggest TEC is involved in the Ras-MAPK pathway and acts between MEK1 and Erk. Conclusions: TEC promotes hepatocyte proliferation and regeneration and is involved in HGF-induced Erk signaling pathway.

  13. Statistical analysis of crystallization database links protein physico-chemical features with crystallization mechanisms.

    PubMed

    Fusco, Diana; Barnum, Timothy J; Bruno, Andrew E; Luft, Joseph R; Snell, Edward H; Mukherjee, Sayan; Charbonneau, Patrick

    2014-01-01

    X-ray crystallography is the predominant method for obtaining atomic-scale information about biological macromolecules. Despite the success of the technique, obtaining well diffracting crystals still critically limits going from protein to structure. In practice, the crystallization process proceeds through knowledge-informed empiricism. Better physico-chemical understanding remains elusive because of the large number of variables involved, hence little guidance is available to systematically identify solution conditions that promote crystallization. To help determine relationships between macromolecular properties and their crystallization propensity, we have trained statistical models on samples for 182 proteins supplied by the Northeast Structural Genomics consortium. Gaussian processes, which capture trends beyond the reach of linear statistical models, distinguish between two main physico-chemical mechanisms driving crystallization. One is characterized by low levels of side chain entropy and has been extensively reported in the literature. The other identifies specific electrostatic interactions not previously described in the crystallization context. Because evidence for two distinct mechanisms can be gleaned both from crystal contacts and from solution conditions leading to successful crystallization, the model offers future avenues for optimizing crystallization screens based on partial structural information. The availability of crystallization data coupled with structural outcomes analyzed through state-of-the-art statistical models may thus guide macromolecular crystallization toward a more rational basis.

  14. Statistical Analysis of Crystallization Database Links Protein Physico-Chemical Features with Crystallization Mechanisms

    PubMed Central

    Fusco, Diana; Barnum, Timothy J.; Bruno, Andrew E.; Luft, Joseph R.; Snell, Edward H.; Mukherjee, Sayan; Charbonneau, Patrick

    2014-01-01

    X-ray crystallography is the predominant method for obtaining atomic-scale information about biological macromolecules. Despite the success of the technique, obtaining well diffracting crystals still critically limits going from protein to structure. In practice, the crystallization process proceeds through knowledge-informed empiricism. Better physico-chemical understanding remains elusive because of the large number of variables involved, hence little guidance is available to systematically identify solution conditions that promote crystallization. To help determine relationships between macromolecular properties and their crystallization propensity, we have trained statistical models on samples for 182 proteins supplied by the Northeast Structural Genomics consortium. Gaussian processes, which capture trends beyond the reach of linear statistical models, distinguish between two main physico-chemical mechanisms driving crystallization. One is characterized by low levels of side chain entropy and has been extensively reported in the literature. The other identifies specific electrostatic interactions not previously described in the crystallization context. Because evidence for two distinct mechanisms can be gleaned both from crystal contacts and from solution conditions leading to successful crystallization, the model offers future avenues for optimizing crystallization screens based on partial structural information. The availability of crystallization data coupled with structural outcomes analyzed through state-of-the-art statistical models may thus guide macromolecular crystallization toward a more rational basis. PMID:24988076

  15. Endocervical glandular involvement, multicentricity, and extent of the disease are features of high-grade cervical intraepithelial neoplasia.

    PubMed

    Güdücü, Nilgün; Sidar, Güliz; Başsüllü, Nuray; Türkmen, Ilknur; Dünder, Ilkkan

    2013-08-01

    This study aimed to evaluate the rate of endocervical glandular involvement, positive surgical margins, multicentricity, and disease extent between low-grade and high-grade cervical intraepithelial lesions after loop electrosurgical excision procedure (LEEP). Pathology medical records of patients who underwent LEEP were reviewed retrospectively. Patients with negative LEEP results were excluded. Loop electrosurgical excision procedure reports of patients with cervical intraepithelial neoplasia (CIN) 1, 2, and 3 were compared. There was no statistically significant difference between patients with CIN 1 (n=24), CIN 2 (n=27), and CIN 3 (n=64) when age and surgical margin positivity were considered. Endocervical glandular involvement, multicentricity, and disease extent were higher in patients with CIN 3 (P=.001, P=.002, and P=.001, respectively). In conclusion, we recommend that patients with endocervical glandular involvement, lesions involving more than two-thirds of the LEEP specimen, and multicentricity be followed up more closely.

  16. Probing the Sites of Interactions of Rotaviral Proteins Involved in Replication

    PubMed Central

    Viskovska, Maria; Anish, Ramakrishnan; Hu, Liya; Chow, Dar-Chone; Hurwitz, Amy M.; Brown, Nicholas G.; Palzkill, Timothy; Estes, Mary K.

    2014-01-01

    ABSTRACT Replication and packaging of the rotavirus genome occur in cytoplasmic compartments called viroplasms, which form during virus infection. These processes are orchestrated by yet-to-be-understood complex networks of interactions involving nonstructural proteins (NSPs) 2, 5, and 6 and structural proteins (VPs) 1, 2, 3, and 6. The multifunctional enzyme NSP2, an octamer with RNA binding activity, is critical for viroplasm formation with its binding partner, NSP5, and for genome replication/packaging through its interactions with replicating RNA, the viral polymerase VP1, and the inner core protein VP2. Using isothermal calorimetry, biolayer interferometry, and peptide array screening, we examined the interactions between NSP2, VP1, VP2, NSP5, and NSP6. These studies provide the first evidence that NSP2 can directly bind to VP1, VP2, and NSP6, in addition to the previously known binding to NSP5. The interacting sites identified from reciprocal peptide arrays were found to be in close proximity to the RNA template entry and double-stranded RNA (dsRNA) exit tunnels of VP1 and near the catalytic cleft and RNA-binding grooves of NSP2; these sites are consistent with the proposed role of NSP2 in facilitating dsRNA synthesis by VP1. Peptide screening of VP2 identified NSP2-binding sites in the regions close to the intersubunit junctions, suggesting that NSP2 binding could be a regulatory mechanism for preventing the premature self-assembly of VP2. The binding sites on NSP2 for NSP6 were found to overlap that of VP1, and the NSP5-binding sites overlap those of VP2 and VP1, suggesting that interaction of these proteins with NSP2 is likely spatially and/or temporally regulated. IMPORTANCE Replication and packaging of the rotavirus genome occur in cytoplasmic compartments called viroplasms that form during virus infection and are orchestrated by complex networks of interactions involving nonstructural proteins (NSPs) and structural proteins (VPs). A multifunctional RNA

  17. Regulatory elements and structural features of Beta vulgaris polygalacturonase-inhibiting protein gene for fungal and pest control

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Polygalacturonase-inhibiting proteins (PGIPs) are involved in plant defense. PGIPs are cell wall leucine-rich repeat (LRR) proteins that are known to inhibit pathogen and pest polygalacturonases (PGs) during the infection process. Several sugar beet (Beta vulgaris L.) PGIP genes (BvPGIP) were clon...

  18. The Chlamydomonas reinhardtii Nar1 Gene Encodes a Chloroplast Membrane Protein Involved in Nitrite Transport

    PubMed Central

    Rexach, Jesus; Fernández, Emilio; Galván, Aurora

    2000-01-01

    A key step for nitrate assimilation in photosynthetic eukaryotes occurs within chloroplasts, where nitrite is reduced to ammonium, which is incorporated into carbon skeletons. The Nar1 gene from Chlamydomonas reinhardtii is clustered with five other genes for nitrate assimilation, all of them regulated by nitrate. Sequence analysis of genomic DNA and cDNA of Nar1 and comparative studies of strains having or lacking Nar1 have been performed. The deduced amino acid sequence indicates that Nar1 encodes a chloroplast membrane protein with substantial identity to putative formate and nitrite transporters in bacteria. Use of antibodies against NAR1 has corroborated its location in the plastidic membrane. Characterization of strains having or lacking this gene suggests that NAR1 is involved in nitrite transport in plastids, which is critical for cell survival under limiting nitrate conditions, and controls the amount of nitrate incorporated by the cells under limiting CO2 conditions. PMID:10948261

  19. AN ODORANT-BINDING PROTEIN INVOLVED IN PERCEPTION OF HOST PLANT ODORANTS IN LOCUST Locusta migratoria.

    PubMed

    Li, Jia; Zhang, Long; Wang, Xiaoqi

    2016-04-01

    Locusts, Locusta migratoria (Orthoptera: Acrididae), are extremely destructive agricultural pests, but very little is known of their molecular aspects of perception to host plant odorants including related odorant-binding proteins (OBPs), though several OBPs have been identified in locust. To elucidate the function of LmigOBP1, the first OBP identified from locust, RNA interference was employed in this study to silence LmigOBP1, which was achieved by injection of dsRNA targeting LmigOBP1 into the hemolymph of male nymphs. Compared with LmigOBP1 normal nymphs, LmigOBP1 knockdown nymphs significantly decreased food (maize leaf, Zea mays) consumption and electro-antennography responses to five maize leaf volatiles, ((Z)-3-hexenol, linalool, nonanal, decanal, and (Z)-3-hexenyl acetate). These suggest that LmigOBP1 is involved in perception of host plant odorants.

  20. Clinicopathologic and molecular features of sporadic early-onset colorectal adenocarcinoma: an adenocarcinoma with frequent signet ring cell differentiation, rectal and sigmoid involvement, and adverse morphologic features.

    PubMed

    Chang, Daniel T; Pai, Rish K; Rybicki, Lisa A; Dimaio, Michael A; Limaye, Maneesha; Jayachandran, Priya; Koong, Albert C; Kunz, Pamela A; Fisher, George A; Ford, James M; Welton, Mark; Shelton, Andrew; Ma, Lisa; Arber, Daniel A; Pai, Reetesh K

    2012-08-01

    Recent literature suggests an increasing incidence of colorectal carcinoma in young patients. We performed a histologic, molecular, and immunophenotypic analysis of patients with sporadic early-onset (≤40 years of age) colorectal carcinoma seen at our institution from the years 2000-2010 and compared these tumors to a cohort of consecutively resected colorectal carcinomas seen in patients >40 years of age. A total of 1160 primary colorectal adenocarcinomas were surgically resected for the years 2000 through 2010. Of these, 75 (6%) were diagnoses in patients ≤40 years of age of which 13 (17%) demonstrated abnormalities in DNA mismatch repair, 4 (5%) were in patients with known germline genetic disorders (two patients with familial adenomatous polyposis, one patient with juvenile polyposis, and one patient with Li-Fraumeni syndrome), and three patients (4%) had long-standing chronic inflammatory bowel disease. The sporadic early-onset colorectal carcinoma group comprised a total of 55 patients (55/1160, 5%) and were compared with a control group comprising 73 consecutively resected colorectal carcinomas with proficient DNA mismatch repair in patients >40 years of age. For the early-onset colorectal carcinoma group, most cases (33/55, 60%) were diagnosed between the age of 35 and 40 years of age. Compared with the control group, the early-onset colorectal carcinoma group was significantly different with respect to tumor location (P<0.007) with 80% (44/55 cases) identified in either the sigmoid colon (24/55, 44%) or rectum (20/55, 36%). Morphologically, early-onset colorectal carcinomas more frequently displayed adverse histologic features compared with the control colorectal carcinoma group such as signet ring cell differentiation (7/55, 13% vs 1/73, 1%, P=0.021), perineural invasion (16/55, 29% vs 8/73, 11%, P=0.009) and venous invasion (12/55, 22% vs 4/73, 6%, P=0.006). A precursor adenomatous lesion was less frequently identified in the early-onset colorectal

  1. Involvement of retinoblastoma-associated protein 48 during photodynamic therapy of cervical cancer cells.

    PubMed

    Wu, Shuxia; Wang, Lijun; Ren, Xingye; Pan, Yulu; Peng, Yan; Zou, Xiaoyan; Shi, Cuige; Zhang, Youzhong

    2017-03-01

    5-Aminolevulinic acid-mediated photodynamic therapy (ALA‑PDT) is an effective treatment option for cervical intraepithelial neoplasia, the precancerous lesion of cervical cancer, and early cervical cancer, particularly for young or nulliparous women who want to remain fertile. A previous report described the involvement of histone deacetylases (HDAC) during ALA‑PDT mediated apoptosis in the cerebral cortex of a mouse model. Retinoblastoma‑associated protein 48 (RbAp48), a highly abundant component of HDACs, is a critical mediator that controls the transforming activity of human papillomavirus 16 in cervical cancer cells. The aim of the present study was to investigate the involvement of RbAp48 in ALA‑PDT‑induced cell death in cervical cancer cells. RbAp48 was significantly upregulated in cervical cancer cell lines treated with ALA‑PDT, including SiHa and HeLa cells. To establish the relevance of RbAp48 and the efficacy of ALA‑PDT in cervical cancer cells, the effect of ALA‑PDT was investigated in SiHa or HeLa cells following the depletion of RbAp48 by small interfering RNA (siRNA). Reduction of RbAp48 led to the reduced suppression of proliferation and apoptosis induced by ALA‑PDT in cervical cancer cells, which was associated with a reduction in tumor suppressor protein 53 (p53), retinoblastoma (Rb), apoptosis‑related enzyme caspase‑3, and increased levels of the oncogenic genes, human papillomavirus E6 and E7. These results provide evidence that RbAp48 is an important contributor to the efficacy of ALA‑PDT in cervical cancer cells. RbAp48 may be a therapeutic target that may help to improve the treatment of cervical cancer.

  2. Matrix Gla protein is involved in elastic fiber calcification in the dermis of pseudoxanthoma elasticum patients.

    PubMed

    Gheduzzi, Dealba; Boraldi, Federica; Annovi, Giulia; DeVincenzi, Chiara Paolinelli; Schurgers, Leon J; Vermeer, Cees; Quaglino, Daniela; Ronchetti, Ivonne Pasquali

    2007-10-01

    Mature MGP (Matrix gamma-carboxyglutamic acid protein) is known to inhibit soft connective tissues calcification. We investigated its possible involvement in pseudoxanthoma elasticum (PXE), a genetic disorder whose clinical manifestations are due to mineralization of elastic fibers. PXE patients have lower serum concentration of total MGP compared to controls (P<0.001). Antibodies specific for the noncarboxylated (Glu-MGP) and for the gamma-carboxylated (Gla-MGP) forms of MGP were assayed on ultrathin sections of dermis from controls and PXE patients. Normal elastic fibers in controls and patients were slightly positive for both forms of MGP, whereas Gla-MGP was more abundant within control's than within patient's elastic fibers (P<0.001). In patients' calcified elastic fibers, Glu-MGP intensively colocalized with mineral precipitates, whereas Gla-MGP precisely localized at the mineralization front. Data suggest that MGP is present within elastic fibers and is associated with calcification of dermal elastic fibers in PXE. To investigate whether local cells produce MGP, dermal fibroblasts were cultured in vitro and MGP was assayed at mRNA and protein levels. In spite of very similar MGP mRNA expression, cells from PXE patients produced 30% less of Gla-MGP compared to controls. Data were confirmed by immunocytochemistry on ultrathin sections. Normal fibroblasts in vitro were positive for both forms of MGP. PXE fibroblasts were positive for Glu-MGP and only barely positive for Gla-MGP (P<0.001). In conclusion, MGP is involved in elastic fiber calcification in PXE. The lower ratio of Gla-MGP over Glu-MGP in pathological fibroblasts compared to controls suggests these cells may play an important role in the ectopic calcification in PXE.

  3. Involvement of eicosanoids and surfactant protein D in extrinsic allergic alveolitis.

    PubMed

    Higashi, A; Higashi, N; Tsuburai, T; Takeuchi, Y; Taniguchi, M; Mita, H; Saito, A; Takatori, K; Arimura, K; Akiyama, K

    2005-12-01

    The pathophysiology of extrinsic allergic alveolitis (EAA) involves oxidative lung damage as well as interstitial and alveolar inflammation. Macrophages and mast cells are inflammatory components of EAA that produce both leukotrienes (LTs) and prostaglandin D2 (PGD2). In addition, PGD2 is also produced by the free-radical-catalysed peroxidation of arachidonic acid during oxidative stress. Urinary 8-iso prostaglandin F2alpha (8-isoPGF2alpha) and serum surfactant protein D (SP-D) are considered appropriate biomarkers of oxidative stress and interstitial lung disease activity, respectively. The present study aimed to assess the association of these biomarkers with the pathophysiology of EAA. Two cases of acute EAA caused by the inhalation of fungi spores were reported. Eight asthmatic patients and six healthy control subjects were also enrolled in the current study. The serum SP-D and urinary eicosanoid (LTE4, PGD2 metabolite (9alpha,11betaPGF2), 8-isoPGF2alpha) concentrations markedly increased during the acute exacerbation phase. These concentrations decreased following corticosteroid therapy in the EAA patients. There was a significant correlation between serum SP-D and urinary 9alpha,11betaPGF2 concentrations in the EAA patients. In conclusion, although the present study proposes that serum surfactant protein-D and urinary eicosanoids are new biomarkers involved in the various immunological responses in extrinsic allergic alveolitis, further large-scale studies are needed to investigate the role of these compounds, not just as biomarkers, but also as biological potentiators of extrinsic allergic alveolitis.

  4. Resveratrol Administration Increases Transthyretin Protein Levels, Ameliorating AD Features: The Importance of Transthyretin Tetrameric Stability

    PubMed Central

    Santos, Luís Miguel; Rodrigues, Daniela; Alemi, Mobina; Silva, Sara Costa; Ribeiro, Carlos Alexandre; Cardoso, Isabel

    2016-01-01

    Previous in vivo work showed that resveratrol has beneficial effects in Alzheimer’s disease (AD) pathology, resulting in increased expression of transthyretin (TTR). TTR binds amyloid-beta (Aβ) peptide, avoiding its aggregation and toxicity, and is reduced in the cerebrospinal fluid (CSF) and plasma in AD. Further, resveratrol binds TTR, stabilizing the native TTR tetrameric structure. To further explore the mechanism of neuroprotection conferred by TTR in AD, resveratrol was administered in the diet to 5- to 8-month-old AD transgenic female mice carrying just 1 copy of the mouse TTR gene for 2 months. Effects in brain Aβ burden were evaluated by immunohistochemistry, and total brain Aβ levels by ELISA, showing a striking decrease in both parameters in treated animals. In addition, total brain lipoprotein-related receptor protein 1 (LRP1) levels were increased in treated animals, although its gene expression was unaltered. To further understand the mechanism(s) underlying such improvement in AD features, we measured TTR plasma levels, showing that TTR increased in resveratrol-treated mice, whereas liver TTR gene transcription was not altered. These results strengthen the stability hypothesis, which postulates that TTR is unstable in AD, leading to accelerated clearance and lower levels. Therefore, resveratrol, which stabilizes the TTR tetramer results in TTR-normalized clearance, increases the protein plasma levels. In turn, stabilized TTR binds more strongly to Aβ peptide, avoiding its aggregation. Our results represent a step forward in the understanding of the mechanism underlying TTR protection in AD and highlight the possibility of using TTR stabilization as a therapeutic target in AD. PMID:27385446

  5. Involvement of protein kinase C in the response of Neurospora crassa to blue light.

    PubMed

    Arpaia, G; Cerri, F; Baima, S; Macino, G

    1999-09-01

    As a first step towards understanding the process of blue light perception, and the signal transduction mechanisms involved, in Neurospora crassa we have used a pharmacological approach to screen a wide range of second messengers and chemical compounds known to interfere with the activity of well-known signal transducing molecules in vivo. We tested the influence of these compounds on the induction of the al-3 gene, a key step in light-induced carotenoid biosynthesis. This approach has implicated protein kinase C (PKC) as a component of the light transduction machinery. The conclusion is based on the effects of specific inhibitors (calphostin C and chelerythrine chloride) and activators of PKC (1,2-dihexanoyl-sn-glycerol). During vegetative growth PKC may be responsible for desensitization to light because inhibitors of the enzyme cause an increase in the total amount of mRNA transcribed after illumination. PKC is therefore proposed here to be an important regulator of transduction of the blue light signal, and may act through modification of the protein White Collar-1, which we show to be a substrate for PKC in N. crassa.

  6. A Wiskott-Aldrich syndrome protein is involved in endocytosis in Aspergillus nidulans.

    PubMed

    Hoshi, Hiro-Omi; Zheng, Lu; Ohta, Akinori; Horiuchi, Hiroyuki

    2016-09-01

    Endocytosis is vital for hyphal tip growth in filamentous fungi and is involved in the tip localization of various membrane proteins. To investigate the function of a Wiskott-Aldrich syndrome protein (WASP) in endocytosis of filamentous fungi, we identified a WASP ortholog-encoding gene, wspA, in Aspergillus nidulans and characterized it. The wspA product, WspA, localized to the tips of germ tubes during germination and actin rings in the subapical regions of mature hyphae. wspA is essential for the growth and functioned in the polarity establishment and maintenance during germination of conidia. We also investigated its function in endocytosis and revealed that endocytosis of SynA, a synaptobrevin ortholog that is known to be endocytosed at the subapical regions of hyphal tips in A. nidulans, did not occur when wspA expression was repressed. These results suggest that WspA plays roles in endocytosis at hyphal tips and polarity establishment during germination.

  7. HTLV-1 Tax-mediated TAK1 activation involves TAB2 adapter protein

    SciTech Connect

    Yu Qingsheng; Minoda, Yasumasa; Yoshida, Ryoko; Yoshida, Hideyuki; Iha, Hidekatsu; Kobayashi, Takashi; Yoshimura, Akihiko; Takaesu, Giichi

    2008-01-04

    Human T cell leukemia virus type 1 (HTLV-1) Tax is an oncoprotein that plays a crucial role in the proliferation and transformation of HTLV-1-infected T lymphocytes. It has recently been reported that Tax activates a MAPKKK family, TAK1. However, the molecular mechanism of Tax-mediated TAK1 activation is not well understood. In this report, we investigated the role of TAK1-binding protein 2 (TAB2) in Tax-mediated TAK1 activation. We found that TAB2 physically interacts with Tax and augments Tax-induced NF-{kappa}B activity. Tax and TAB2 cooperatively activate TAK1 when they are coexpressed. Furthermore, TAK1 activation by Tax requires TAB2 binding as well as ubiquitination of Tax. We also found that the overexpression of TRAF2, 5, or 6 strongly induces Tax ubiquitination. These results suggest that TAB2 may be critically involved in Tax-mediated activation of TAK1 and that NF-{kappa}B-activating TRAF family proteins are potential cellular E3 ubiquitin ligases toward Tax.

  8. Cellular COPII Proteins Are Involved in Production of the Vesicles That Form the Poliovirus Replication Complex

    PubMed Central

    Rust, René C.; Landmann, Lukas; Gosert, Rainer; Tang, Bor Luen; Hong, Wanjin; Hauri, Hans-Peter; Egger, Denise; Bienz, Kurt

    2001-01-01

    Poliovirus (PV) replicates its genome in association with membranous vesicles in the cytoplasm of infected cells. To elucidate the origin and mode of formation of PV vesicles, immunofluorescence labeling with antibodies against the viral vesicle marker proteins 2B and 2BC, as well as cellular markers of the endoplasmic reticulum (ER), anterograde transport vesicles, and the Golgi complex, was performed in BT7-H cells. Optical sections obtained by confocal laser scanning microscopy were subjected to a deconvolution process to enhance resolution and signal-to-noise ratio and to allow for a three-dimensional representation of labeled membrane structures. The mode of formation of the PV vesicles was, on morphological grounds, similar to the formation of anterograde membrane traffic vesicles in uninfected cells. ER-resident membrane markers were excluded from both types of vesicles, and the COPII components Sec13 and Sec31 were both found to be colocalized on the vesicular surface, indicating the presence of a functional COPII coat. PV vesicle formation during early time points of infection did not involve the Golgi complex. The expression of PV protein 2BC or the entire P2 and P3 genomic region led to the production of vesicles carrying a COPII coat and showing the same mode of formation as vesicles produced after PV infection. These results indicate that PV vesicles are formed at the ER by the cellular COPII budding mechanism and thus are homologous to the vesicles of the anterograde membrane transport pathway. PMID:11559814

  9. Involvement of the cellular prion protein in the migration of brain microvascular endothelial cells.

    PubMed

    Watanabe, Takuya; Yasutaka, Yuki; Nishioku, Tsuyoshi; Kusakabe, Sae; Futagami, Koujiro; Yamauchi, Atsushi; Kataoka, Yasufumi

    2011-06-01

    The conversion of cellular prion protein (PrP(C)) to its protease-resistant isoform is involved in the pathogenesis of prion disease. Although PrP(C) is a ubiquitous glycoprotein that is present in various cell types, the physiological role of PrP(C) remains obscure. The present study aimed to determine whether PrP(C) mediates migration of brain microvascular endothelial cells. Small interfering RNAs (siRNAs) targeting PrP(C) were transfected into a mouse brain microvascular endothelial cell line (bEND.3 cells). siPrP1, selected among three siRNAs, reduced mRNA and protein levels of PrP(C) in bEND.3 cells. Cellular migration was evaluated with a scratch-wound assay. siPrP1 suppressed migration without significantly affecting cellular proliferation. This study provides the first evidence that PrP(C) may be necessary for brain microvascular endothelial cells to migrate into damaged regions in the brain. This function of PrP(C) in the brain endothelium may be a mechanism by which the neurovascular unit recovers from an injury such as an ischemic insult.

  10. Equatorial segment protein (ESP) is a human alloantigen involved in sperm-egg binding and fusion.

    PubMed

    Wolkowicz, M J; Digilio, L; Klotz, K; Shetty, J; Flickinger, C J; Herr, J C

    2008-01-01

    The equatorial segment of the sperm head is known to play a role in fertilization; however, the specific sperm molecules contributing to the integrity of the equatorial segment and in binding and fusion at the oolemma remain incomplete. Moreover, identification of molecular mediators of fertilization that are also immunogenic in humans is predicted to advance both the diagnosis and treatment of immune infertility. We previously reported the cloning of Equatorial Segment Protein (ESP), a protein localized to the equatorial segment of ejaculated human sperm. ESP is a biomarker for a subcompartment of the acrosomal matrix that can be traced through all stages of acrosome biogenesis (Wolkowicz et al, 2003). In the present study, ESP immunoreacted on Western blots with 4 (27%) of 15 antisperm antibody (ASA)-positive serum samples from infertile male patients and 2 (40%) of 5 ASA-positive female sera. Immunofluorescent studies revealed ESP in the equatorial segment of 89% of acrosome-reacted sperm. ESP persisted as a defined equatorial segment band on 100% of sperm tightly bound to the oolemma of hamster eggs. Antisera to recombinant human ESP inhibited both oolemmal binding and fusion of human sperm in the hamster egg penetration assay. The results indicate that ESP is a human alloantigen involved in sperm-egg binding and fusion. Defined recombinant sperm immunogens, such as ESP, may offer opportunities for differential diagnosis of immune infertility.

  11. Gene expression in primary cultured astrocytes affected by aluminum: alteration of chaperons involved in protein folding

    PubMed Central

    Aremu, David A.; Ezomo, Ojeiru F.

    2010-01-01

    Objectives Aluminum is notorious as a neurotoxic metal. The aim of our study was to determine whether endoplasmic reticulum (ER) stress is involved in aluminum-induced apoptosis in astrocytes. Methods Mitochondrial RNA (mRNA) was analyzed by reverse transcription (RT)-PCR following pulse exposure of aluminum glycinate to primary cultured astrocytes. Tunicamycin was used as a positive control. Results Gene expression analysis revealed that Ire1β was up-regulated in astrocytes exposed to aluminum while Ire1α was up-regulated by tunicamycin. Exposure to aluminum glycinate, in contrast to tunicamycin, seemed to down-regulate mRNA expression of many genes, including the ER resident molecular chaperone BiP/Grp78 and Ca2+-binding chaperones (calnexin and calreticulin), as well as stanniocalcin 2 and OASIS. The down-regulation or non-activation of the molecular chaperons, whose expressions are known to be protective by increasing protein folding, may spell doom for the adaptive response. Exposure to aluminum did not have any significant effects on the expression of Bax and Bcl2 in astrocytes. Conclusions The results of this study demonstrate that aluminum may induce apoptosis in astrocytes via ER stress by impairing the protein-folding machinery. PMID:21432213

  12. ZAS: C2H2 zinc finger proteins involved in growth and development.

    PubMed

    Wu, Lai-Chu

    2002-01-01

    A ZAS gene encodes a large protein with two separate C2H2 zinc finger pairs that independently bind to specific DNA sequences, including the kappaB motif. Three paralogous mammalian genes, ZAS1, ZAS2, and ZAS3, and a related Drosophila gene, Schnurri, have been cloned and characterized. The ZAS genes encode transcriptional proteins that activate or repress the transcription of a variety of genes involved in growth, development, and metastasis. In addition, ZAS3 associates with a TNF receptor-associated factor to inhibit NF-kappaB- and JNK/ SAPK-mediated signaling of TNF-alpha. Genetic experiments show that ZAS3 deficiency leads to proliferation of cells and tumor formation in mice. The data suggest that ZAS3 is important in controlling cell growth, apoptosis, and inflammation. The potent vasoactive hormone endothelin and transcription factor AP2 gene families also each consist of three members. The ZAS, endothelin, and transcription factor AP2 genes form several linkage groups. Knowledge of the chromosomal locations of these genes provides valuable clues to the evolution of the vertebrate genome.

  13. Sendai virus trailer RNA binds TIAR, a cellular protein involved in virus-induced apoptosis.

    PubMed

    Iseni, Frédéric; Garcin, Dominique; Nishio, Machiko; Kedersha, Nancy; Anderson, Paul; Kolakofsky, Daniel

    2002-10-01

    Sendai virus (SeV) leader (le) and trailer (tr) RNAs are short transcripts generated during abortive antigenome and genome synthesis, respectively. Recom binant SeV (rSeV) that express tr-like RNAs from the leader region are non-cytopathic and, moreover, prevent wild-type SeV from inducing apoptosis in mixed infections. These rSeV thus appear to have gained a function. Here we report that tr RNA binds to a cellular protein with many links to apoptosis (TIAR) via the AU-rich sequence 5' UUUUAAAUUUU. Duplication of this AU-rich sequence alone within the le RNA confers TIAR binding on this le* RNA and a non-cytopathic phenotype to these rSeV in cell culture. Transgenic overexpression of TIAR during SeV infection promotes apoptosis and reverses the anti-apoptotic effects of le* RNA expression. More over, TIAR overexpression and SeV infection act synergistically to induce apoptosis. These short viral RNAs may act by sequestering TIAR, a multivalent RNA recognition motif (RRM) family RNA-binding protein involved in SeV-induced apoptosis. In this view, tr RNA is not simply a by-product of abortive genome synthesis, but is also an antigenome transcript that modulates the cellular antiviral response.

  14. The RNA binding protein TIAR is involved in the regulation of human iNOS expression.

    PubMed

    Fechir, M; Linker, K; Pautz, A; Hubrich, T; Kleinert, H

    2005-09-05

    Human inducible NO synthase (iNOS) expression is regulated by post-transcriptional mechanisms. The 3'-untranslated region (3'-UTR) of the human iNOS mRNA contains AU-rich elements (ARE), which are known to be important for the regulation of mRNA stability. The 3'-UTR of the human iNOS mRNA has been shown to regulate human iNOS mRNA expression post-transcriptionally. One RNA-binding protein known to interact with AREs and to regulate mRNA stability is the T cell intracellular antigen-1-related protein (TIAR). In RNA binding studies TIAR displayed high affinity binding to the human iNOS 3'-UTR sequence. In RNase protection experiments, the cytokine incubation needed for iNOS expression did not change TIAR expression in DLD-1 cells. However, overexpression of TIAR in human DLD-1 colon carcinoma cells resulted in enhanced cytokine-induced iNOS expression. In conclusion, TIAR seems to be involved in the post-transcriptional regulation of human iNOS expression.

  15. Adenylate cyclase-associated protein 1 overexpressed in pancreatic cancers is involved in cancer cell motility.

    PubMed

    Yamazaki, Ken; Takamura, Masaaki; Masugi, Yohei; Mori, Taisuke; Du, Wenlin; Hibi, Taizo; Hiraoka, Nobuyoshi; Ohta, Tsutomu; Ohki, Misao; Hirohashi, Setsuo; Sakamoto, Michiie

    2009-04-01

    Pancreatic cancer has the worst prognosis among cancers due to the difficulty of early diagnosis and its aggressive behavior. To characterize the aggressiveness of pancreatic cancers on gene expression, pancreatic cancer xenografts transplanted into severe combined immunodeficient mice served as a panel for gene-expression profiling. As a result of profiling, the adenylate cyclase-associated protein 1 (CAP1) gene was shown to be overexpressed in all of the xenografts. The expression of CAP1 protein in all 73 cases of pancreatic cancer was recognized by immunohistochemical analyses. The ratio of CAP1-positive tumor cells in clinical specimens was correlated with the presence of lymph node metastasis and neural invasion, and also with the poor prognosis of patients. Immunocytochemical analyses in pancreatic cancer cells demonstrated that CAP1 colocalized to the leading edge of lamellipodia with actin. Knockdown of CAP1 by RNA interference resulted in the reduction of lamellipodium formation, motility, and invasion of pancreatic cancer cells. This is the first report demonstrating the overexpression of CAP1 in pancreatic cancers and suggesting the involvement of CAP1 in the aggressive behavior of pancreatic cancer cells.

  16. Activation of Holliday junction recognizing protein involved in the chromosomal stability and immortality of cancer cells.

    PubMed

    Kato, Tatsuya; Sato, Nagato; Hayama, Satoshi; Yamabuki, Takumi; Ito, Tomoo; Miyamoto, Masaki; Kondo, Satoshi; Nakamura, Yusuke; Daigo, Yataro

    2007-09-15

    We identified a novel gene HJURP (Holliday junction-recognizing protein) whose activation seemed to play a pivotal role in the immortality of cancer cells. HJURP was considered a possible downstream target for ataxia telangiectasia mutated signaling, and its expression was increased by DNA double-strand breaks (DSB). HJURP was involved in the homologous recombination pathway in the DSB repair process through interaction with hMSH5 and NBS1, which is a part of the MRN protein complex. HJURP formed nuclear foci in cells at S phase and those subjected to DNA damage. In vitro assays implied that HJURP bound directly to the Holliday junction and rDNA arrays. Treatment of cancer cells with small interfering RNA (siRNA) against HJURP caused abnormal chromosomal fusions and led to genomic instability and senescence. In addition, HJURP overexpression was observed in a majority of lung cancers and was associated with poor prognosis as well. We suggest that HJURP is an indispensable factor for chromosomal stability in immortalized cancer cells and is a potential novel therapeutic target for the development of anticancer drugs.

  17. Molecular heterogeneity in major urinary proteins of Mus musculus subspecies: potential candidates involved in speciation

    PubMed Central

    Hurst, Jane L.; Beynon, Robert J.; Armstrong, Stuart D.; Davidson, Amanda J.; Roberts, Sarah A.; Gómez-Baena, Guadalupe; Smadja, Carole M.; Ganem, Guila

    2017-01-01

    When hybridisation carries a cost, natural selection is predicted to favour evolution of traits that allow assortative mating (reinforcement). Incipient speciation between the two European house mouse subspecies, Mus musculus domesticus and M.m.musculus, sharing a hybrid zone, provides an opportunity to understand evolution of assortative mating at a molecular level. Mouse urine odours allow subspecific mate discrimination, with assortative preferences evident in the hybrid zone but not in allopatry. Here we assess the potential of MUPs (major urinary proteins) as candidates for signal divergence by comparing MUP expression in urine samples from the Danish hybrid zone border (contact) and from allopatric populations. Mass spectrometric characterisation identified novel MUPs in both subspecies involving mostly new combinations of amino acid changes previously observed in M.m.domesticus. The subspecies expressed distinct MUP signatures, with most MUPs expressed by only one subspecies. Expression of at least eight MUPs showed significant subspecies divergence both in allopatry and contact zone. Another seven MUPs showed divergence in expression between the subspecies only in the contact zone, consistent with divergence by reinforcement. These proteins are candidates for the semiochemical barrier to hybridisation, providing an opportunity to characterise the nature and evolution of a putative species recognition signal. PMID:28337988

  18. Hepatitis C virus nonstructural protein 5B is involved in virus morphogenesis.

    PubMed

    Gouklani, Hamed; Bull, Rowena A; Beyer, Claudia; Coulibaly, Fasséli; Gowans, Eric J; Drummer, Heidi E; Netter, Hans J; White, Peter A; Haqshenas, Gholamreza

    2012-05-01

    The p7 protein of hepatitis C virus (HCV) is a viroporin that is dispensable for viral genome replication but plays a critical role in virus morphogenesis. In this study, we generated a JFH1-based intergenotypic chimeric genome that encoded a heterologous genotype 1b (GT1b) p7. The parental intergenotypic chimeric genome was nonviable in human hepatoma cells, and infectious chimeric virions were produced only when cells transfected with the chimeric genomes were passaged several times. Sequence analysis of the entire polyprotein-coding region of the recovered chimeric virus revealed one predominant amino acid substitution in nonstructural protein 2 (NS2), T23N, and one in NS5B, K151R. Forward genetic analysis demonstrated that each of these mutations per se restored the infectivity of the parental chimeric genome, suggesting that interactions between p7, NS2, and NS5B were required for virion assembly/maturation. p7 and NS5B colocalized in cellular compartments, and the NS5B mutation did not affect the colocalization pattern. The NS5B K151R mutation neither increased viral RNA replication in human hepatoma cells nor altered the polymerase activity of NS5B in an in vitro assay. In conclusion, this study suggests that HCV NS5B is involved in virus morphogenesis.

  19. Hepatitis C Virus Nonstructural Protein 5B Is Involved in Virus Morphogenesis

    PubMed Central

    Gouklani, Hamed; Bull, Rowena A.; Beyer, Claudia; Coulibaly, Fasséli; Gowans, Eric J.; Drummer, Heidi E.; Netter, Hans J.; White, Peter A.

    2012-01-01

    The p7 protein of hepatitis C virus (HCV) is a viroporin that is dispensable for viral genome replication but plays a critical role in virus morphogenesis. In this study, we generated a JFH1-based intergenotypic chimeric genome that encoded a heterologous genotype 1b (GT1b) p7. The parental intergenotypic chimeric genome was nonviable in human hepatoma cells, and infectious chimeric virions were produced only when cells transfected with the chimeric genomes were passaged several times. Sequence analysis of the entire polyprotein-coding region of the recovered chimeric virus revealed one predominant amino acid substitution in nonstructural protein 2 (NS2), T23N, and one in NS5B, K151R. Forward genetic analysis demonstrated that each of these mutations per se restored the infectivity of the parental chimeric genome, suggesting that interactions between p7, NS2, and NS5B were required for virion assembly/maturation. p7 and NS5B colocalized in cellular compartments, and the NS5B mutation did not affect the colocalization pattern. The NS5B K151R mutation neither increased viral RNA replication in human hepatoma cells nor altered the polymerase activity of NS5B in an in vitro assay. In conclusion, this study suggests that HCV NS5B is involved in virus morphogenesis. PMID:22345449

  20. A review of the electrophilic reaction chemistry involved in covalent protein binding relevant to toxicity.

    PubMed

    Enoch, S J; Ellison, C M; Schultz, T W; Cronin, M T D

    2011-10-01

    Several pieces of legislation have led to an increased interest in the use of in silico methods, specifically the formation of chemical categories for the assessment of toxicological endpoints. For a number of endpoints, this requires a detailed knowledge of the electrophilic reaction chemistry that governs the ability of an exogenous chemical to form a covalent adduct. Historically, this chemistry has been defined as compilations of structural alerts without documenting the associated electrophilic chemistry mechanisms. To address this, this article has reviewed the literature defining the structural alerts associated with covalent protein binding and detailed the associated electrophilic reaction chemistry. This information is useful to both toxicologists and regulators when using the chemical category approach to fill data gaps for endpoints involving covalent protein binding. The structural alerts and associated electrophilic reaction chemistry outlined in this review have been incorporated into the OECD (Q)SAR Toolbox, a freely available software tool designed to fill data gaps in a regulatory environment without the need for further animal testing.

  1. Estimation of Position Specific Energy as a Feature of Protein Residues from Sequence Alone for Structural Classification

    PubMed Central

    Iqbal, Sumaiya; Hoque, Md Tamjidul

    2016-01-01

    A set of features computed from the primary amino acid sequence of proteins, is crucial in the process of inducing a machine learning model that is capable of accurately predicting three-dimensional protein structures. Solutions for existing protein structure prediction problems are in need of features that can capture the complexity of molecular level interactions. With a view to this, we propose a novel approach to estimate position specific estimated energy (PSEE) of a residue using contact energy and predicted relative solvent accessibility (RSA). Furthermore, we demonstrate PSEE can be reasonably estimated based on sequence information alone. PSEE is useful in identifying the structured as well as unstructured or, intrinsically disordered region of a protein by computing favorable and unfavorable energy respectively, characterized by appropriate threshold. The most intriguing finding, verified empirically, is the indication that the PSEE feature can effectively classify disorder versus ordered residues and can segregate different secondary structure type residues by computing the constituent energies. PSEE values for each amino acid strongly correlate with the hydrophobicity value of the corresponding amino acid. Further, PSEE can be used to detect the existence of critical binding regions that essentially undergo disorder-to-order transitions to perform crucial biological functions. Towards an application of disorder prediction using the PSEE feature, we have rigorously tested and found that a support vector machine model informed by a set of features including PSEE consistently outperforms a model with an identical set of features with PSEE removed. In addition, the new disorder predictor, DisPredict2, shows competitive performance in predicting protein disorder when compared with six existing disordered protein predictors. PMID:27588752

  2. An Ehrlichia chaffeensis tandem repeat protein interacts with multiple host targets involved in cell signaling, transcriptional regulation, and vesicle trafficking.

    PubMed

    Wakeel, Abdul; Kuriakose, Jeeba A; McBride, Jere W

    2009-05-01

    Ehrlichia chaffeensis is an obligately intracellular bacterium that exhibits tropism for mononuclear phagocytes forming cytoplasmic membrane-bound microcolonies called morulae. To survive and replicate within phagocytes, E. chaffeensis exploits the host cell by modulating a number of host cell processes, but the ehrlichial effector proteins involved are unknown. In this study, we determined that p47, a secreted, differentially expressed, tandem repeat (TR) protein, interacts with multiple host proteins associated with cell signaling, transcriptional regulation, and vesicle trafficking. Yeast two-hybrid analysis revealed that p47 interacts with polycomb group ring finger 5 (PCGF5) protein, Src protein tyrosine kinase FYN (FYN), protein tyrosine phosphatase non-receptor type 2 (PTPN2), and adenylate cyclase-associated protein 1 (CAP1). p47 interaction with these proteins was further confirmed by coimmunoprecipitation assays and colocalization in HeLa cells transfected with p47-green fluorescent fusion protein (AcGFP1-p47). Moreover, confocal microscopy demonstrated p47-expressing dense-cored (DC) ehrlichiae colocalized with PCGF5, FYN, PTPN2, and CAP1. An amino-terminally truncated form of p47 containing TRs interacted only with PCGF5 and not with FYN, PTPN2, and CAP1, indicating differences in p47 domains that are involved in these interactions. These results demonstrate that p47 is involved in a complex network of interactions involving numerous host cell proteins. Furthermore, this study provides a new insight into the molecular and functional distinction of DC ehrlichiae, as well as the effector proteins involved in facilitating ehrlichial survival in mononuclear phagocytes.

  3. Quantitative characterization of protein–protein complexes involved in base excision DNA repair

    PubMed Central

    Moor, Nina A.; Vasil'eva, Inna A.; Anarbaev, Rashid O.; Antson, Alfred A.; Lavrik, Olga I.

    2015-01-01

    Base Excision Repair (BER) efficiently corrects the most common types of DNA damage in mammalian cells. Step-by-step coordination of BER is facilitated by multiple interactions between enzymes and accessory proteins involved. Here we characterize quantitatively a number of complexes formed by DNA polymerase β (Polβ), apurinic/apyrimidinic endonuclease 1 (APE1), poly(ADP-ribose) polymerase 1 (PARP1), X-ray repair cross-complementing protein 1 (XRCC1) and tyrosyl-DNA phosphodiesterase 1 (TDP1), using fluorescence- and light scattering-based techniques. Direct physical interactions between the APE1-Polβ, APE1-TDP1, APE1-PARP1 and Polβ-TDP1 pairs have been detected and characterized for the first time. The combined results provide strong evidence that the most stable complex is formed between XRCC1 and Polβ. Model DNA intermediates of BER are shown to induce significant rearrangement of the Polβ complexes with XRCC1 and PARP1, while having no detectable influence on the protein–protein binding affinities. The strength of APE1 interaction with Polβ, XRCC1 and PARP1 is revealed to be modulated by BER intermediates to different extents, depending on the type of DNA damage. The affinity of APE1 for Polβ is higher in the complex with abasic site-containing DNA than after the APE1-catalyzed incision. Our findings advance understanding of the molecular mechanisms underlying coordination and regulation of the BER process. PMID:26013813

  4. PARP12, an interferon-stimulated gene involved in the control of protein translation and inflammation.

    PubMed

    Welsby, Iain; Hutin, David; Gueydan, Cyril; Kruys, Veronique; Rongvaux, Anthony; Leo, Oberdan

    2014-09-19

    Transcriptome analyses have recently identified PARP12, a member of a large family of ADP-ribosyl transferases, as an interferon-induced gene (ISG), whose function remains incompletely characterized. We demonstrate herein that PARP12 is a genuine ISG, whose expressed protein displays at least two distinct subcellular locations and related functions. Upon ectopic expression or exposure to oxidative stress, PARP12 is recruited to stress-granules (SGs), known sites of mRNA translational arrest. Accordingly, PARP12 was found to block mRNA translation, possibly upon association to the translational machinery. Both the N-terminal domain (containing an RNA-binding domain characterized by the presence of five CCCH-type Zn-fingers) and integrity of the catalytic domain are required for this suppressive function. In contrast, stimulation with LPS leads to the localization of PARP12 to p62/SQSTM1 (an adaptor protein involved in innate signaling and autophagy) containing structures, unrelated to SGs. Deletion of the N-terminal domain promotes the association of the protein to p62/SQSTM1, suggesting that the RNA-binding domain is responsible for the subcellular localization of PARP12. Association to p62/SQSTM1 was found to correlate with increased NF-κB signaling, suggesting a role for PARP12 in inflammation. Collectively, these observations suggest that PARP12 can alternate between two distinct subcellular compartments associated to two distinct cellular functions. The present work therefore identifies PARP12 as an ISG with a potential role in cellular defenses against viral infections.

  5. Human DCXR - another 'moonlighting protein' involved in sugar metabolism, carbonyl detoxification, cell adhesion and male fertility?

    PubMed

    Ebert, Bettina; Kisiela, Michael; Maser, Edmund

    2015-02-01

    Dicarbonyl/L-xylulose reductase (DCXR; SDR20C1), a member of the short-chain dehydrogenase/reductase (SDR) superfamily catalyzes the reduction of α-dicarbonyl compounds and monosaccharides. Its role in the metabolism of L-xylulose has been known since 1970, when essential pentosuria was found to be associated with DCXR deficiency. Despite its early discovery, our knowledge about the role of human DCXR in normal physiology and pathophysiology is still incomplete. Sporadic studies have demonstrated aberrant expression in several cancers, but their physiological significance is unknown. In reproductive medicine, where DCXR is commonly referred to as 'sperm surface protein P34H', it serves as marker for epididymal sperm maturation and is essential for gamete interaction and successful fertilization. DCXR exhibits a multifunctional nature, both acting as a carbonyl reductase and also performing non-catalytic functions, possibly resulting from interactions with other proteins. Recent observations associate DCXR with a role in cell adhesion, pointing to a novel function involving tumour progression and possibly metastasis. This review summarizes the current knowledge about human DCXR and its orthologs from mouse and Caenorhabditis elegans (DHS-21) with an emphasis on its multifunctional characteristics. Due to its close structural relationship with DCXR, carbonyl reductase 2 (Cbr2), a tetrameric enzyme found in several non-primate species is also discussed. Similar to human DCXR, Cbr2 from golden hamster (P26h) and cow (P25b) is essential for sperm-zona pellucida interaction and fertilization. Because of the apparent similarity of these two proteins and the inconsistent use of alternative names previously, we provide an overview of the systematic classification of DCXR and Cbr2 and a phylogenetic analysis to illustrate their ancestry.

  6. Involvement of Arabidopsis RACK1 in Protein Translation and Its Regulation by Abscisic Acid

    SciTech Connect

    Guo, Jianjun; Wang, Shucai; Valerius, Oliver; Hall, Hardy; Zeng, Qingning; Li, Jian-Feng; Weston, David; Ellis, Brian; Chen, Jay

    2011-01-01

    Earlier studies have shown that RACK1 functions as a negative regulator of ABA responses in Arabidopsis, but the molecular mechanism of the action of RACK1 in these processes remains elusive. Global gene expression profiling revealed that approximately 40% of the genes affected by ABA treatment were affected in a similar manner by the rack1 mutation, supporting the view that RACK1 is an important regulator of ABA responses. On the other hand, co-expression analysis revealed that >80% of the genes co-expressed with RACK1 encode ribosome proteins, implying a close relationship between RACK1 s function and the ribosome complex. These results implied that the regulatory role for RACK1 in ABA responses may be partially due to its putative function in protein translation, which is one of the major cellular processes that mammalian and yeast RACK1 is involved in. Consistently, all three Arabidopsis RACK1 homologous genes, namely RACK1A, RACK1B and RACK1C, complemented the growth defects of the S. cerevisiae cpc2/rack1 mutant. In addition, RACK1 physically interacts with Arabidopsis Eukaryotic Initiation Factor 6 (eIF6), whose mammalian homologue is a key regulator of 80S ribosome assembly. Moreover, rack1 mutants displayed hypersensitivity to anisomycin, an inhibitor of protein translation, and displayed characteristics of impaired 80S functional ribosome assembly and 60S ribosomal subunit biogenesis in a ribosome profiling assay. Gene expression analysis revealed that ABA inhibits the expression of both RACK1 and eIF6. Taken together, these results suggest that RACK1 may be required for normal production of 60S and 80S ribosomes and that its action in these processes may be regulated by ABA.

  7. Arabidopsis microtubule destabilizing protein40 is involved in brassinosteroid regulation of hypocotyl elongation.

    PubMed

    Wang, Xianling; Zhang, Jin; Yuan, Ming; Ehrhardt, David W; Wang, Zhiyong; Mao, Tonglin

    2012-10-01

    The brassinosteroid (BR) phytohormones play crucial roles in regulating plant cell growth and morphogenesis, particularly in hypocotyl cell elongation. The microtubule cytoskeleton is also known to participate in the regulation of hypocotyl elongation. However, it is unclear if BR regulation of hypocotyl elongation involves the microtubule cytoskeleton. In this study, we demonstrate that BRs mediate hypocotyl cell elongation by influencing the orientation and stability of cortical microtubules. Further analysis identified the previously undiscovered Arabidopsis thaliana microtubule destabilizing protein40 (MDP40) as a positive regulator of hypocotyl cell elongation. Brassinazole-resistant1, a key transcription factor in the BR signaling pathway, directly targets and upregulates MDP40. Overexpression of MDP40 partially rescued the shorter hypocotyl phenotype in BR-deficient mutant de-etiolated-2 seedlings. Reorientation of the cortical microtubules in the cells of MDP40 RNA interference transgenic lines was less sensitive to BR. These findings demonstrate that MDP40 is a key regulator in BR regulation of cortical microtubule reorientation and mediates hypocotyl growth. This study reveals a mechanism involving BR regulation of microtubules through MDP40 to mediate hypocotyl cell elongation.

  8. Involvement of calcitonin gene-related peptide and receptor component protein in experimental autoimmune encephalomyelitis

    PubMed Central

    Sardi, Claudia; Zambusi, Laura; Finardi, Annamaria; Ruffini, Francesca; Tolun, Adviye A.; Dickerson, Ian M.; Righi, Marco; Zacchetti, Daniele; Grohovaz, Fabio; Provini, Luciano; Furlan, Roberto; Morara, Stefano

    2015-01-01

    Calcitonin Gene-Related Peptide (CGRP) inhibits microglia inflammatory activation in vitro. We here analyzed the involvement of CGRP and Receptor Component Protein (RCP) in experimental autoimmune encephalomyelitis (EAE). Alpha-CGRP deficiency increased EAE scores which followed the scale alpha-CGRP null > heterozygote > wild type. In wild type mice, CGRP delivery into the cerebrospinal fluid (CSF) 1) reduced chronic EAE (C-EAE) signs, 2) inhibited microglia activation (revealed by quantitative shape analysis), and 3) did not alter GFAP expression, cell density, lymphocyte infiltration, and peripheral lymphocyte production of IFN-gamma, TNF-alpha, IL-17, IL-2, and IL-4. RCP (probe for receptor involvement) was expressed in white matter microglia, astrocytes, oligodendrocytes, and vascular-endothelial cells: in EAE, also in infiltrating lymphocytes. In relapsing–remitting EAE (R-EAE) RCP increased during relapse, without correlation with lymphocyte density. RCP nuclear localization (stimulated by CGRP in vitro) was I) increased in microglia and decreased in astrocytes (R-EAE), and II) increased in microglia by CGRP CSF delivery (C-EAE). Calcitonin like receptor was rarely localized in nuclei of control and relapse mice. CGRP increased in motoneurons. In conclusion, CGRP can inhibit microglia activation in vivo in EAE. CGRP and its receptor may represent novel protective factors in EAE, apparently acting through the differential cell-specific intracellular translocationof RCP. PMID:24746422

  9. The MagA Protein of Magnetospirilla Is Not Involved in Bacterial Magnetite Biomineralization

    PubMed Central

    Uebe, René; Henn, Verena

    2012-01-01

    Magnetotactic bacteria have the ability to orient along geomagnetic field lines based on the formation of magnetosomes, which are intracellular nanometer-sized, membrane-enclosed magnetic iron minerals. The formation of these unique bacterial organelles involves several processes, such as cytoplasmic membrane invagination and magnetosome vesicle formation, the accumulation of iron in the vesicles, and the crystallization of magnetite. Previous studies suggested that the magA gene encodes a magnetosome-directed ferrous iron transporter with a supposedly essential function for magnetosome formation in Magnetospirillum magneticum AMB-1 that may cause magnetite biomineralization if expressed in mammalian cells. However, more recent studies failed to detect the MagA protein among polypeptides associated with the magnetosome membrane and did not identify magA within the magnetosome island, a conserved genomic region that is essential for magnetosome formation in magnetotactic bacteria. This raised increasing doubts about the presumptive role of magA in bacterial magnetosome formation, which prompted us to reassess MagA function by targeted deletion in Magnetospirillum magneticum AMB-1 and Magnetospirillum gryphiswaldense MSR-1. Contrary to previous reports, magA mutants of both strains still were able to form wild-type-like magnetosomes and had no obvious growth defects. This unambiguously shows that magA is not involved in magnetosome formation in magnetotactic bacteria. PMID:22194451

  10. BcSUN1, a B. cinerea SUN-Family Protein, Is Involved in Virulence

    PubMed Central

    Pérez-Hernández, Alicia; González, Mario; González, Celedonio; van Kan, Jan A. L.; Brito, Nélida

    2017-01-01

    BcSUN1 is a glycoprotein secreted by Botrytis cinerea, an important plant pathogen that causes severe losses in agriculture worldwide. In this work, the role of BcSUN1 in different aspects of the B. cinerea biology was studied by phenotypic analysis of Bcsun1 knockout strains. We identified BcSUN1 as the only member of the Group-I SUN family of proteins encoded in the B. cinerea genome, which is expressed both in axenic culture and during infection. BcSUN1 is also weakly attached to the cellular surface and is involved in maintaining the structure of the cell wall and/or the extracellular matrix. Disruption of the Bcsun1 gene produces different cell surface alterations affecting the production of reproductive structures and adhesion to plant surface, therefore reducing B. cinerea virulence. BcSUN1 is the first member of the SUN family reported to be involved in the pathogenesis of a filamentous fungus. PMID:28163701

  11. Tracking structural features leading to resistance of activated protein C to alpha 1-antitrypsin.

    PubMed

    Shen, L; Dahlbäck, B; Villoutreix, B O

    2000-03-21

    Activated protein C (APC) is a multi-modular anticoagulant serine protease, which degrades factor V/Va and factor VIIIa. Human APC (hAPC) is inhibited by human alpha 1-antitrypsin (AAT), while the bovine enzyme (bAPC) is fully resistant to this serpin. Structural features in the catalytic domains between the two species cause this difference, but detailed knowledge about the causal molecular difference is missing. To gain insight into the APC-AAT interaction and to create a human protein C resistant to AAT inhibition, we have used molecular modeling and site-directed mutagenesis. First, a structural model for bAPC based on the Gla-domainless X-ray structure of hAPC was built. Screening the molecular surface of the human and bovine APC enzymes suggested that a hAPC molecule resistant to AAT inhibition could be constructed by substituting only a few amino acids. We thus produced recombinant hAPC molecules with a single mutation (S173E, the numbering follows the chymotrypsinogen nomenclature), two mutations (E60aS/S61R) or a combination of all these substitutions (E60aS/S61R/S173E). Amidolytic and anticoagulant activities of the three mutant APC molecules were similar to those of wild-type hAPC. Inhibition of wild-type hAPC by AAT was characterized by a second-order rate constant (k2) of 2.71 M-1 s-1. The amino acid substitution at position 173 (S173E mutant) led to partial resistance to AAT (k2 = 0.84 M-1 s-1). The E60aS/S61R mutant displayed mild resistance to AAT inhibition (k2 = 1.70 M-1 s-1), whereas the E60aS/S61R/S173E mutant was inefficiently inactivated by AAT (k2 = 0.40 M-1 s-1). Inhibition of recombinant APC molecules by the serpin protein C inhibitor (PCI) in the presence and absence of heparin was also investigated.

  12. Oxidation by Neutrophils-Derived HOCl Increases Immunogenicity of Proteins by Converting Them into Ligands of Several Endocytic Receptors Involved in Antigen Uptake by Dendritic Cells and Macrophages

    PubMed Central

    Biedroń, Rafał; Konopiński, Maciej K.; Marcinkiewicz, Janusz; Józefowski, Szczepan

    2015-01-01

    The initiation of adaptive immune responses to protein antigens has to be preceded by their uptake by antigen presenting cells and intracellular proteolytic processing. Paradoxically, endocytic receptors involved in antigen uptake do not bind the majority of proteins, which may be the main reason why purified proteins stimulate at most weak immune responses. A shared feature of different types of adjuvants, capable of boosting immunogenicity of protein vaccines, is their ability to induce acute inflammation, characterized by early influx of activated neutrophils. Neutrophils are also rapidly recruited to sites of tissue injury or infection. These cells are the source of potent oxidants, including hypochlorous acid (HOCl), causing oxidation of proteins present in inflammatory foci. We demonstrate that oxidation of proteins by endogenous, neutrophils-derived HOCl increases their immunogenicity. Upon oxidation, different, randomly chosen simple proteins (yeast alcohol dehydrogenase, human and bovine serum albumin) and glycoproteins (human apo-transferrin, ovalbumin) gain the ability to bind with high affinity to several endocytic receptors on antigen presenting cells, which seems to be the major mechanism of their increased immunogenicity. The mannose receptor (CD206), scavenger receptors A (CD204) and CD36 were responsible for the uptake and presentation of HOCl-modified proteins by murine dendritic cells and macrophages. Other scavenger receptors, SREC-I and LOX-1, as well as RAGE were also able to bind HOCl-modified proteins, but they did not contribute significantly to these ligands uptake by dendritic cells because they were either not expressed or exhibited preference for more heavily oxidised proteins. Our results indicate that oxidation by neutrophils-derived HOCl may be a physiological mechanism of conferring immunogenicity on proteins which in their native forms do not bind to endocytic receptors. This mechanism might enable the immune system to detect

  13. Involvement of hypothalamic AMP-activated protein kinase in leptin-induced sympathetic nerve activation.

    PubMed

    Tanida, Mamoru; Yamamoto, Naoki; Shibamoto, Toshishige; Rahmouni, Kamal

    2013-01-01

    In mammals, leptin released from the white adipose tissue acts on the central nervous system to control feeding behavior, cardiovascular function, and energy metabolism. Central leptin activates sympathetic nerves that innervate the kidney, adipose tissue, and some abdominal organs in rats. AMP-activated protein kinase (AMPK) is essential in the intracellular signaling pathway involving the activation of leptin receptors (ObRb). We investigated the potential of AMPKα2 in the sympathetic effects of leptin using in vivo siRNA injection to knockdown AMPKα2 in rats, to produce reduced hypothalamic AMPKα2 expression. Leptin effects on body weight, food intake, and blood FFA levels were eliminated in AMPKα2 siRNA-treated rats. Leptin-evoked enhancements of the sympathetic nerve outflows to the kidney, brown and white adipose tissues were attenuated in AMPKα2 siRNA-treated rats. To check whether AMPKα2 was specific to sympathetic changes induced by leptin, we examined the effects of injecting MT-II, a melanocortin-3 and -4 receptor agonist, on the sympathetic nerve outflows to the kidney and adipose tissue. MT-II-induced sympatho-excitation in the kidney was unchanged in AMPKα2 siRNA-treated rats. However, responses of neural activities involving adipose tissue to MT-II were attenuated in AMPKα2 siRNA-treated rats. These results suggest that hypothalamic AMPKα2 is involved not only in appetite and body weight regulation but also in the regulation of sympathetic nerve discharges to the kidney and adipose tissue. Thus, AMPK might function not only as an energy sensor, but as a key molecule in the cardiovascular, thermogenic, and lipolytic effects of leptin through the sympathetic nervous system.

  14. Toxicant-induced acceleration of epididymal sperm transit: androgen-dependent proteins may be involved.

    PubMed

    Klinefelter, G R; Suarez, J D

    1997-01-01

    protein profile in homogenates of the caput/corpus epididymidis revealed treatment-related diminutions in two proteins CC9 (M(r) = 42 kDa, pI = 4.2) and CC34 (M(r) = 35 kDa, pI = 5.5), and the level of each of these proteins in the caput/corpus was significantly correlated with the decrease in caput/corpus sperm number. Thus, both CEMS and HFLUT accelerate sperm transit through the proximal segment of the epididymis; and, while this effect is not dependent on the testis, it may involve a lesion in androgen-dependent epididymal function.

  15. Application of Ict and Rubrics to the Assessment Process where Professional Judgement Is Involved: The Features of an Emarking Tool

    ERIC Educational Resources Information Center

    Campbell, Alistair

    2005-01-01

    This paper describes a new ICT assessment tool that reduces multihandling of marks, comments and scores specifically where professional judgement is involved. Whereas previous ICT applications in most fields of education have focused on student learning, this tool focuses on the lecturer task of the assessment process. Unlike many ICT based…

  16. Amyloid precursor protein in Drosophila glia regulates sleep and genes involved in glutamate recycling.

    PubMed

    Farca Luna, Abud Jose; Perier, Magali; Seugnet, Laurent

    2017-03-17

    The Amyloid Precursor Protein (App) plays a crucial role in Alzheimer disease (AD) via the production and deposition of toxic β-amyloid peptides. App is heavily expressed in neurons where the vast majority of studies investigating its function have been carried out, while almost nothing is known about its function in glia, where it is also expressed, and can potentially participate in the regulation of neuronal physiology. In this report, we investigated whether Appl, the Drosophila homolog of App, could influence sleep-wake regulation when its function is manipulated in glial cells. Appl inhibition in astrocyte-like and cortex glia resulted in higher sleep amounts and longer sleep bout duration during the night, while overexpression had the opposite effect. These sleep phenotypes were not the result of developmental defects, and were correlated with changes in expression in Glutamine Synthetase (GS) in astrocyte-like glia, and in changes in the gap-junction component innexin2 in cortex glia. Downregulating both GS and innexin2, but not either one individually, resulted in higher sleep amounts, similarly to Appl inhibition. Consistent with these results the expression of GS and innexin2 are increased following sleep deprivation indicating that these two genes are dynamically linked to vigilance states. Interestingly, the reduction of GS expression and the sleep phenotype observed upon Appl inhibition could be rescued by increasing the expression of the glutamate transporter dEaat1. In contrast, reducing dEaat1 expression severely disrupted sleep. These results associate glutamate recycling, sleep and a glial function for the App family proteins.StatementThe Amyloid Precursor Protein (App) has been intensively studied for its implication in Alzheimer Disease (AD). The attributed functions of App are linked to the physiology and cellular biology of neurons where the protein is predominantly expressed. Consequences on glia in AD are generally thought to be secondary

  17. Proteins involved on TGF-β pathway are up-regulated during the acute phase of experimental Chagas disease.

    PubMed

    Ferreira, Roberto Rodrigues; de Souza, Elen Mello; de Oliveira, Fabiane Loiola; Ferrão, Patrícia Mello; Gomes, Leonardo Henrique Ferreira; Mendonça-Lima, Leila; Meuser-Batista, Marcelo; Bailly, Sabine; Feige, Jean Jacques; de Araujo-Jorge, Tania Cremonini; Waghabi, Mariana Caldas

    2016-05-01

    Studies developed by our group in the last years have shown the involvement of TGF-β in acute and chronic Chagas heart disease, with elevated plasma levels and activated TGF-β cell signaling pathway as remarkable features of patients in the advanced stages of this disease, when high levels of cardiac fibrosis is present. Imbalance in synthesis and degradation of extracellular matrix components is the basis of pathological fibrosis and TGF-β is considered as one of the key regulators of this process. In the present study, we investigated the activity of the TGF-β signaling pathway, including receptors and signaling proteins activation in the heart of animals experimentally infected with Trypanosoma cruzi during the period that mimics the acute phase of Chagas disease. We observed that T. cruzi-infected animals presented increased expression of TGF-β receptors. Overexpression of receptors was followed by an increased phosphorylation of Smad2/3, p38 and ERK. Furthermore, we correlated these activities with cellular factors involved in the fibrotic process induced by TGF-β. We observed that the expression of collagen I, fibronectin and CTGF were increased in the heart of infected animals on day 15 post-infection. Correlated with the increased TGF-β activity in the heart, we found that serum levels of total TGF-β were significantly higher during acute infection. Taken together, our data suggest that the commitment of the heart associates with increased activity of TGF-β pathway and expression of its main components. Our results, confirm the importance of this cytokine in the development and maintenance of cardiac damage caused by T. cruzi infection.

  18. A novel Drosophila Girdin-like protein is involved in Akt pathway control of cell size

    SciTech Connect

    Puseenam, Aekkachai; Yoshioka, Yasuhide; Nagai, Rika; Hashimoto, Reina; Suyari, Osamu; Itoh, Masanobu; Enomoto, Atsushi; Takahashi, Masahide; Yamaguchi, Masamitsu

    2009-11-15

    The Akt signaling pathway is well known to regulate cell proliferation and growth. Girdin, a novel substrate of Akt, plays a crucial role in organization of the actin cytoskeleton and cell motility under the control of Akt. We here identified a novel Girdin-like protein in Drosophila (dGirdin), which has two isoforms, dGirdin PA and dGirdin PB. dGirdin shows high homology with human Girdin in the N-terminal and coiled-coil domains, while diverging at the C-terminal domain. On establishment of transgenic fly lines, featuring knockdown or overexpression of dGirdin in vivo, overexpression in the wing disc cells induced ectopic apoptosis, implying a role in directing apoptosis. Knockdown of dGirdin in the Drosophila wing imaginal disc cells resulted in reduction of cell size. Furthermore, this was enhanced by half reduction of the Akt gene dose, suggesting that Akt positively regulates dGirdin. In the wing disc, cells in which dGirdin was knocked down exhibited disruption of actin filaments. From these in vivo analyses, we conclude that dGirdin is required for actin organization and regulation of appropriate cell size under control of the Akt signaling pathway.

  19. Extensive and mutilating craniofacial trauma involving defleshing and decapitation: unusual features of fatal dog attacks in the young.

    PubMed

    Tsokos, Michael; Byard, Roger W; Püschel, Klaus

    2007-06-01

    Four cases of fatal dog attacks are reported in 3 children aged 6, 10, and 11 years and in an infant aged 3 weeks. The cases were all characterized by extensive and mutilative stripping of soft tissues from the face and scalp, progressing to decapitation in the infant. The attacks were highly focused, involving 2 dogs in all but 1 case, with the area of trauma localized to the craniofacial region. The injuries resembled those found after postmortem animal depredation. The involvement of more than 1 dog may account for the severity of the injuries due to "pack" behavior. Deaths were due to exsanguination, air embolism, and decapitation. Necropsy examination of the attacking dogs revealed tissues from the victims in 2 of the animals' stomachs. These cases demonstrate the vulnerability of infants and young children to fatal dog attacks, with an unusual concentration of severe injuries to the head regions. Necropsy of the canine assailant, with collaboration between pathologists and veterinarians, is an important part of such investigations as it may provide information helping to establish the identity and ownership of the animal, along with trace evidence confirming that the dog was involved in the attack.

  20. A Systematic Review of the Effects of Plant Compared with Animal Protein Sources on Features of Metabolic Syndrome.

    PubMed

    Chalvon-Demersay, Tristan; Azzout-Marniche, Dalila; Arfsten, Judith; Egli, Léonie; Gaudichon, Claire; Karagounis, Leonidas G; Tomé, Daniel

    2017-03-01

    Dietary protein may play an important role in the prevention of metabolic dysfunctions. However, the way in which the protein source affects these dysfunctions has not been clearly established. The aim of the current systematic review was to compare the impact of plant- and animal-sourced dietary proteins on several features of metabolic syndrome in humans. The PubMed database was searched for both chronic and acute interventional studies, as well as observational studies, in healthy humans or those with metabolic dysfunctions, in which the impact of animal and plant protein intake was compared while using the following variables: cholesterolemia and triglyceridemia, blood pressure, glucose homeostasis, and body composition. Based on data extraction, we observed that soy protein consumption (with isoflavones), but not soy protein alone (without isoflavones) or other plant proteins (pea and lupine proteins, wheat gluten), leads to a 3% greater decrease in both total and LDL cholesterol compared with animal-sourced protein ingestion, especially in individuals with high fasting cholesterol concentrations. This observation was made when animal proteins were provided as a whole diet rather than given supplementally. Some observational studies reported an inverse association between plant protein intake and systolic and diastolic blood pressure, but this was not confirmed by intervention studies. Moreover, plant protein (wheat gluten, soy protein) intake as part of a mixed meal resulted in a lower postprandial insulin response than did whey. This systematic review provides some evidence that the intake of soy protein associated with isoflavones may prevent the onset of risk factors associated with cardiovascular disease, i.e., hypercholesterolemia and hypertension, in humans. However, we were not able to draw any further conclusions from the present work on the positive effects of plant proteins relating to glucose homeostasis and body composition.

  1. Characterization of carbohydrate structural features recognized by anti-arabinogalactan-protein monoclonal antibodies.

    PubMed

    Yates, E A; Valdor, J F; Haslam, S M; Morris, H R; Dell, A; Mackie, W; Knox, J P

    1996-03-01

    Arabinogalactan-proteins (AGPs) are a diverse class of plant cell surface proteoglycans implicated in a range of fundamental processes associated with plant cell development. Anti-AGP monoclonal antibodies have been used extensively for the investigation of the developmental regulation of AGPs although virtually nothing is known about the structure of the carbohydrate epitopes recognised by these antibodies. In this report, a series of methyl glycosides of monosaccharides and a range of oligosaccharides that are elements of the carbohydrate component of AGPs have been investigated for recognition by previously derived anti-AGP monoclonal antibodies. No clear evidence was obtained for the involvement of terminal arabinofuranosides, nor of the galactan backbone, in the recognition of the glycan structure of AGPs by any of the antibodies used in this study. Interestingly, the most effective inhibitor of the binding of the monoclonal antibodies MAC207, JIM4 and JIM13 to exudate gum antigens was an acidic trisaccharide, isolated from a partial acid hydrolysate of gum karaya which has the structure: GlcA beta(1-->3) GalA alpha(1-->2)Rha, determined by a combination of FAB-MS, GC-MS and NMR spectroscopy.

  2. The Eucalyptus Tonoplast Intrinsic Protein (TIP) Gene Subfamily: Genomic Organization, Structural Features, and Expression Profiles

    PubMed Central

    Rodrigues, Marcela I.; Takeda, Agnes A. S.; Bravo, Juliana P.; Maia, Ivan G.

    2016-01-01

    Plant aquaporins are water channels implicated in various physiological processes, including growth, development and adaptation to stress. In this study, the Tonoplast Intrinsic Protein (TIP) gene subfamily of Eucalyptus, an economically important woody species, was investigated and characterized. A genome-wide survey of the Eucalyptus grandis genome revealed the presence of eleven putative TIP genes (referred as EgTIP), which were individually assigned by phylogeny to each of the classical TIP1–5 groups. Homology modeling confirmed the presence of the two highly conserved NPA (Asn-Pro-Ala) motifs in the identified EgTIPs. Residue variations in the corresponding selectivity filters, that might reflect differences in EgTIP substrate specificity, were observed. All EgTIP genes, except EgTIP5.1, were transcribed and the majority of them showed organ/tissue-enriched expression. Inspection of the EgTIP promoters revealed the presence of common cis-regulatory elements implicated in abiotic stress and hormone responses pointing to an involvement of the identified genes in abiotic stress responses. In line with these observations, additional gene expression profiling demonstrated increased expression under polyethylene glycol-imposed osmotic stress. Overall, the results obtained suggest that these novel EgTIPs might be functionally implicated in eucalyptus adaptation to stress. PMID:27965702

  3. Human Suv3 protein reveals unique features among SF2 helicases

    PubMed Central

    Jedrzejczak, Robert; Wang, Jiawei; Dauter, Miroslawa; Szczesny, Roman J.; Stepien, Piotr P.; Dauter, Zbigniew

    2011-01-01

    Suv3 is a helicase that is involved in efficient turnover and surveillance of RNA in eukaryotes. In vitro studies show that human Suv3 (hSuv3) in complex with human polynucleotide phosphorylase has RNA degradosome activity. The enzyme is mainly localized in mitochondria, but small fractions are found in cell nuclei. Here, two X-ray crystallographic structures of human Suv3 in complex with AMPPNP, a nonhydrolysable analog of ATP, and with a short five-nucleotide strand of RNA are presented at resolutions of 2.08 and 2.9 Å, respectively. The structure of the enzyme is very similar in the two complexes and consists of four domains. Two RecA-like domains form the tandem typical of all helicases from the SF2 superfamily which together with the C-terminal all-helical domain makes a ring structure through which the nucleotide strand threads. The mostly helical N-terminal domain is positioned externally with respect to the core of the enzyme. Most of the typical helicase motifs are present in hSuv3, but the protein shows certain unique characteristics, suggesting that Suv3 enzymes may constitute a separate subfamily of helicases. PMID:22101826

  4. Human Suv3 protein reveals unique features among SF2 helicases

    SciTech Connect

    Jedrzejczak, Robert; Wang, Jiawei; Dauter, Miroslawa; Szczesny, Roman J.; Stepien, Piotr P.; Dauter, Zbigniew

    2011-11-01

    Crystal structures of the human mitochondrial helicase hSuv3 in complex with AMPPNP and with a short strand of RNA are presented. Suv3 is a helicase that is involved in efficient turnover and surveillance of RNA in eukaryotes. In vitro studies show that human Suv3 (hSuv3) in complex with human polynucleotide phosphorylase has RNA degradosome activity. The enzyme is mainly localized in mitochondria, but small fractions are found in cell nuclei. Here, two X-ray crystallographic structures of human Suv3 in complex with AMPPNP, a nonhydrolysable analog of ATP, and with a short five-nucleotide strand of RNA are presented at resolutions of 2.08 and 2.9 Å, respectively. The structure of the enzyme is very similar in the two complexes and consists of four domains. Two RecA-like domains form the tandem typical of all helicases from the SF2 superfamily which together with the C-terminal all-helical domain makes a ring structure through which the nucleotide strand threads. The mostly helical N-terminal domain is positioned externally with respect to the core of the enzyme. Most of the typical helicase motifs are present in hSuv3, but the protein shows certain unique characteristics, suggesting that Suv3 enzymes may constitute a separate subfamily of helicases.

  5. Human Suv3 protein reveals unique features among SF2 helicases

    SciTech Connect

    Jedrzejczak, Robert; Wang, Jiawei; Dauter, Miroslawa; Szczesny, Roman J.; Stepien, Piotr P.; Dauter, Zbigniew

    2012-03-16

    Suv3 is a helicase that is involved in efficient turnover and surveillance of RNA in eukaryotes. In vitro studies show that human Suv3 (hSuv3) in complex with human polynucleotide phosphorylase has RNA degradosome activity. The enzyme is mainly localized in mitochondria, but small fractions are found in cell nuclei. Here, two X-ray crystallographic structures of human Suv3 in complex with AMPPNP, a nonhydrolysable analog of ATP, and with a short five-nucleotide strand of RNA are presented at resolutions of 2.08 and 2.9 {angstrom}, respectively. The structure of the enzyme is very similar in the two complexes and consists of four domains. Two RecA-like domains form the tandem typical of all helicases from the SF2 superfamily which together with the C-terminal all-helical domain makes a ring structure through which the nucleotide strand threads. The mostly helical N-terminal domain is positioned externally with respect to the core of the enzyme. Most of the typical helicase motifs are present in hSuv3, but the protein shows certain unique characteristics, suggesting that Suv3 enzymes may constitute a separate subfamily of helicases.

  6. The effects of (-)-epicatechin on endothelial cells involve the G protein-coupled estrogen receptor (GPER).

    PubMed

    Moreno-Ulloa, Aldo; Mendez-Luna, David; Beltran-Partida, Ernesto; Castillo, Carmen; Guevara, Gustavo; Ramirez-Sanchez, Israel; Correa-Basurto, José; Ceballos, Guillermo; Villarreal, Francisco

    2015-10-01

    We have provided evidence that the stimulatory effects of (-)-epicatechin ((-)-EPI) on endothelial cell nitric oxide (NO) production may involve the participation of a cell-surface receptor. Thus far, such entity(ies) has not been fully elucidated. The G protein-coupled estrogen receptor (GPER) is a cell-surface receptor that has been linked to protective effects on the cardiovascular system and activation of intracellular signaling pathways (including NO production) similar to those reported with (-)-EPI. In bovine coronary artery endothelial cells (BCAEC) by the use of confocal imaging, we evidence the presence of GPER at the cell-surface and on F-actin filaments. Using in silico studies we document the favorable binding mode between (-)-EPI and GPER. Such binding is comparable to that of the GPER agonist, G1. By the use of selective blockers, we demonstrate that the activation of ERK 1/2 and CaMKII by (-)-EPI is dependent on the GPER/c-SRC/EGFR axis mimicking those effects noted with G1. We also evidence by the use of siRNA the role that GPER has on mediating ERK1/2 activation by (-)-EPI. GPER appears to be coupled to a non Gαi/o or Gαs, protein subtype. To extrapolate our findings to an ex vivo model, we employed phenylephrine pre-contracted aortic rings evidencing that (-)-EPI can mediate vasodilation through GPER activation. In conclusion, we provide evidence that suggests the GPER as a potential mediator of (-)-EPI effects and highlights the important role that GPER may have on cardiovascular system protection.

  7. Development of Novel In Vivo Chemical Probes to Address CNS Protein Kinase Involvement in Synaptic Dysfunction

    PubMed Central

    Watterson, D. Martin; Grum-Tokars, Valerie L.; Roy, Saktimayee M.; Schavocky, James P.; Bradaric, Brinda Desai; Bachstetter, Adam D.; Xing, Bin; Dimayuga, Edgardo; Saeed, Faisal; Zhang, Hong; Staniszewski, Agnieszka; Pelletier, Jeffrey C.; Minasov, George; Anderson, Wayne F.; Arancio, Ottavio; Van Eldik, Linda J.

    2013-01-01

    Serine-threonine protein kinases are critical to CNS function, yet there is a dearth of highly selective, CNS-active kinase inhibitors for in vivo investigations. Further, prevailing assumptions raise concerns about whether single kinase inhibitors can show in vivo efficacy for CNS pathologies, and debates over viable approaches to the development of safe and efficacious kinase inhibitors are unsettled. It is critical, therefore, that these scientific challenges be addressed in order to test hypotheses about protein kinases in neuropathology progression and the potential for in vivo modulation of their catalytic activity. Identification of molecular targets whose in vivo modulation can attenuate synaptic dysfunction would provide a foundation for future disease-modifying therapeutic development as well as insight into cellular mechanisms. Clinical and preclinical studies suggest a critical link between synaptic dysfunction in neurodegenerative disorders and the activation of p38αMAPK mediated signaling cascades. Activation in both neurons and glia also offers the unusual potential to generate enhanced responses through targeting a single kinase in two distinct cell types involved in pathology progression. However, target validation has been limited by lack of highly selective inhibitors amenable to in vivo use in the CNS. Therefore, we employed high-resolution co-crystallography and pharmacoinformatics to design and develop a novel synthetic, active site targeted, CNS-active, p38αMAPK inhibitor (MW108). Selectivity was demonstrated by large-scale kinome screens, functional GPCR agonist and antagonist analyses of off-target potential, and evaluation of cellular target engagement. In vitro and in vivo assays demonstrated that MW108 ameliorates beta-amyloid induced synaptic and cognitive dysfunction. A serendipitous discovery during co-crystallographic analyses revised prevailing models about active site targeting of inhibitors, providing insights that will

  8. Signatures of nitrogen limitation in the elemental composition of the proteins involved in the metabolic apparatus.

    PubMed

    Acquisti, Claudia; Kumar, Sudhir; Elser, James J

    2009-07-22

    Nitrogen (N) is a fundamental component of nucleotides and amino acids and is often a limiting nutrient in natural ecosystems. Thus, study of the N content of biomolecules may establish important connections between ecology and genomics. However, while significant differences in the elemental composition of whole organisms are well documented, how the flux of nutrients in the cell has shaped the evolution of different cellular processes remains poorly understood. By examining the elemental composition of major functional classes of proteins in four multicellular eukaryotic model organisms, we find that the catabolic machinery shows substantially lower N content than the anabolic machinery and the rest of the proteome. This pattern suggests that ecological selection for N conservation specifically targets cellular components that are highly expressed in response to nutrient limitation. We propose that the RNA component of the anabolic machineries is the mechanistic force driving the elemental imbalance we found, and that RNA functions as an intracellular nutrient reservoir that is degraded and recycled during starvation periods. A comparison of the elemental composition of the anabolic and catabolic machineries in species that have experienced different levels of N limitation in their evolutionary history (animals versus plants) suggests that selection for N conservation has preferentially targeted the catabolic machineries of plants, resulting in a lower N content of the proteins involved in their catabolic processes. These findings link the composition of major cellular components to the environmental factors that trigger the activation of those components, suggesting that resource availability has constrained the atomic composition and the molecular architecture of the biotic processes that enable cells to respond to reduced nutrient availability.

  9. A highly accurate protein structural class prediction approach using auto cross covariance transformation and recursive feature elimination.

    PubMed

    Li, Xiaowei; Liu, Taigang; Tao, Peiying; Wang, Chunhua; Chen, Lanming

    2015-12-01

    Structural class characterizes the overall folding type of a protein or its domain. Many methods have been proposed to improve the prediction accuracy of protein structural class in recent years, but it is still a challenge for the low-similarity sequences. In this study, we introduce a feature extraction technique based on auto cross covariance (ACC) transformation of position-specific score matrix (PSSM) to represent a protein sequence. Then support vector machine-recursive feature elimination (SVM-RFE) is adopted to select top K features according to their importance and these features are input to a support vector machine (SVM) to conduct the prediction. Performance evaluation of the proposed method is performed using the jackknife test on three low-similarity datasets, i.e., D640, 1189 and 25PDB. By means of this method, the overall accuracies of 97.2%, 96.2%, and 93.3% are achieved on these three datasets, which are higher than those of most existing methods. This suggests that the proposed method could serve as a very cost-effective tool for predicting protein structural class especially for low-similarity datasets.

  10. Calmodulin and Ca2+/calmodulin-binding proteins are involved in Tetrahymena thermophila phagocytosis.

    PubMed

    Gonda, K; Komatsu, M; Numata, O

    2000-08-01

    The ciliated protist, Tetrahymena thermophila, possesses one oral apparatus for phagocytosis, one of the most important cell functions, in the anterior cell cortex. The apparatus comprises four membrane structures which consist of ciliated and unciliated basal bodies, a cytostome where food is collected by oral ciliary motility, and a cytopharynx where food vacuoles are formed. The food vacuole is thought to be transported into the cytoplasm by a deep fiber which connects with the oral apparatus. Although a large number of studies have been done on the structure of the oral apparatus, the molecular mechanisms of phagocytosis in Tetrahymena thermophila are not well understood. In this study, using indirect immunofluorescence, we demonstrated that the deep fiber consisted of actin, CaM, and Ca2+/CaM-binding proteins, p85 and EF-1alpha, which are closely involved in cytokinesis. Moreover, we showed that CaM, p85, and EF-1alpha are colocalized in the cytostome and the cytopharynx of the oral apparatus. Next, we examined whether Ca2+/CaM signal regulates Tetrahymena thermophila phagocytosis, using Ca2+/CaM inhibitors chlorpromazine, trifluoperazine, N-(6-aminohexyl)-1-naphthalenesulfonamide, and N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide HCI. In Tetrahymena, it is known that Ca2+/CaM signal is closely involved in ciliary motility and cytokinesis. The results showed that one of the inhibitors, N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide HCl, inhibited the food vacuole formation rather than the ciliary motility, while the other three inhibitors effectively prevented the ciliary motility. Considering the colocalization of CaM, p85, and EF-1alpha to the cytopharynx, these results suggest that the Ca2+/CaM signal plays a pivotal role in Tetrahymena thermophila food vacuole formation.

  11. Translation of CGA codon repeats in yeast involves quality control components and ribosomal protein L1.

    PubMed

    Letzring, Daniel P; Wolf, Andrew S; Brule, Christina E; Grayhack, Elizabeth J

    2013-09-01

    Translation of CGA codon repeats in the yeast Saccharomyces cerevisiae is inefficient, resulting in dose-dependent reduction in expression and in production of an mRNA cleavage product, indicative of a stalled ribosome. Here, we use genetics and translation inhibitors to understand how ribosomes respond to CGA repeats. We find that CGA codon repeats result in a truncated polypeptide that is targeted for degradation by Ltn1, an E3 ubiquitin ligase involved in nonstop decay, although deletion of LTN1 does not improve expression downstream from CGA repeats. Expression downstream from CGA codons at residue 318, but not at residue 4, is improved by deletion of either ASC1 or HEL2, previously implicated in inhibition of translation by polybasic sequences. Thus, translation of CGA repeats likely causes ribosomes to stall and exploits known quality control systems. Expression downstream from CGA repeats at amino acid 4 is improved by paromomycin, an aminoglycoside that relaxes decoding specificity. Paromomycin has no effect if native tRNA(Arg(ICG)) is highly expressed, consistent with the idea that failure to efficiently decode CGA codons might occur in part due to rejection of the cognate tRNA(Arg(ICG)). Furthermore, expression downstream from CGA repeats is improved by inactivation of RPL1B, one of two genes encoding the universally conserved ribosomal protein L1. The effects of rpl1b-Δ and of either paromomycin or tRNA(Arg(ICG)) on CGA decoding are additive, suggesting that the rpl1b-Δ mutant suppresses CGA inhibition by means other than increased acceptance of tRNA(Arg(ICG)). Thus, inefficient decoding of CGA likely involves at least two independent defects in translation.

  12. Involvement of Cyclic Guanosine Monophosphate-Dependent Protein Kinase I in Renal Antifibrotic Effects of Serelaxin

    PubMed Central

    Wetzl, Veronika; Schinner, Elisabeth; Kees, Frieder; Hofmann, Franz; Faerber, Lothar; Schlossmann, Jens

    2016-01-01

    Introduction: Kidney fibrosis has shown to be ameliorated through the involvement of cyclic guanosine monophosphate (cGMP) and its dependent protein kinase I (cGKI). Serelaxin, the recombinant form of human relaxin-II, increases cGMP levels and has shown beneficial effects on kidney function in acute heart failure patients. Antifibrotic properties of serelaxin are supposed to be mediated via relaxin family peptide receptor 1 and subsequently enhanced nitric oxide/cGMP to inhibit transforming growth factor-β (TGF-β) signaling. This study examines the involvement of cGKI in the antifibrotic signaling of serelaxin. Methods and Results: Kidney fibrosis was induced by unilateral ureteral obstruction in wildtype (WT) and cGKI knock-out (KO) mice. After 7 days, renal antifibrotic effects of serelaxin were assessed. Serelaxin treatment for 7 days significantly increased cGMP in the kidney of WT and cGKI-KO. In WT, renal fibrosis was reduced through decreased accumulation of collagen1A1, total collagen, and fibronectin. The profibrotic connective tissue growth factor as well as myofibroblast differentiation were reduced and matrix metalloproteinases-2 and -9 were positively modulated after treatment. Moreover, Smad2 as well as extracellular signal-regulated kinase 1 (ERK1) phosphorylation were decreased, whereas phosphodiesterase (PDE) 5a phosphorylation was increased. However, these effects were not observed in cGKI-KO. Conclusion: Antifibrotic renal effects of serelaxin are mediated via cGMP/cGKI to inhibit Smad2- and ERK1-dependent TGF-β signaling and increased PDE5a phosphorylation. PMID:27462268

  13. Gadolinium-enhanced MRI features of acute gouty arthritis on top of chronic gouty involvement in different joints.

    PubMed

    Emad, Yasser; Ragab, Yasser; El-Naggar, Ahmed; El-Shaarawy, Nashwa; Abd-Allah, Mayada A; Gamal, Rania M; Fathy, Ahmed; Hawass, Mona; Rasker, Johannes J

    2015-11-01

    The aims of the current study are to describe gadolinium-enhanced MRI features of an acute flare of established gouty arthritis in different joints and to examine a possible association between serum uric acid and MRI signs indicative of ongoing inflammation and/or structural joint damage as well as association with disease characteristics and laboratory findings. Twenty-seven male patients with established chronic gout agreed to participate, mean age 47.6 years, and mean disease duration in months 43.2 (±31.8). For all patients, detailed demographic, disease characteristics, and laboratory findings were obtained and correlated with MRI findings. In 27 patients with established gout, a total of 50 MRI studies were performed of the following joints: feet joints (n = 23), ankles (n = 18), knees (n = 5), and hand and wrist joints (n = 4). MRI revealed capsular thickening in 19 patients, bone marrow edema (BME) in 15, soft tissue edema (STE) in 20, joint effusion in 21, bone erosions in 17, cartilaginous erosions in 4, and tenosynovitis in 9 cases. In 17 cases, tophaceous lesions were found. Post contrast MRI showed synovial thickening in seven cases. Positive correlations were observed between serum uric acid levels and the following MRI findings: capsular thickening (r = 0.552, p = 0.003), BME (r = 0.668, p ≤ 0.0001), STE (r = 0.559, p = 0.002), and tenosynovitis (r = 0.513, p = 0.006). Using MRI in chronic gout, important features can be detected like BME, minute cartilaginous erosions, and hypertrophic synovial inflammation in post contrast MR images. Serum uric acid (SUA) was positively correlated with capsular thickening, BME, STE, and tenosynovitis.

  14. A cathepsin F-like peptidase involved in barley grain protein mobilization, HvPap-1, is modulated by its own propeptide and by cystatins

    PubMed Central

    Diaz, Isabel

    2012-01-01

    Among the C1A cysteine proteases, the plant cathepsin F-like group has been poorly studied. This paper describes the molecular and functional characterization of the HvPap-1 cathepsin F-like protein from barley. This peptidase is N-glycosylated and has to be processed to become active by its own propeptide being an important modulator of the peptidase activity. The expression pattern of its mRNA and protein suggest that it is involved in different proteolytic processes in the barley plant. HvPap-1 peptidase has been purified in Escherichia coli and the recombinant protein is able to degrade different substrates, including barley grain proteins (hordeins, albumins, and globulins) stored in the barley endosperm. It has been localized in protein bodies and vesicles of the embryo and it is induced in aleurones by gibberellin treatment. These three features support the implication of HvPap-1 in storage protein mobilization during grain germination. In addition, a complex regulation exerted by the barley cystatins, which are cysteine protease inhibitors, and by its own propeptide, is also described PMID:22791822

  15. A cathepsin F-like peptidase involved in barley grain protein mobilization, HvPap-1, is modulated by its own propeptide and by cystatins.

    PubMed

    Cambra, Ines; Martinez, Manuel; Dáder, Beatriz; González-Melendi, Pablo; Gandullo, Jacinto; Santamaría, M Estrella; Diaz, Isabel

    2012-07-01

    Among the C1A cysteine proteases, the plant cathepsin F-like group has been poorly studied. This paper describes the molecular and functional characterization of the HvPap-1 cathepsin F-like protein from barley. This peptidase is N-glycosylated and has to be processed to become active by its own propeptide being an important modulator of the peptidase activity. The expression pattern of its mRNA and protein suggest that it is involved in different proteolytic processes in the barley plant. HvPap-1 peptidase has been purified in Escherichia coli and the recombinant protein is able to degrade different substrates, including barley grain proteins (hordeins, albumins, and globulins) stored in the barley endosperm. It has been localized in protein bodies and vesicles of the embryo and it is induced in aleurones by gibberellin treatment. These three features support the implication of HvPap-1 in storage protein mobilization during grain germination. In addition, a complex regulation exerted by the barley cystatins, which are cysteine protease inhibitors, and by its own propeptide, is also described.

  16. Classification of membrane protein types using Voting Feature Interval in combination with Chou's Pseudo Amino Acid Composition.

    PubMed

    Ali, Farman; Hayat, Maqsood

    2015-11-07

    Membrane protein is a major constituent of cell, performing numerous crucial functions in the cell. These functions are mostly concerned with membrane protein's types. Initially, membrane proteins types are classified through traditional methods and reasonable results were obtained using these methods. However, due to large exploration of protein sequences in databases, it is very difficult or sometimes impossible to classify through conventional methods, because it is laborious and wasting of time. Therefore, a new powerful discriminating model is indispensable for classification of membrane protein's types with high precision. In this work, a quite promising classification model is developed having effective discriminating power of membrane protein's types. In our classification model, silent features of protein sequences are extracted via Pseudo Amino Acid Composition. Five classification algorithms were utilized. Among these classification algorithms Voting Feature Interval has obtained outstanding performance in all the three datasets. The accuracy of proposed model is 93.9% on dataset S1, 89.33% on S2 and 86.9% on dataset S3, respectively, applying 10-fold cross validation test. The success rates revealed that our proposed model has obtained the utmost outcomes than other existing models in literatures so far and will be played a substantial role in the fields of drug design and pharmaceutical industry.

  17. cDNA Library Screening Identifies Protein Interactors Potentially Involved in Non-Telomeric Roles of Arabidopsis Telomerase.

    PubMed

    Dokládal, Ladislav; Honys, David; Rana, Rajiv; Lee, Lan-Ying; Gelvin, Stanton B; Sýkorová, Eva

    2015-01-01

    Telomerase-reverse transcriptase (TERT) plays an essential catalytic role in maintaining telomeres. However, in animal systems telomerase plays additional non-telomeric functional roles. We previously screened an Arabidopsis cDNA library for proteins that interact with the C-terminal extension (CTE) TERT domain and identified a nuclear-localized protein that contains an RNA recognition motif (RRM). This RRM-protein forms homodimers in both plants and yeast. Mutation of the gene encoding the RRM-protein had no detectable effect on plant growth and development, nor did it affect telomerase activity or telomere length in vivo, suggesting a non-telomeric role for TERT/RRM-protein complexes. The gene encoding the RRM-protein is highly expressed in leaf and reproductive tissues. We further screened an Arabidopsis cDNA library for proteins that interact with the RRM-protein and identified five interactors. These proteins are involved in numerous non-telomere-associated cellular activities. In plants, the RRM-protein, both alone and in a complex with its interactors, localizes to nuclear speckles. Transcriptional analyses in wild-type and rrm mutant plants, as well as transcriptional co-analyses, suggest that TERT, the RRM-protein, and the RRM-protein interactors may play important roles in non-telomeric cellular functions.

  18. Collagen-induced platelet activation mainly involves the protein kinase C pathway.

    PubMed Central

    Karniguian, A; Grelac, F; Levy-Toledano, S; Legrand, Y J; Rendu, F

    1990-01-01

    This study analyses early biochemical events in collagen-induced platelet activation. An early metabolic event occurring during the lag phase was the activation of PtdIns(4,5)P2-specific phospholipase C. Phosphatidic acid (PtdOH) formation, phosphorylation of P43 and P20, thromboxane B2 (TXB2) synthesis and platelet secretion began after the lag phase, and were similarly time-dependent, except for TXB2 synthesis, which was delayed. Collagen induced extensive P43 phosphorylation, whereas P20 phosphorylation was weak and always lower than with thrombin. The dose-response curves of P43 phosphorylation and granule secretion were similar, and both reached a peak at 7.5 micrograms of collagen/ml, a dose which induced half-maximal PtdOH and TXB2 formation. Sphingosine, assumed to inhibit protein kinase C, inhibited P43 phosphorylation and secretion in parallel. However, sphingosine was not specific for protein kinase C, since a 15 microM concentration, which did not inhibit P43 phosphorylation, blocked TXB2 synthesis by 50%. Sphingosine did not affect PtdOH formation at all, even at 100 microM, suggesting that collagen itself induced this PtdOH formation, independently of TXB2 generation. The absence of external Ca2+ allowed the cleavage of polyphosphoinositides and the accumulation of InsP3 to occur, but impaired P43 phosphorylation, PtdOH and TXB2 formation, and secretion; these were only restored by adding 0.11 microM-Ca2+. In conclusion, stimulation of platelet membrane receptors for collagen initiates a PtdInsP2-specific phospholipase C activation, which is independent of external Ca2+, and might be the immediate receptor-linked response. A Ca2+ influx is indispensable to the triggering of subsequent platelet responses. This stimulation predominantly involves the protein kinase C pathway associated with secretion, and appears not to be mediated by TXB2, at least during its initial stage. Images Fig. 6. PMID:2163606

  19. A mammalian germ cell-specific RNA-binding protein interacts with ubiquitously expressed proteins involved in splice site selection

    NASA Astrophysics Data System (ADS)

    Elliott, David J.; Bourgeois, Cyril F.; Klink, Albrecht; Stévenin, James; Cooke, Howard J.

    2000-05-01

    RNA-binding motif (RBM) genes are found on all mammalian Y chromosomes and are implicated in spermatogenesis. Within human germ cells, RBM protein shows a similar nuclear distribution to components of the pre-mRNA splicing machinery. To address the function of RBM, we have used protein-protein interaction assays to test for possible physical interactions between these proteins. We find that RBM protein directly interacts with members of the SR family of splicing factors and, in addition, strongly interacts with itself. We have mapped the protein domains responsible for mediating these interactions and expressed the mouse RBM interaction region as a bacterial fusion protein. This fusion protein can pull-down several functionally active SR protein species from cell extracts. Depletion and add-back experiments indicate that these SR proteins are the only splicing factors bound by RBM which are required for the splicing of a panel of pre-mRNAs. Our results suggest that RBM protein is an evolutionarily conserved mammalian splicing regulator which operates as a germ cell-specific cofactor for more ubiquitously expressed pre-mRNA splicing activators.

  20. Surface Proteins of Gram-Positive Pathogens: Using Crystallography to Uncover Novel Features in Drug and Vaccine Candidates

    NASA Astrophysics Data System (ADS)

    Baker, Edward N.; Proft, Thomas; Kang, Haejoo

    Proteins displayed on the cell surfaces of pathogenic organisms are the front-line troops of bacterial attack, playing critical roles in colonization, infection and virulence. Although such proteins can often be recognized from genome sequence data, through characteristic sequence motifs, their functions are often unknown. One such group of surface proteins is attached to the cell surface of Gram-positive pathogens through the action of sortase enzymes. Some of these proteins are now known to form pili: long filamentous structures that mediate attachment to human cells. Crystallographic analyses of these and other cell surface proteins have uncovered novel features in their structure, assembly and stability, including the presence of inter- and intramolecular isopeptide crosslinks. This improved understanding of structures on the bacterial cell surface offers opportunities for the development of some new drug targets and for novel approaches to vaccine design.

  1. HybridGO-Loc: mining hybrid features on gene ontology for predicting subcellular localization of multi-location proteins.

    PubMed

    Wan, Shibiao; Mak, Man-Wai; Kung, Sun-Yuan

    2014-01-01

    Protein subcellular localization prediction, as an essential step to elucidate the functions in vivo of proteins and identify drugs targets, has been extensively studied in previous decades. Instead of only determining subcellular localization of single-label proteins, recent studies have focused on predicting both single- and multi-location proteins. Computational methods based on Gene Ontology (GO) have been demonstrated to be superior to methods based on other features. However, existing GO-based methods focus on the occurrences of GO terms and disregard their relationships. This paper proposes a multi-label subcellular-localization predictor, namely HybridGO-Loc, that leverages not only the GO term occurrences but also the inter-term relationships. This is achieved by hybridizing the GO frequencies of occurrences and the semantic similarity between GO terms. Given a protein, a set of GO terms are retrieved by searching against the gene ontology database, using the accession numbers of homologous proteins obtained via BLAST search as the keys. The frequency of GO occurrences and semantic similarity (SS) between GO terms are used to formulate frequency vectors and semantic similarity vectors, respectively, which are subsequently hybridized to construct fusion vectors. An adaptive-decision based multi-label support vector machine (SVM) classifier is proposed to classify the fusion vectors. Experimental results based on recent benchmark datasets and a new dataset containing novel proteins show that the proposed hybrid-feature predictor significantly outperforms predictors based on individual GO features as well as other state-of-the-art predictors. For readers' convenience, the HybridGO-Loc server, which is for predicting virus or plant proteins, is available online at http://bioinfo.eie.polyu.edu.hk/HybridGoServer/.

  2. HybridGO-Loc: Mining Hybrid Features on Gene Ontology for Predicting Subcellular Localization of Multi-Location Proteins

    PubMed Central

    Wan, Shibiao; Mak, Man-Wai; Kung, Sun-Yuan

    2014-01-01

    Protein subcellular localization prediction, as an essential step to elucidate the functions in vivo of proteins and identify drugs targets, has been extensively studied in previous decades. Instead of only determining subcellular localization of single-label proteins, recent studies have focused on predicting both single- and multi-location proteins. Computational methods based on Gene Ontology (GO) have been demonstrated to be superior to methods based on other features. However, existing GO-based methods focus on the occurrences of GO terms and disregard their relationships. This paper proposes a multi-label subcellular-localization predictor, namely HybridGO-Loc, that leverages not only the GO term occurrences but also the inter-term relationships. This is achieved by hybridizing the GO frequencies of occurrences and the semantic similarity between GO terms. Given a protein, a set of GO terms are retrieved by searching against the gene ontology database, using the accession numbers of homologous proteins obtained via BLAST search as the keys. The frequency of GO occurrences and semantic similarity (SS) between GO terms are used to formulate frequency vectors and semantic similarity vectors, respectively, which are subsequently hybridized to construct fusion vectors. An adaptive-decision based multi-label support vector machine (SVM) classifier is proposed to classify the fusion vectors. Experimental results based on recent benchmark datasets and a new dataset containing novel proteins show that the proposed hybrid-feature predictor significantly outperforms predictors based on individual GO features as well as other state-of-the-art predictors. For readers' convenience, the HybridGO-Loc server, which is for predicting virus or plant proteins, is available online at http://bioinfo.eie.polyu.edu.hk/HybridGoServer/. PMID:24647341

  3. Nicotine-induced plasticity in the retinocollicular pathway: Evidence for involvement of amyloid precursor protein.

    PubMed

    Gonçalves, R G J; Vasques, J F; Trindade, P; Serfaty, C A; Campello-Costa, P; Faria-Melibeu, A C

    2016-01-28

    During early postnatal development retinocollicular projections undergo activity-dependent synaptic refinement that results in the formation of precise topographical maps in the visual layers of the superior colliculus (SC). Amyloid Precursor Protein (APP) is a widely expressed transmembrane glycoprotein involved in the regulation of several aspects of neural development, such as neurite outgrowth, synapse formation and plasticity. Stimulation of cholinergic system has been found to alter the expression and processing of APP in different cell lines. Herein, we investigated the effect of nicotine on the development of retinocollicular pathway and on APP metabolism in the SC of pigmented rats. Animals were submitted to intracranial Elvax implants loaded with nicotine or phosphate-buffered saline (vehicle) at postnatal day (PND) 7. The ipsilateral retinocollicular pathway of control and experimental groups was anterogradely labeled either 1 or 3 weeks after surgery (PND 14 or PND 28). Local nicotine exposure produces a transitory sprouting of uncrossed retinal axons outside their main terminal zones. Nicotine also increases APP content and its soluble neurotrophic fragment sAPPα. Furthermore, nicotine treatment upregulates nicotinic acetylcholine receptor α7 and β2 subunits. Taken together, these data indicate that nicotine disrupts the ordering and topographic mapping of axons in the retinocollicular pathway and facilitates APP processing through the nonamyloidogenic pathway, suggesting that sAPPα may act as a trophic agent that mediates nicotine-induced morphological plasticity.

  4. The involvement of heat-shock proteins in the pathogenesis of autoimmune arthritis: a critical appraisal

    PubMed Central

    Huang, Min-Nung; Yu, Hua; Moudgil, Kamal D.

    2012-01-01

    Objectives To review the literature on the role of heat-shock proteins (HSPs) in the pathogenesis of autoimmune arthritis in animal models ans patients with rheumatoid arthritis (RA). Methods The published literature in Medline (PubMed), including our published work on the cell-mediated as well as humoral immune response to various HSPs was reviewed. Studies in both the pre-clinical animal models of arthritis as well as RA were examined critically and the data presented. Results In experimental arthritis, disease induction by different arthritogenic stimuli, including an adjuvant, led to immune response to mycobacterial HSP65 (BHSP65). However, attempts to induce arthritis by a purified HSP have not met with success. There are several reports of a significant immune response to HSP65 in RA patients. But, the issue of cause and effect is difficult to address. Nevertheless, several studies in animal models and a couple of clinical trials in RA patients have shown the beneficial effect of HSPs against autoimmune arthritis. Conclusions There is a clear association between immune response to HSPs, particularly HSP65, and the initiation and propagation of autoimmune arthritis in experimental models. The correlation is relatively less convincing in RA patients. In both cases, the ability of HSPs to modulate arthritis offers support, albeit an indirect one, for the involvement of these antigens in the disease process. PMID:19969325

  5. The Lowe syndrome protein OCRL1 is involved in primary cilia assembly.

    PubMed

    Coon, Brian G; Hernandez, Victor; Madhivanan, Kayalvizhi; Mukherjee, Debarati; Hanna, Claudia B; Barinaga-Rementeria Ramirez, Irene; Lowe, Martin; Beales, Philip L; Aguilar, R Claudio

    2012-04-15

    Lowe syndrome (LS) is a devastating, X-linked genetic disease characterized by the presence of congenital cataracts, profound learning disabilities and renal dysfunction. Unfortunately, children affected with LS often die early of health complications including renal failure. Although this syndrome was first described in the early 1950s and the affected gene, OCRL1, was identified more than 17 years ago, the mechanism by which Ocrl1 defects lead to LS's symptoms remains unknown. Here we show that LS display characteristics of a ciliopathy. Specifically, we found that patients' cells have defects in the assembly of primary cilia and this phenotype was reproduced in cell lines by knock-down of Ocrl1. Importantly, this defect could be rescued by re-introduction of WT Ocrl1 in both patient and Ocrl1 knock-down cells. In addition, a zebrafish animal model of LS exhibited cilia defects and multiple morphological and anatomical abnormalities typically seen in ciliopathies. Mechanistically, we show that Ocrl1 is involved in protein trafficking to the primary cilia in an Rab8-and IPIP27/Ses-dependent manner. Taking into consideration the relevance of the signaling pathways hosted by the primary cilium, our results suggest hitherto unrecognized mechanisms by which Ocrl1 deficiency may contribute to the phenotypic characteristics of LS. This conceptual change in our understanding of the disease etiology may provide an alternative avenue for the development of therapies.

  6. Protein SUMOylation is Involved in Cell-cycle Progression and Cell Morphology in Giardia lamblia.

    PubMed

    Di Genova, Bruno M; da Silva, Richard C; da Cunha, Júlia P C; Gargantini, Pablo R; Mortara, Renato A; Tonelli, Renata R

    2016-11-19

    The unicellular protozoa Giardia lamblia is a food- and waterborne parasite that causes giardiasis. This illness is manifested as acute and self-limited diarrhea and can evolve to long-term complications. Successful establishment of infection by Giardia trophozoites requires adhesion to host cells and colonization of the small intestine, where parasites multiply by mitotic division. The tight binding of trophozoites to host cells occurs by means of the ventral adhesive disc, a spiral array of microtubules and associated proteins such as giardins. In this work we show that knock down of the Small Ubiquitin-like MOdifier (SUMO) results in less adhesive trophzoites, decreased cell proliferation and deep morphological alterations, including at the ventral disc. Consistent with the reduced proliferation, SUMO knocked-down trophozoites were arrested in G1 and in S phases of the cell cycle. Mass spectrometry analysis of anti-SUMO immunoprecipitates was performed to identify SUMO substrates possibly involved in these events. Among the identified SUMOylation targets, α-tubulin was further validated by Western blot and confirmed to be a SUMO target in Giardia trophozoites.

  7. The involvement of multidrug and toxin extrusion protein 1 in the distribution and excretion of berberine.

    PubMed

    Xiao, Ling; Xue, Yaru; Zhang, Cuifeng; Wang, Le; Lin, Yunfei; Pan, Guoyu

    2017-03-16

    1. Berberine (BBR), an isoquinoline alkaloid, has demonstrated multiple clinical pharmacological actions. As a substrate of multiple transporters in the liver, BBR is rarely excreted into the bile but can be found in the urine. The purpose of the present study was to investigate the role of multidrug and toxin extrusion protein 1 (MATE1) in the transport of BBR in the liver and kidney. 2. Using human MATE1 (hMATE1)-transfected HEK293 cells, BBR was shown to be a substrate of hMATE1 (Km = 4.28 ± 2.18 μM). In primary rat hepatocytes, pH-dependent uptake and efflux studies suggested that the transport of BBR was driven by the exchange of H(+) and involved Mate1. In rats, we found that pyrimethamine (PYR), an inhibitor of Mate1, increased hepatic and renal distribution of BBR and decreased systematic excretion of BBR. 3. These findings indicated that BBR is a substrate of MATE1 and that hepatic and renal Mate1 promote excretion of BBR into bile and urine, respectively. In conclusion, Mate1 plays a key role in the distribution and excretion of BBR, and we speculate that drug-drug interactions (DDIs) caused by MATE1 may occur between BBR and other co-administered drugs.

  8. Aminophospholipid translocation in erythrocytes: Evidence for the involvement of a specific transporter and an endofacial protein

    SciTech Connect

    Connor, J.; Schroit, A.J. )

    1990-01-09

    The transport of exogenously supplied fluorescent analogues of aminophospholipids from the outer to inner leaflet in red blood cells (RBC) is dependent upon the oxidative status of membrane sulfhydryls. Oxidation of a sulfhydryl on a 32-kDa membrane protein by pyridyldithioethylamine (PDA) has been previously shown to inhibit the transport of NBD-labeled phosphatidylserine (NBD-PS). In the present study, other sulfhydryl oxidants were examined to determine whether additional sites are involved in the transport process. The results show that diamide inhibits the transport of NBD-PS via a mechanism that is independent of the 32-kDa site. This is shown by the inability of diamide to block labeling of the 32-kDa sulfhydryl with {sup 125}I-labeled PDA and to protect against PDA-mediated inhibition of NBD-PS transport. Diamide-mediated inhibition, but not PDA-mediated inhibition, could be reversed by reduction with cysteamine or endogenous glutathione. Once established, the asymmetric distribution of NBD-PS could not be altered by oxidation of either site. These data indicate that a second site critical to the transport of aminophospholipids residues on the endofacial surface and suggest that the transport of aminophospholipids across the bilayer membrane of RBC depends on a coordinated and complementary process between a cytoskeletal component and the 32-kDa membrane polypeptide; both must be operative for transport to proceed.

  9. Large-scale study of the interactions between proteins involved in type IV pilus biology in Neisseria meningitidis: characterization of a subcomplex involved in pilus assembly.

    PubMed

    Georgiadou, Michaella; Castagnini, Marta; Karimova, Gouzel; Ladant, Daniel; Pelicic, Vladimir

    2012-06-01

    The functionally versatile type IV pili (Tfp) are one of the most widespread virulence factors in bacteria. However, despite generating much research interest for decades, the molecular mechanisms underpinning the various aspects of Tfp biology remain poorly understood, mainly because of the complexity of the system. In the human pathogen Neisseria meningitidis for example, 23 proteins are dedicated to Tfp biology, 15 of which are essential for pilus biogenesis. One of the important gaps in our knowledge concerns the topology of this multiprotein machinery. Here we have used a bacterial two-hybrid system to identify and quantify the interactions between 11 Pil proteins from N. meningitidis. We identified 20 different binary interactions, many of which are novel. This represents the most complex interaction network between Pil proteins reported to date and indicates, among other things, that PilE, PilM, PilN and PilO, which are involved in pilus assembly, indeed interact. We focused our efforts on this subset of proteins and used a battery of assays to determine the membrane topology of PilN and PilO, map the interaction domains between PilE, PilM, PilN and PilO, and show that a widely conserved N-terminal motif in PilN is essential for both PilM-PilN interactions and pilus assembly. Finally, we show that PilP (another protein involved in pilus assembly) forms a complex with PilM, PilN and PilO. Taken together, these findings have numerous implications for understanding Tfp biology and provide a useful blueprint for future studies.

  10. Quantitative feature extraction reveals the status quo of protein fibrillation in the cell nucleus.

    PubMed

    Arnhold, Florian; von Mikecz, Anna

    2011-07-01

    Stepwise fibrillation of otherwise soluble proteins to insoluble amyloid-like protein aggregates is a hallmark of neurodegenerative protein-misfolding diseases including Alzheimer's, polyglutamine diseases, and the prion encephalopathies. Investigation of protein aggregation mechanisms has considerably advanced in vitro due to recent technical innovation, whereas the development of analyses tools for intracellular protein fibrillation remains a major challenge. Here, we introduce a method that enables monitoring of the protein fibrillation status in the cell nucleus. We show that the amyloid indicator Congo red can be induced to bind to distinct nucleoplasmic microdomains that are describable by application of discrete mathematics on the image information. Since formation of Congo red-binding nuclear microdomains (CRBDs) correlates with increased amyloid formation and decreased solubility of endogenous proteins with homopolymeric polyglutamine (polyQ) stretches we introduce the idea that different protein fibrillation steps can be characterized intracellularly by graph theory-aided pattern recognition.

  11. Using co-expression analysis and stress-based screens to uncover Arabidopsis peroxisomal proteins involved in drought response

    DOE PAGES

    Li, Jiying; Hu, Jianping; Bassham, Diane

    2015-09-14

    Peroxisomes are essential organelles that house a wide array of metabolic reactions important for plant growth and development. However, our knowledge regarding the role of peroxisomal proteins in various biological processes, including plant stress response, is still incomplete. Recent proteomic studies of plant peroxisomes significantly increased the number of known peroxisomal proteins and greatly facilitated the study of peroxisomes at the systems level. The objectives of this study were to determine whether genes that encode peroxisomal proteins with related functions are co-expressed in Arabidopsis and identify peroxisomal proteins involved in stress response using in silico analysis and mutant screens. Usingmore » microarray data from online databases, we performed hierarchical clustering analysis to generate a comprehensive view of transcript level changes for Arabidopsis peroxisomal genes during development and under abiotic and biotic stress conditions. Many genes involved in the same metabolic pathways exhibited co-expression, some genes known to be involved in stress response are regulated by the corresponding stress conditions, and function of some peroxisomal proteins could be predicted based on their coexpression pattern. Since drought caused expression changes to the highest number of genes that encode peroxisomal proteins, we subjected a subset of Arabidopsis peroxisomal mutants to a drought stress assay. Mutants of the LON2 protease and the photorespiratory enzyme hydroxypyruvate reductase 1 (HPR1) showed enhanced susceptibility to drought, suggesting the involvement of peroxisomal quality control and photorespiration in drought resistance. Lastly, our study provided a global view of how genes that encode peroxisomal proteins respond to developmental and environmental cues and began to reveal additional peroxisomal proteins involved in stress response, thus opening up new avenues to investigate the role of peroxisomes in plant adaptation to

  12. Using co-expression analysis and stress-based screens to uncover Arabidopsis peroxisomal proteins involved in drought response

    SciTech Connect

    Li, Jiying; Hu, Jianping; Bassham, Diane

    2015-09-14

    Peroxisomes are essential organelles that house a wide array of metabolic reactions important for plant growth and development. However, our knowledge regarding the role of peroxisomal proteins in various biological processes, including plant stress response, is still incomplete. Recent proteomic studies of plant peroxisomes significantly increased the number of known peroxisomal proteins and greatly facilitated the study of peroxisomes at the systems level. The objectives of this study were to determine whether genes that encode peroxisomal proteins with related functions are co-expressed in Arabidopsis and identify peroxisomal proteins involved in stress response using in silico analysis and mutant screens. Using microarray data from online databases, we performed hierarchical clustering analysis to generate a comprehensive view of transcript level changes for Arabidopsis peroxisomal genes during development and under abiotic and biotic stress conditions. Many genes involved in the same metabolic pathways exhibited co-expression, some genes known to be involved in stress response are regulated by the corresponding stress conditions, and function of some peroxisomal proteins could be predicted based on their coexpression pattern. Since drought caused expression changes to the highest number of genes that encode peroxisomal proteins, we subjected a subset of Arabidopsis peroxisomal mutants to a drought stress assay. Mutants of the LON2 protease and the photorespiratory enzyme hydroxypyruvate reductase 1 (HPR1) showed enhanced susceptibility to drought, suggesting the involvement of peroxisomal quality control and photorespiration in drought resistance. Lastly, our study provided a global view of how genes that encode peroxisomal proteins respond to developmental and environmental cues and began to reveal additional peroxisomal proteins involved in stress response, thus opening up new avenues to investigate the role of peroxisomes in plant adaptation to

  13. Prediction of protein subcellular localization by incorporating multiobjective PSO-based feature subset selection into the general form of Chou's PseAAC.

    PubMed

    Mandal, Monalisa; Mukhopadhyay, Anirban; Maulik, Ujjwal

    2015-04-01

    In this article, the possible subcellular location of a protein is predicted using multiobjective particle swarm optimization-based feature selection technique. In general form of pseudo-amino acid composition, the protein sequences are used for constructing protein features. Here, the different amino acids compositions are used to construct the feature sets. Therefore, the data are presented as sample of protein versus amino acid compositions as features. The proposed algorithm tries to maximize the feature relevance and minimize the feature redundancy simultaneously. After proposed algorithm is executed on the multiclass dataset, some features are selected. On this resultant feature subset, tenfold cross-validation is applied and corresponding accuracy, F score, entropy, representation entropy and average correlation are calculated. The performance of the proposed method is compared with that of its single objective versions, sequential forward search, sequential backward search, minimum redundancy maximum relevance with two schemes, CFS, CBFS, [Formula: see text], Fisher discriminant and a Cluster-based technique.

  14. Fibronectin-binding protein of Streptococcus pyogenes: sequence of the binding domain involved in adherence of streptococci to epithelial cells.

    PubMed Central

    Talay, S R; Valentin-Weigand, P; Jerlström, P G; Timmis, K N; Chhatwal, G S

    1992-01-01

    The sequence of the fibronectin-binding domain of the fibronectin-binding protein of Streptococcus pyogenes (Sfb protein) was determined, and its role in streptococcal adherence was investigated by use of an Sfb fusion protein in adherence studies. A 1-kb DNA fragment coding for the binding domain of Sfb protein was cloned into the expression vector pEX31 to produce an Sfb fusion protein consisting of the N-terminal part of MS2 polymerase and a C-terminal fragment of the streptococcal protein. Induction of the vector promoter resulted in hyperexpression of fibronectin-binding fusion protein in the cytoplasm of the recombinant Escherichia coli cells. Sequence determination of the cloned 1-kb fragment revealed an in-frame reading frame for a 268-amino-acid peptide composed of a 37-amino-acid sequence which is completely repeated three times and incompletely repeated a fourth time. Cloning of one repeat into pEX31 resulted in expression of small fusion peptides that show fibronectin-binding activity, indicating that one repeat contains at least one binding domain. Each repeat exhibits two charged domains and shows high homology with the 38-amino-acid D3 repeat of the fibronectin-binding protein of Staphylococcus aureus. Sequence comparison with other streptococcal ligand-binding surface proteins, including M protein, failed to reveal significant homology, which suggests that Sfb protein represents a novel type of functional protein in S. pyogenes. The Sfb fusion protein isolated from the cytoplasm of recombinant cells was purified by fast protein liquid chromatography. It showed a strong competitive inhibition of fibronectin binding to S. pyogenes and of the adherence of bacteria to cultured epithelial cells. In contrast, purified streptococcal lipoteichoic acid showed only a weak inhibition of fibronectin binding and streptococcal adherence. These results demonstrate that Sfb protein is directly involved in the fibronectin-mediated adherence of S. pyogenes to

  15. Systematic analysis of non-structural protein features for the prediction of PTM function potential by artificial neural networks.

    PubMed

    Dewhurst, Henry M; Torres, Matthew P

    2017-01-01

    Post-translational modifications (PTMs) provide an extensible framework for regulation of protein behavior beyond the diversity represented within the genome alone. While the rate of identification of PTMs has rapidly increased in recent years, our knowledge of PTM functionality encompasses less than 5% of this data. We previously developed SAPH-ire (Structural Analysis of PTM Hotspots) for the prioritization of eukaryotic PTMs based on function potential of discrete modified alignment positions (MAPs) in a set of 8 protein families. A proteome-wide expansion of the dataset to all families of PTM-bearing, eukaryotic proteins with a representational crystal structure and the application of artificial neural network (ANN) models demonstrated the broader applicability of this approach. Although structural features of proteins have been repeatedly demonstrated to be predictive of PTM functionality, the availability of adequately resolved 3D structures in the Protein Data Bank (PDB) limits the scope of these methods. In order to bridge this gap and capture the larger set of PTM-bearing proteins without an available, homologous structure, we explored all available MAP features as ANN inputs to identify predictive models that do not rely on 3D protein structural data. This systematic, algorithmic approach explores 8 available input features in exhaustive combinations (247 models; size 2-8). To control for potential bias in random sampling for holdback in training sets, we iterated each model across 100 randomized, sample training and testing sets-yielding 24,700 individual ANNs. The size of the analyzed dataset and iterative generation of ANNs represents the largest and most thorough investigation of predictive models for PTM functionality to date. Comparison of input layer combinations allows us to quantify ANN performance with a high degree of confidence and subsequently select a top-ranked, robust fit model which highlights 3,687 MAPs, including 10,933 PTMs with a high

  16. Systematic analysis of non-structural protein features for the prediction of PTM function potential by artificial neural networks

    PubMed Central

    2017-01-01

    Post-translational modifications (PTMs) provide an extensible framework for regulation of protein behavior beyond the diversity represented within the genome alone. While the rate of identification of PTMs has rapidly increased in recent years, our knowledge of PTM functionality encompasses less than 5% of this data. We previously developed SAPH-ire (Structural Analysis of PTM Hotspots) for the prioritization of eukaryotic PTMs based on function potential of discrete modified alignment positions (MAPs) in a set of 8 protein families. A proteome-wide expansion of the dataset to all families of PTM-bearing, eukaryotic proteins with a representational crystal structure and the application of artificial neural network (ANN) models demonstrated the broader applicability of this approach. Although structural features of proteins have been repeatedly demonstrated to be predictive of PTM functionality, the availability of adequately resolved 3D structures in the Protein Data Bank (PDB) limits the scope of these methods. In order to bridge this gap and capture the larger set of PTM-bearing proteins without an available, homologous structure, we explored all available MAP features as ANN inputs to identify predictive models that do not rely on 3D protein structural data. This systematic, algorithmic approach explores 8 available input features in exhaustive combinations (247 models; size 2–8). To control for potential bias in random sampling for holdback in training sets, we iterated each model across 100 randomized, sample training and testing sets—yielding 24,700 individual ANNs. The size of the analyzed dataset and iterative generation of ANNs represents the largest and most thorough investigation of predictive models for PTM functionality to date. Comparison of input layer combinations allows us to quantify ANN performance with a high degree of confidence and subsequently select a top-ranked, robust fit model which highlights 3,687 MAPs, including 10,933 PTMs with a

  17. Featured Article: Transcriptional landscape analysis identifies differently expressed genes involved in follicle-stimulating hormone induced postmenopausal osteoporosis.

    PubMed

    Maasalu, Katre; Laius, Ott; Zhytnik, Lidiia; Kõks, Sulev; Prans, Ele; Reimann, Ene; Märtson, Aare

    2017-01-01

    Osteoporosis is a disorder associated with bone tissue reorganization, bone mass, and mineral density. Osteoporosis can severely affect postmenopausal women, causing bone fragility and osteoporotic fractures. The aim of the current study was to compare blood mRNA profiles of postmenopausal women with and without osteoporosis, with the aim of finding different gene expressions and thus targets for future osteoporosis biomarker studies. Our study consisted of transcriptome analysis of whole blood serum from 12 elderly female osteoporotic patients and 12 non-osteoporotic elderly female controls. The transcriptome analysis was performed with RNA sequencing technology. For data analysis, the edgeR package of R Bioconductor was used. Two hundred and fourteen genes were expressed differently in osteoporotic compared with non-osteoporotic patients. Statistical analysis revealed 20 differently expressed genes with a false discovery rate of less than 1.47 × 10(-4) among osteoporotic patients. The expression of 10 genes were up-regulated and 10 down-regulated. Further statistical analysis identified a potential osteoporosis mRNA biomarker pattern consisting of six genes: CACNA1G, ALG13, SBK1, GGT7, MBNL3, and RIOK3. Functional ingenuity pathway analysis identified the strongest candidate genes with regard to potential involvement in a follicle-stimulating hormone activated network of increased osteoclast activity and hypogonadal bone loss. The differentially expressed genes identified in this study may contribute to future research of postmenopausal osteoporosis blood biomarkers.

  18. The RNase R from Campylobacter jejuni has unique features and is involved in the first steps of infection.

    PubMed

    Haddad, Nabila; Matos, Rute G; Pinto, Teresa; Rannou, Pauline; Cappelier, Jean-Michel; Prévost, Hervé; Arraiano, Cecília M

    2014-10-03

    Bacterial pathogens must adapt/respond rapidly to changing environmental conditions. Ribonucleases (RNases) can be crucial factors contributing to the fast adaptation of RNA levels to different environmental demands. It has been demonstrated that the exoribonuclease polynucleotide phosphorylase (PNPase) facilitates survival of Campylobacter jejuni in low temperatures and favors swimming, chick colonization, and cell adhesion/invasion. However, little is known about the mechanism of action of other ribonucleases in this microorganism. Members of the RNB family of enzymes have been shown to be involved in virulence of several pathogens. We have searched C. jejuni genome for homologues and found one candidate that displayed properties more similar to RNase R (Cj-RNR). We show here that Cj-RNR is important for the first steps of infection, the adhesion and invasion of C. jejuni to eukaryotic cells. Moreover, Cj-RNR proved to be active in a wide range of conditions. The results obtained lead us to conclude that Cj-RNR has an important role in the biology of this foodborne pathogen.

  19. Comparison of clinical features of left-sided infective endocarditis involving previously normal versus previously abnormal valves.

    PubMed

    Olmos, Carmen; Vilacosta, Isidre; Fernández, Cristina; Sarriá, Cristina; López, Javier; Del Trigo, María; Ferrera, Carlos; Vivas, David; Maroto, Luis; Hernández, Miguel; Rodríguez, Enrique; San Román, José Alberto

    2014-07-15

    Native valve infective endocarditis (IE) in patients with normal valves has increased in the last decades. Whether patients with normal valves present a similar prognosis to those with pathologic valves is unresolved. Our aim is to describe epidemiologic and clinical differences between patients with left-sided IE and normal valves and those with native pathologic valves. We analyzed 945 consecutive episodes of IE, 435 of which involved left-sided nonprosthetic IE. They were classified into 2 groups: episodes in normal valves (normal group, n=173) and episodes in pathologic valves (abnormal group, n=262). Patients in the normal group were younger, Staphylococcus aureus and Streptococcus bovis were more frequently isolated, and vegetations were more frequently found. Heart failure, septic shock, and the need for surgery or death were more common. Multivariate analysis identified the following as factors independently associated with normal valve IE: age<65 years, S bovis, S aureus, heart failure, and vegetation detection. Factors independently associated with in-hospital events included S aureus, periannular complications, heart failure, and septic shock development. In conclusion, compared with patients with abnormal valve IE, patients with IE on normal valves were younger, had a more virulent microbiological profile, developed heart failure and septic shock more frequently, needed more surgical procedures, and had worse prognosis.

  20. Promoter-specific trans activation and repression by human cytomegalovirus immediate-early proteins involves common and unique protein domains.

    PubMed Central

    Stenberg, R M; Fortney, J; Barlow, S W; Magrane, B P; Nelson, J A; Ghazal, P

    1990-01-01

    trans activation of promoters by viral regulatory proteins provides a useful tool to study coordinate control of gene expression. Immediate-early (IE) regions 1 and 2 of human cytomegalovirus (CMV) code for a series of proteins that originate from differentially spliced mRNAs. These IE proteins are proposed to regulate the temporal expression of the viral genome. To examine the structure and function of the IE proteins, we used linker insertion mutagenesis of the IE gene region as well as cDNA expression vector cloning of the abundant IE mRNAs. We showed that IE1 and IE2 proteins of CMV exhibit promoter-specific differences in their modes of action by either trans activating early and IE promoters or repressing the major IE promoter (MIEP). Transient cotransfection experiments with permissive human cells revealed a synergistic interaction between the 72- and the 86-kilodalton (kDa) IE proteins in trans activating an early promoter. In addition, transfection studies revealed that the 72-kDa protein was capable of trans activating the MIEP. In contrast, the 86-kDa protein specifically repressed the MIEP and this repression was suppressed by the 72-kDa protein. Furthermore, observations based on the primary sequence structure revealed a modular arrangement of putative regulatory motifs that could either potentiate or repress gene expression. These modular domains are either shared or unique among the IE proteins. From these data, we propose a model for IE protein function in the coordinate control of CMV gene expression. Images PMID:2157043

  1. [Septic arthritis in children with normal initial C-reactive protein: clinical and biological features].

    PubMed

    Basmaci, R; Ilharreborde, B; Bonacorsi, S; Kahil, M; Mallet, C; Aupiais, C; Doit, C; Dugué, S; Lorrot, M

    2014-11-01

    Septic arthritis has to be suspected in children with joint effusion and fever so as to perform joint aspiration, which will confirm the diagnosis by bacteriological methods, and to perform surgical treatment by joint lavage. Since development of current molecular methods, such as real-time PCR, Kingella kingae has become the first microbial agent of osteoarticular infections in young children, whereas Staphylococcus aureus is second. C-reactive protein (CRP) is an aid used to diagnose septic arthritis, but its elevation could be moderate. In a previous study, conducted at our hospital, 10% of children hospitalized for S. aureus or K. kingae septic arthritis had a CRP level<10 mg/L. To determine if diagnosis of septic arthritis could be made by other parameters, we analyzed the clinical and biologic features of these patients and compared them to those of children hospitalized for septic arthritis with initial CRP ≥10 mg/L. Among the 89 children with septic arthritis, 10% (n=9) had initial CRP<10 mg/L (K. kingae, n=5/63 ; S. aureus, n=4/26). Initial temperature and fibrinogen were significantly lower in the CRP<10 mg/L group than in the other (37.3°C vs. 37.9°C, P=0.039 and 4.19 vs. 5.72 g/L, P=0.003, respectively). Age, symptom duration before diagnosis, as well as leukocyte and platelet counts were similar in both groups. Two children (2/89=2.2%) with S. aureus septic arthritis had no fever, CRP elevation, or fibrinogen elevation. In the CRP-negative group, three of four children with S. aureus arthritis and one of five with K. kingae arthritis had a high CRP level (34, 40, 61, and 13 mg/L, respectively) 3 days after surgery and antibiotic treatment. One child with K. kingae septic arthritis and initial CRP<10 mg/L needed a second surgical drainage because of relapse of arthritis. In the S. aureus arthritis group, none of the children with initial CRP<10 mg/L experienced complications, while six of those with initial CRP≥10 mg/L needed a second surgical act

  2. The mechanism of binding staphylococcal protein A to immunoglobin G does not involve helix unwinding.

    PubMed

    Jendeberg, L; Tashiro, M; Tejero, R; Lyons, B A; Uhlén, M; Montelione, G T; Nilsson, B

    1996-01-09

    Structural changes in staphylococcal protein A (SpA) upon its binding to the constant region (Fc) of immunoglobulin G (IgG) have been studied by nuclear magnetic resonance and circular dichroism (CD) spectroscopy. The NMR solution structure of the engineered IgG-binding domain of SpA, the Z domain (an analogue of the B domain of SpA), has been determined by simulated annealing with molecular dynamics, using 599 distance and dihedral angle constraints. Domain Z contains three alpha-helices in the polypeptide segments Lys7 to His18 (helix 1), Glu25 to Asp36 (helix 2), and Ser41 to Ala54 (helix 3). The overall chain fold is an antiparallel three-helical bundle. This is in contrast to the previously determined X-ray structure of the similar SpA domain B in complex with Fc, where helix 3 is not observed in the electron density map [Deisenhofer, J. (1981) Biochemistry 20, 2361-2370], but similar to the solution NMR structure of domain B, which is also a three-helical bundle structure [Gouda, H., et al. (1992) Biochemistry 31, 9665-9672]. In order to characterize possible secondary structural changes associated with IgG binding, far-UV CD spectra were collected for the Z domain, an engineered repeat of this molecule (ZZ), recombinant Fc from IgG subclass 1 (Fc1), recombinant Fc from IgG subclass 3 (Fc3), and mixtures of Z/Fc1, Z/Fc3, ZZ/Fc1, and ZZ/Fc3. Fc3 was included as a control for possible changes of the CD spectrum in the mixture of noncomplexed molecules, since SpA is known not to bind Fc3. From these CD spectra, it was concluded that the third alpha-helix in Z is not disrupted in its complexes with Fc1. Similar results were obtained for the ZZ molecule. However, in both Z and ZZ there are some perturbations in CD spectra at high energy wavelengths (i.e., lambda < 215 nm) accompanying complex formation. On the basis of the combined CD and NMR results, as well as previously described binding studies of Z mutant proteins to Fc1, we conclude that the Z domain

  3. PIAS proteins are involved in the SUMO-1 modification, intracellular translocation and transcriptional repressive activity of RET finger protein

    SciTech Connect

    Matsuura, Tetsuo; Shimono, Yohei; Kawai, Kumi; Murakami, Hideki; Urano, Takeshi; Niwa, Yasumasa; Goto, Hidemi; Takahashi, Masahide . E-mail: mtakaha@med.nagoya-u.ac.jp

    2005-08-01

    Ret finger protein (RFP) is a nuclear protein that is highly expressed in testis and in various tumor cell lines. RFP functions as a transcriptional repressor and associates with Enhancer of Polycomb 1 (EPC1), a member of the Polycomb group proteins, and Mi-2{beta}, a main component of the nucleosome remodeling and deacetylase (NuRD) complex. We show that RFP binds with PIAS (protein inhibitor of activated STAT) proteins, PIAS1, PIAS3, PIASx{alpha} and PIASy at their carboxyl-terminal region and is covalently modified by SUMO-1 (sumoylation). PIAS proteins enhance the sumoylation of RFP in a dose-dependent manner and induce the translocation of RFP into nuclear bodies reminiscent of the PML bodies. In addition, co-expression of PIAS proteins or SUMO-1 strengthened the transcriptional repressive activity of RFP. Finally, our immunohistochemical results show that RFP, SUMO-1 and PIASy localize in a characteristic nuclear structure juxtaposed with the inner nuclear membrane (XY body) of primary spermatocytes in mouse testis. These results demonstrate that the intracellular location and the transcriptional activity of RFP are modified by PIAS proteins which possess SUMO E3 ligase activities and suggest that they may play a co-operative role in spermatogenesis.

  4. The microtubule motor protein KIF13A is involved in intracellular trafficking of the Lassa virus matrix protein Z.

    PubMed

    Fehling, Sarah Katharina; Noda, Takeshi; Maisner, Andrea; Lamp, Boris; Conzelmann, Karl-Klaus; Kawaoka, Yoshihiro; Klenk, Hans-Dieter; Garten, Wolfgang; Strecker, Thomas

    2013-02-01

    The small matrix protein Z of arenaviruses has been identified as the main driving force to promote viral particle production at the plasma membrane. Although multiple functions of Z in the arenaviral life cycle have been uncovered, the mechanism of intracellular transport of Z to the site of virus budding is poorly understood and cellular motor proteins that mediate Z trafficking remain to be identified. In the present study, we report that the Z protein of the Old World arenavirus Lassa virus (LASV) interacts with the kinesin family member 13A (KIF13A), a plus-end-directed microtubule-dependent motor protein. Plasmid-driven overexpression of KIF13A results in relocalization of Z to the cell periphery, while functional blockage of endogenous KIF13A by overexpression of a dominant-negative mutant or KIF13A-specific siRNA causes a perinuclearaccumulation and decreased production of both Z-induced virus-like particles and infectious LASV. The interaction of KIF13A with Z proteins from both Old and New World arenaviruses suggests a conserved intracellular transport mechanism. In contrast, the intracellular distribution of the matrix proteins of prototypic members of the paramyxo- and rhabdovirus family is independent of KIF13A. In summary, our studies identify for the first time a molecular motor protein as a critical mediator for intracellular microtubule-dependent transport of arenavirus matrix proteins.

  5. Platyhelminth Venom Allergen-Like (VAL) proteins: revealing structural diversity, class-specific features and biological associations across the phylum

    PubMed Central

    CHALMERS, IAIN W.; HOFFMANN, KARL F.

    2012-01-01

    SUMMARY During platyhelminth infection, a cocktail of proteins is released by the parasite to aid invasion, initiate feeding, facilitate adaptation and mediate modulation of the host immune response. Included amongst these proteins is the Venom Allergen-Like (VAL) family, part of the larger sperm coating protein/Tpx-1/Ag5/PR-1/Sc7 (SCP/TAPS) superfamily. To explore the significance of this protein family during Platyhelminthes development and host interactions, we systematically summarize all published proteomic, genomic and immunological investigations of the VAL protein family to date. By conducting new genomic and transcriptomic interrogations to identify over 200 VAL proteins (228) from species in all 4 traditional taxonomic classes (Trematoda, Cestoda, Monogenea and Turbellaria), we further expand our knowledge related to platyhelminth VAL diversity across the phylum. Subsequent phylogenetic and tertiary structural analyses reveal several class-specific VAL features, which likely indicate a range of roles mediated by this protein family. Our comprehensive analysis of platyhelminth VALs represents a unifying synopsis for understanding diversity within this protein family and a firm context in which to initiate future functional characterization of these enigmatic members. PMID:22717097

  6. Expression, stabilization and purification of membrane proteins via diverse protein synthesis systems and detergents involving cell-free associated with self-assembly peptide surfactants.

    PubMed

    Zheng, Xuan; Dong, Shuangshuang; Zheng, Jie; Li, Duanhua; Li, Feng; Luo, Zhongli

    2014-01-01

    G-protein coupled receptors (GPCRs) are involved in regulating most of physiological actions and metabolism in the bodies, which have become most frequently addressed therapeutic targets for various disorders and diseases. Purified GPCR-based drug discoveries have become routine that approaches to structural study, novel biophysical and biochemical function analyses. However, several bottlenecks that GPCR-directed drugs need to conquer the problems including overexpression, solubilization, and purification as well as stabilization. The breakthroughs are to obtain efficient protein yield and stabilize their functional conformation which are both urgently requiring of effective protein synthesis system methods and optimal surfactants. Cell-free protein synthesis system is superior to the high yields and post-translation modifications, and early signs of self-assembly peptide detergents also emerged to superiority in purification of membrane proteins. We herein focus several predominant protein synthesis systems and surfactants involving the novel peptide detergents, and uncover the advantages of cell-free protein synthesis system with self-assembling peptide detergents in purification of functional GPCRs. This review is useful to further study in membrane proteins as well as the new drug exploration.

  7. Proteomic profile of carbonylated proteins in rat liver: exercise attenuated oxidative stress may be involved in fatty liver improvement.

    PubMed

    Hu, Xiaofei; Duan, Zhigui; Hu, Hui; Li, Guolin; Yan, Siyu; Wu, Jinfeng; Wang, Jun; Yin, Dazhong; Xie, Qingji

    2013-05-01

    To screen target proteins of oxidative stress which mediate the effects of exercise on preventing nonalcoholic fatty liver disease (NAFLD), the methods for selecting carbonylated proteins were modified, and carbonylated proteins were profiled. The results showed that treadmill training reduced oxidative stress and the levels of intrahepatic triglyceride (IHTG). The changes in IHTG showed a significant positive correlation with oxidative stress as indicated by malondialdehyde level. Further results from proteomics illustrated that 17 functional proteins were susceptible to oxidative modification, and exercise protected three proteins from carbonylation. The latter three proteins may serve as both direct target proteins of oxidative stress and mediators contributing to the beneficial effects of exercise. In particular, a long-chain specific acyl-CoA dehydrogenase (ACADL) which was a key enzyme in lipid metabolism was not carbonylated and with higher activities in exercise group. These findings indicate that this modified technique is practical and powerful in selecting carbonylated proteins. Long-term treadmill training is effective in ameliorating oxidative stress and preventing the accumulation of IHTG. Among the 17 target proteins of oxidative modification, three proteins contribute to the beneficial effects of exercise. Preventing ACADL from carbonylation may be involved in the physiological mechanism of exercise-induced NAFLD improvement.

  8. Zinc Is Involved in Depression by Modulating G Protein-Coupled Receptor Heterodimerization.

    PubMed

    Tena-Campos, Mercè; Ramon, Eva; Lupala, Cecylia S; Pérez, Juan J; Koch, Karl-W; Garriga, Pere

    2016-04-01

    5-Hydroxytryptamine 1A receptor and galanin receptor 1 belong to the G protein-coupled receptors superfamily, and they have been described to heterodimerize triggering an anomalous physiological state that would underlie depression. Zinc supplementation has been widely reported to improve treatment against major depressive disorder. Our work has focused on the study and characterization of these receptors and its relationships with zinc both under purified conditions and in cell culture. To this aim, we have designed a strategy to purify the receptors in a conformationally active state. We have used receptors tagged with the monoclonal Rho-1D4 antibody and employed ligand-assisted purification in order to successfully purify both receptors in a properly folded and active state. The interaction between both purified receptors has been analyzed by surface plasmon resonance in order to determine the kinetics of dimerization. Zinc effect on heteromer has also been tested using the same methodology but exposing the 5-hydroxytryptamine 1A receptor to zinc before the binding experiment. These results, combined with Förster resonance energy transfer (FRET) measurements, in the absence and presence of zinc, suggest that this ion is capable of disrupting this interaction. Moreover, molecular modeling suggests that there is a coincidence between zinc-binding sites and heterodimerization interfaces for the serotonin receptor. Our results establish a rational explanation for the role of zinc in the molecular processes associated with receptor-receptor interactions and its relationship with depression, in agreement with previously reported evidence for the positive effects of zinc in depression treatment, and the involvement of our target dimer in the same disease.

  9. Overexpression of G protein-coupled receptors in cancer cells: involvement in tumor progression.

    PubMed

    Li, Shuyu; Huang, Shuguang; Peng, Sheng-Bin

    2005-11-01

    G protein-coupled receptors (GPCRs) play important roles in a variety of biological and pathological processes. They are considered among the most desirable targets for drug development. Recent studies have demonstrated that many GPCRs, such as endothelin receptors, chemokine receptors and lysophosphatidic acid receptors have been implicated in the tumorigenesis and metastasis of multiple human cancers. In this study, we conducted an in silico analysis of GPCR gene expression in primary human tumors by analyzing some publicly available gene expression profiling data. Statistical analysis was performed on eight microarray data sets of non-small cell lung cancer, breast cancer, prostate cancer, melanoma, gastric cancer and diffused large B cell lymphoma to identify GPCRs that are up-regulated in primary or metastatic cancer cells. Our analysis has demonstrated overexpression of several GPCRs in primary tumor cells, including chemokine receptors and protease-activated receptors that were shown to be important for tumorigenesis by previous studies. In addition, we have uncovered several GPCRs, such as neuropeptide receptors, adenosine A2B receptor, P2Y purinoceptor, calcium-sensing receptor and metabotropic glutamate receptors, that are expressed at a significantly higher level in some cancer tissue and may play a role in cancer progression. Analysis of cancer samples in different disease stages also suggests that some GPCRs, such as endothelin receptor A, may be involved in early tumor progression and others, such as CXCR4, may play a critical role in tumor invasion and metastasis. The present study demonstrates the value of publicly available microarray data as a resource to gain more understanding of cancer biology, to validate previous findings from in vitro experiments, and to identify potential novel anticancer targets and biomarkers.

  10. Structural and functional features of a collagen-binding matrix protein from the mussel byssus.

    PubMed

    Suhre, Michael H; Gertz, Melanie; Steegborn, Clemens; Scheibel, Thomas

    2014-02-26

    Blue mussels adhere to surfaces by the byssus, a holdfast structure composed of individual threads representing a collagen fibre reinforced composite. Here, we present the crystal structure and function of one of its matrix proteins, the proximal thread matrix protein 1, which is present in the proximal section of the byssus. The structure reveals two von Willebrand factor type A domains linked by a two-β-stranded linker yielding a novel structural arrangement. In vitro, the protein binds heterologous collagens with high affinity and affects collagen assembly, morphology and arrangement of its fibrils. By providing charged surface clusters as well as insufficiently coordinated metal ions, the proximal thread matrix protein 1 might interconnect other byssal proteins and thereby contribute to the integrity of the byssal threads in vivo. Moreover, the protein could be used for adjusting the mechanical properties of collagen materials, a function likely important in the natural byssus.

  11. A brief history of the search for the protein(s) involved in the acute regulation of steroidogenesis.

    PubMed

    Stocco, Douglas M; Zhao, Amy H; Tu, Lan N; Morohaku, Kanako; Selvaraj, Vimal

    2017-02-05

    The synthesis of steroid hormones occurs in specific cells and tissues in the body in response to trophic hormones and other signals. In order to synthesize steroids de novo, cholesterol, the precursor of all steroid hormones, must be mobilized from cellular stores to the inner mitochondrial membrane (IMM) to be converted into the first steroid formed, pregnenolone. This delivery of cholesterol to the IMM is the rate-limiting step in this process, and has long been known to require the rapid synthesis of a new protein(s) in response to stimulation. Although several possibilities for this protein have arisen over the past few decades, most of the recent attention to fill this role has centered on the candidacies of the proteins the Translocator Protein (TSPO) and the Steroidogenic Acute Regulatory Protein (StAR). In this review, the process of regulating steroidogenesis is briefly described, the characteristics of the candidate proteins and the data supporting their candidacies summarized, and some recent findings that propose a serious challenge for the role of TSPO in this process are discussed.

  12. Export of virulence proteins by malaria-infected erythrocytes involves remodeling of host actin cytoskeleton.

    PubMed

    Rug, Melanie; Cyrklaff, Marek; Mikkonen, Antti; Lemgruber, Leandro; Kuelzer, Simone; Sanchez, Cecilia P; Thompson, Jennifer; Hanssen, Eric; O'Neill, Matthew; Langer, Christine; Lanzer, Michael; Frischknecht, Friedrich; Maier, Alexander G; Cowman, Alan F

    2014-11-27

    Following invasion of human red blood cells (RBCs) by the malaria parasite, Plasmodium falciparum, a remarkable process of remodeling occurs in the host cell mediated by trafficking of several hundred effector proteins to the RBC compartment. The exported virulence protein, P falciparum erythrocyte membrane protein 1 (PfEMP1), is responsible for cytoadherence of infected cells to host endothelial receptors. Maurer clefts are organelles essential for protein trafficking, sorting, and assembly of protein complexes. Here we demonstrate that disruption of PfEMP1 trafficking protein 1 (PfPTP1) function leads to severe alterations in the architecture of Maurer's clefts. Furthermore, 2 major surface antigen families, PfEMP1 and STEVOR, are no longer displayed on the host cell surface leading to ablation of cytoadherence to host receptors. PfPTP1 functions in a large complex of proteins and is required for linking of Maurer's clefts to the host actin cytoskeleton.

  13. NMR Identification of the Binding Surfaces Involved in the Salmonella and Shigella Type III Secretion Tip-Translocon Protein-Protein Interactions

    PubMed Central

    McShan, Andrew C.; Kaur, Kawaljit; Chatterjee, Srirupa; Knight, Kevin M.; De Guzman, Roberto N.

    2017-01-01

    The type III secretion system (T3SS) is essential for the pathogenesis of many bacteria including Salmonella and Shigella, which together are responsible for millions of deaths worldwide each year. The structural component of the T3SS consists of the needle apparatus, which is assembled in part by the protein-protein interaction between the tip and the translocon. The atomic detail of the interaction between the tip and the translocon proteins is currently unknown. Here, we used NMR methods to identify that the N-terminal domain of the Salmonella SipB translocon protein interacts with the SipD tip protein at a surface at the distal region of the tip formed by the mixed α/β domain and a portion of its coiled-coil domain. Likewise, the Shigella IpaB translocon protein and the IpaD tip protein interact with each other using similar surfaces identified for the Salmonella homologs. Furthermore, removal of the extreme N-terminal residues of the translocon protein, previously thought to be important for the interaction, had little change on the binding surface. Finally, mutations at the binding surface of SipD reduced invasion of Salmonella into human intestinal epithelial cells. Together, these results reveal the binding surfaces involved in the tip-translocon protein-protein interaction and advance our understanding of the assembly of the T3SS needle apparatus. PMID:27093649

  14. Computational analysis of protein-protein interfaces involving an alpha helix: insights for terphenyl–like molecules binding

    PubMed Central

    2013-01-01

    Background Protein-Protein Interactions (PPIs) are key for many cellular processes. The characterization of PPI interfaces and the prediction of putative ligand binding sites and hot spot residues are essential to design efficient small-molecule modulators of PPI. Terphenyl and its derivatives are small organic molecules known to mimic one face of protein-binding alpha-helical peptides. In this work we focus on several PPIs mediated by alpha-helical peptides. Method We performed computational sequence- and structure-based analyses in order to evaluate several key physicochemical and surface properties of proteins known to interact with alpha-helical peptides and/or terphenyl and its derivatives. Results Sequence-based analysis revealed low sequence identity between some of the analyzed proteins binding alpha-helical peptides. Structure-based analysis was performed to calculate the volume, the fractal dimension roughness and the hydrophobicity of the binding regions. Besides the overall hydrophobic character of the binding pockets, some specificities were detected. We showed that the hydrophobicity is not uniformly distributed in different alpha-helix binding pockets that can help to identify key hydrophobic hot spots. Conclusions The presence of hydrophobic cavities at the protein surface with a more complex shape than the entire protein surface seems to be an important property related to the ability of proteins to bind alpha-helical peptides and low molecular weight mimetics. Characterization of similarities and specificities of PPI binding sites can be helpful for further development of small molecules targeting alpha-helix binding proteins. PMID:23768251

  15. The intrinsic conformational features of amino acids from a protein coil library and their applications in force field development.

    PubMed

    Jiang, Fan; Han, Wei; Wu, Yun-Dong

    2013-03-14

    The local conformational (φ, ψ, χ) preferences of amino acid residues remain an active research area, which are important for the development of protein force fields. In this perspective article, we first summarize spectroscopic studies of alanine-based short peptides in aqueous solution. While most studies indicate a preference for the P(II) conformation in the unfolded state over α and β conformations, significant variations are also observed. A statistical analysis from various coil libraries of high-resolution protein structures is then summarized, which gives a more coherent view of the local conformational features. The φ, ψ, χ distributions of the 20 amino acids have been obtained from a protein coil library, considering both backbone and side-chain conformational preferences. The intrinsic side-chain χ(1) rotamer preference and χ(1)-dependent Ramachandran plot can be generally understood by combining the interaction of the side-chain Cγ/Oγ atom with two neighboring backbone peptide groups. Current all-atom force fields such as AMBER ff99sb-ILDN, ff03 and OPLS-AA/L do not reproduce these distributions well. A method has been developed by combining the φ, ψ plot of alanine with the influence of side-chain χ(1) rotamers to derive the local conformational features of various amino acids. It has been further applied to improve the OPLS-AA force field. The modified force field (OPLS-AA/C) reproduces experimental (3)J coupling constants for various short peptides quite well. It also better reproduces the temperature-dependence of the helix-coil transition for alanine-based peptides. The new force field can fold a series of peptides and proteins with various secondary structures to their experimental structures. MD simulations of several globular proteins using the improved force field give significantly less deviation (RMSD) to experimental structures. The results indicate that the local conformational features from coil libraries are valuable for

  16. Mal-Lys: prediction of lysine malonylation sites in proteins integrated sequence-based features with mRMR feature selection

    PubMed Central

    Xu, Yan; Ding, Ya-Xin; Ding, Jun; Wu, Ling-Yun; Xue, Yu

    2016-01-01

    Lysine malonylation is an important post-translational modification (PTM) in proteins, and has been characterized to be associated with diseases. However, identifying malonyllysine sites still remains to be a great challenge due to the labor-intensive and time-consuming experiments. In view of this situation, the establishment of a useful computational method and the development of an efficient predictor are highly desired. In this study, a predictor Mal-Lys which incorporated residue sequence order information, position-specific amino acid propensity and physicochemical properties was proposed. A feature selection method of minimum Redundancy Maximum Relevance (mRMR) was used to select optimal ones from the whole features. With the leave-one-out validation, the value of the area under the curve (AUC) was calculated as 0.8143, whereas 6-, 8- and 10-fold cross-validations had similar AUC values which showed the robustness of the predictor Mal-Lys. The predictor also showed satisfying performance in the experimental data from the UniProt database. Meanwhile, a user-friendly web-server for Mal-Lys is accessible at http://app.aporc.org/Mal-Lys/. PMID:27910954

  17. Structural and functional features of the NAD(P) dependent Gfo/Idh/MocA protein family oxidoreductases.

    PubMed

    Taberman, Helena; Parkkinen, Tarja; Rouvinen, Juha

    2016-04-01

    The Gfo/Idh/MocA protein family contains a number of different proteins, which almost exclusively consist of NAD(P)-dependent oxidoreductases that have a diverse set of substrates, typically pyranoses. In this study, to clarify common structural features that would contribute to their function, the available crystal structures of the members of this family have been analyzed. Despite a very low sequence identity, the central features of the three-dimensional structures of the proteins are surprisingly similar. The members of the protein family have a two-domain structure consisting of a N-terminal nucleotide-binding domain and a C-terminal α/β-domain. The C-terminal domain contributes to the substrate binding and catalysis, and contains a βα-motif with a central α-helix carrying common essential amino acid residues. The β-sheet of the α/β-domain contributes to the oligomerization in most of the proteins in the family.

  18. Ligand-induced association of surface immunoglobulin with the detergent insoluble cytoskeleton may involve an 89K protein

    SciTech Connect

    Gupta, S.K.; Woda, B.

    1986-03-01

    Membrane immunoglobulin of B-lymphocytes is thought to play an important role in antigen recognition and cellular activation. Binding of cross-linking ligands to surface immunoglobulin (SIg) on intact cells converts it to a detergent insoluble state, and this conversion is associated with the transmission of a mitogenic signal. Insolubilized membrane proteins may be solubilized by incubating the detergent insoluble cytoskeletons in buffers which convert F-actin to G-actin ((Buffer 1), 0.34M sucrose, 0.5mM ATP, 0.5mM Dithiothrietol and lmM EDTA). Immunoprecipitation of SIg from the detergent soluble fraction of /sup 35/S-methionine labeled non ligand treated rat B-cells results in the co-isolation of an 89K protein and a 44K protein, presumably actin. The 89K protein is not associated with the fraction of endogenous detergent insoluble SIg. On treatment of rat B cells with cross-linking ligand (anti-Ig) the 89K protein becomes detergent insoluble along with most of the SIg and co-isolates with SIg on immunoprecipitation of the detergent insoluble, buffer l solubilized fraction. The migration of the SIg-associated 89K protein from the detergent soluble fraction to the detergent insoluble fraction after ligand treatment, suggests that this protein might be involved in linking SIg to the underlying cytoskeleton and could be involved in the transmission of a mitogenic signal.

  19. Proteomic analysis of differentially expressed proteins involved in ethylene-induced chilling tolerance in harvested banana fruit

    PubMed Central

    Li, Taotao; Yun, Ze; Zhang, Dandan; Yang, Chengwei; Zhu, Hong; Jiang, Yueming; Duan, Xuewu

    2015-01-01

    To better understand the mechanism involved in ethylene-induced chilling tolerance in harvested banana fruit, a gel-based proteomic study followed by MALDI-TOF-TOF MS was carried out. Banana fruit were treated with 500 ppm ethylene for 12 h and then stored at 6°C. During cold storage, the chilling tolerance was assessed and the proteins from the peel were extracted for proteomic analysis. It was observed that ethylene pretreatment significantly induced the chilling tolerance in harvested banana fruit, manifesting as increases in maximal chlorophyll fluorescence (Fv/Fm) and decreased electrolyte leakage. Sixty-four proteins spots with significant differences in abundance were identified, most of which were induced by ethylene pretreatment during cold storage. The up-regulated proteins induced by ethylene pretreatment were mainly related to energy metabolism, stress response and defense, methionine salvage cycle and protein metabolism. These proteins were involved in ATP synthesis, ROS scavenging, protective compounds synthesis, protein refolding and degradation, and polyamine biosynthesis. It is suggested that these up-regulated proteins might play a role in the ethylene-induced chilling tolerance in harvested banana fruit. PMID:26528309

  20. Features of Motion of Soliton Transported Bio-energy in Aperiodic α-Helix Protein Molecules with Three Channels

    NASA Astrophysics Data System (ADS)

    Pang, Xiao-Feng; Liu, Mei-Jie

    2009-01-01

    The structure aperiodicities can influence seriously the features of motion of soliton excited in the α-helix protein molecules with three channels. We study the influence of structure aperiodicities on the features of the soliton in the improved model by numerical simulation and Runge-Kulta method. The results obtained show that the new soliton is very robust against the structure aperiodicities including large disorder in the sequence of mass of the amino acids and fluctuations of spring constant, coupling constant, dipole-dipole interactional constant, ground state energy and chain-chain interaction. However, very strong structure aperiodicities can also destroy the stability of the soliton in the α-helix protein molecules.

  1. Conservation of proteins involved in oocyst wall formation in Eimeria maxima, Eimeria tenella and Eimeria acervulina.

    PubMed

    Belli, Sabina I; Ferguson, David J P; Katrib, Marilyn; Slapetova, Iveta; Mai, Kelly; Slapeta, Jan; Flowers, Sarah A; Miska, Kate B; Tomley, Fiona M; Shirley, Martin W; Wallach, Michael G; Smith, Nicholas C

    2009-08-01

    Vaccination with proteins from gametocytes of Eimeria maxima protects chickens, via transfer of maternal antibodies, against infection with several species of Eimeria. Antibodies to E. maxima gametocyte proteins recognise proteins in the wall forming bodies of macrogametocytes and oocyst walls of E. maxima, Eimeria tenella and Eimeria acervulina. Homologous genes for two major gametocyte proteins - GAM56 and GAM82 - were found in E. maxima, E. tenella and E. acervulina. Alignment of the predicted protein sequences of these genes reveals that, as well as sharing regions of tyrosine richness, strong homology exists in their amino-terminal regions, where protective antibodies bind. This study confirms the conservation of the roles of GAM56 and GAM82 in oocyst wall formation and shows that antibodies to gametocyte antigens of E. maxima cross-react with homologous proteins in other species, helping to explain cross-species maternal immunity.

  2. Involvement of histone phosphorylation in thymocyte apoptosis by protein phosphatase inhibitors.

    PubMed

    Lee, E; Nakatsuma, A; Hiraoka, R; Ishikawa, E; Enomoto, R; Yamauchi, A

    1999-07-01

    Incubation of rat thymocytes with the inhibitors of protein phosphatase such as calyculin A and okadaic acid resulted in an increase in DNA fragmentation. These effects were dependent on the concentration of the inhibitors and the incubation time. Analyses of the fragmented DNA revealed the production of approximately 50 kbp of DNA and a 180 bp DNA ladder. In addition, a laser scanning-microscopic analysis showed that these compounds caused nuclear condensation. Thus, these results demonstrated that protein phosphatase inhibitors induced thymocyte apoptosis. The inhibitors of protein phosphatase increased the phosphorylation of proteins of approximately 15 kDa. The phosphorylation of proteins preceded the DNA fragmentation induced by these inhibitors. Judging from acetic acid-urea-Triton X-100 gel electrophoresis, the phosphorylated proteins were histone H1 and H2A/H3. Therefore, these results suggest that phosphorylation of histones triggers the DNA fragmentation of thymocytes undergoing apoptosis.

  3. Biochemical localization of a protein involved in Gluconacetobacter hansenii cellulose synthesis

    SciTech Connect

    Iyer, Prashanti R; Catchmark, Jeffrey M; Brown, Nicole Robitaille; Tien, Ming

    2011-02-08

    Using subcellular fractionation and Western blot methods, we have shown that AcsD, one of the proteins encoded by the Acetobacter cellulose synthase (acs) operon, is localized in the periplasmic region of the cell. AcsD protein was heterologously expressed in Escherichia coli and purified using histidine tag affinity methods. The purified protein was used to obtain rabbit polyclonal antibodies. The purity of the subcellular fractions was assessed by marker enzyme assays.

  4. Dynamics of Arabidopsis SUN proteins during mitosis and their involvement in nuclear shaping.

    PubMed

    Oda, Yoshihisa; Fukuda, Hiroo

    2011-05-01

    The nuclear envelope (NE) is a highly active structure with a specific set of nuclear envelope proteins acting in diverse cellular events. SUN proteins are conserved NE proteins among eukaryotes. Although they form nucleocytoplasmic linkage complexes in metazoan cells, their functions in the plant kingdom are unknown. To understand the function of plant SUN proteins, in this study we first investigated the dynamics of Arabidopsis SUN proteins during mitosis in Arabidopsis roots and cultured cells. For this purpose, we performed dual and triple visualization of these proteins, microtubules, chromosomes, and endoplasmic reticulum (ER) in cultured cells, and observed their dynamics during mitosis using a high-speed spinning disk confocal microscope. The localizations of SUN proteins changed dynamically during mitosis, tightly coupled with NE dynamics. Moreover, NE re-formation marked with SUN proteins is temporally and spatially coordinated with plant-specific microtubule structures such as phragmoplasts. Finally, the analysis with gene knockdowns of AtSUN1 and AtSUN2 indicated that they are necessary for the maintenance and/or formation of polarized nuclear shape in root hairs. These results suggest that Arabidopsis SUN proteins function in the maintenance or formation of nuclear shape as components of the nucleocytoskeletal complex.

  5. Identification of domains of the Tomato spotted wilt virus NSm protein involved in tubule formation, movement and symptomatology

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Deletion and alanine-substitution mutants of the Tomato spotted wilt virus NSm protein were generated to identify domains involved in tubule formation, movement and symptomatology, using a heterologous expression system derived from Tobacco mosaic virus. Two regions of NSm were required for both tub...

  6. Proteins involved in wine aroma compounds metabolism by a Saccharomyces cerevisiae flor-velum yeast strain grown in two conditions.

    PubMed

    Moreno-García, Jaime; García-Martínez, Teresa; Millán, M Carmen; Mauricio, Juan Carlos; Moreno, Juan

    2015-10-01

    A proteomic and exometabolomic study was conducted on Saccharomyces cerevisiae flor yeast strain growing under biofilm formation condition (BFC) with ethanol and glycerol as carbon sources and results were compared with those obtained under no biofilm formation condition (NBFC) containing glucose as carbon source. By using modern techniques, OFFGEL fractionator and LTQ-Orbitrap for proteome and SBSE-TD-GC-MS for metabolite analysis, we quantified 84 proteins including 33 directly involved in the metabolism of glycerol, ethanol and 17 aroma compounds. Contents in acetaldehyde, acetic acid, decanoic acid, 1,1-diethoxyethane, benzaldehyde and 2-phenethyl acetate, changed above their odor thresholds under BFC, and those of decanoic acid, ethyl octanoate, ethyl decanoate and isoamyl acetate under NBFC. Of the twenty proteins involved in the metabolism of ethanol, acetaldehyde, acetoin, 2,3-butanediol, 1,1-diethoxyethane, benzaldehyde, organic acids and ethyl esters, only Adh2p, Ald4p, Cys4p, Fas3p, Met2p and Plb1p were detected under BFC and as many Acs2p, Ald3p, Cem1p, Ilv2p, Ilv6p and Pox1p, only under NBFC. Of the eight proteins involved in glycerol metabolism, Gut2p was detected only under BFC while Pgs1p and Rhr2p were under NBFC. Finally, of the five proteins involved in the metabolism of higher alcohols, Thi3p was present under BFC, and Aro8p and Bat2p were under NBFC.

  7. One common structural feature of "words" in protein sequences and human texts.

    PubMed

    Zemková, M; Trifonov, E N; Zahradník, D

    2014-01-01

    Frequently discussed analogy between genetic and human texts is explored by comparison of alternation of polar and non-polar amino-acid residues in proteins and alternation of consonants and vowels in human texts. In human languages, the usage of possible combinations of consonants and vowels is influenced by pronounceability of the combinations. Similarly, oligopeptide composition of proteins is influenced by requirements of protein folding and stability. One special type of structure often present in proteins is amphipathic α-helices in which polar and non-polar amino acids alternate with the period 3.5 residues, not unlike alternation of consonants and vowels. In this study, we evaluated the contribution made by amphipathic alternations to the protein sequence texts (20-24%). Their proportion is lower than respective values for alternating words in human texts (57-89%). The proteomes (full sets of proteins for selected organisms) were transformed into ranked sequences of n-grams (words of length n), including periodical amphipathic structures. Similarly, human texts were transformed into sequences of alternating consonants and vowels. Analysis of the vocabularies shows that in both types of texts (human languages and proteins) the alternating words are dominant or highly preferred, thus, strengthening the analogy between these two types of texts. The contribution of amphipathic words in the upper parts of the ranked lists for 10 analyzed proteomes varies between 58 and 74%. In human texts respective values range between 90 and 100%.

  8. Morphine-Induced Preconditioning: Involvement of Protein Kinase A and Mitochondrial Permeability Transition Pore

    PubMed Central

    Dorsch, Marianne; Behmenburg, Friederike; Raible, Miriam; Blase, Dominic; Grievink, Hilbert; Hollmann, Markus W.; Heinen, André; Huhn, Ragnar

    2016-01-01

    hearts STAT3 inhibitor Stattic completely abolished morphine-induced preconditioning. Administration of Stattic and mPTP inhibitor cyclosporine A reduced infarct size to 31±6% (Stat+CsA, P<0.05 vs. Con). Cyclosporine A alone reduced infarct size to 26±7% (CsA P<0.05 vs. Con). In cardiomyocytes, PKA activity was increased by morphine. Conclusion Our data suggest that morphine-induced cardioprotection is mediated by STAT3-activation and inhibition of mPTP, with STA3 located upstream of mPTP. There is some evidence that protein kinase A is involved within the signalling pathway. PMID:26968004

  9. "Features of two proteins of Leptospira interrogans with potential role in host-pathogen interactions"

    PubMed Central

    2012-01-01

    Background Leptospirosis is considered a re-emerging infectious disease caused by pathogenic spirochaetes of the genus Leptospira. Pathogenic leptospires have the ability to survive and disseminate to multiple organs after penetrating the host. Leptospires were shown to express surface proteins that interact with the extracellular matrix (ECM) and to plasminogen (PLG). This study examined the interaction of two putative leptospiral proteins with laminin, collagen Type I, collagen Type IV, cellular fibronectin, plasma fibronectin, PLG, factor H and C4bp. Results We show that two leptospiral proteins encoded by LIC11834 and LIC12253 genes interact with laminin in a dose - dependent and saturable mode, with dissociation equilibrium constants (KD) of 367.5 and 415.4 nM, respectively. These proteins were named Lsa33 and Lsa25 (Leptospiral surface adhesin) for LIC11834 and LIC12253, respectively. Metaperiodate - treated laminin reduced Lsa25 - laminin interaction, suggesting that sugar moieties of this ligand participate in this interaction. The Lsa33 is also PLG - binding receptor, with a KD of 23.53 nM, capable of generating plasmin in the presence of an activator. Although in a weak manner, both proteins interact with C4bp, a regulator of complement classical route. In silico analysis together with proteinase K and immunoflorescence data suggest that these proteins might be surface exposed. Moreover, the recombinant proteins partially inhibited leptospiral adherence to immobilized laminin and PLG. Conclusions We believe that these multifunctional proteins have the potential to participate in the interaction of leptospires to hosts by mediating adhesion and by helping the bacteria to escape the immune system and to overcome tissue barriers. To our knowledge, Lsa33 is the first leptospiral protein described to date with the capability of binding laminin, PLG and C4bp in vitro. PMID:22463075

  10. RNA-Binding Proteins in Trichomonas vaginalis: Atypical Multifunctional Proteins Involved in a Posttranscriptional Iron Regulatory Mechanism

    PubMed Central

    Figueroa-Angulo, Elisa E.; Calla-Choque, Jaeson S.; Mancilla-Olea, Maria Inocente; Arroyo, Rossana

    2015-01-01

    Iron homeostasis is highly regulated in vertebrates through a regulatory system mediated by RNA-protein interactions between the iron regulatory proteins (IRPs) that interact with an iron responsive element (IRE) located in certain mRNAs, dubbed the IRE-IRP regulatory system. Trichomonas vaginalis, the causal agent of trichomoniasis, presents high iron dependency to regulate its growth, metabolism, and virulence properties. Although T. vaginalis lacks IRPs or proteins with aconitase activity, possesses gene expression mechanisms of iron regulation at the transcriptional and posttranscriptional levels. However, only one gene with iron regulation at the transcriptional level has been described. Recently, our research group described an iron posttranscriptional regulatory mechanism in the T. vaginalis tvcp4 and tvcp12 cysteine proteinase mRNAs. The tvcp4 and tvcp12 mRNAs have a stem-loop structure in the 5'-coding region or in the 3'-UTR, respectively that interacts with T. vaginalis multifunctional proteins HSP70, α-Actinin, and Actin under iron starvation condition, causing translation inhibition or mRNA stabilization similar to the previously characterized IRE-IRP system in eukaryotes. Herein, we summarize recent progress and shed some light on atypical RNA-binding proteins that may participate in the iron posttranscriptional regulation in T. vaginalis. PMID:26703754

  11. The involvement of an integrin-like protein and protein kinase C in amoebic adhesion to fibronectin and amoebic cytotoxicity.

    PubMed

    Han, Kyu-Lee; Lee, Hyun-Ju; Shin, Myeong Heon; Shin, Ho-Joon; Im, Kyung-Il; Park, Soon-Jung

    2004-09-01

    Adherence of a pathogen to the host cell is one of the critical steps in microbial infections. Naegleria fowleri, a causative agent of primary amoebic meningoencephalitis in humans, is expected to interact with extracellular components of the host, such as fibronectin, in a receptor-mediated mode. In this study, we investigated the interaction between N. fowleri and fibronectin to understand its cytopathology. In binding assays using immobilized fibronectin, the number of amoebae bound to fibronectin was increased compared to the controls, and was dependent on the amount of coated fibronectin present. A fibronectin binding protein of 60 kDa was found in extracts of N. fowleri. Western blot and immunolocalization assays using integrin alpha(5)/FnR antibodies showed that a 60 kDa protein reacted with the antibodies in extracts of N. fowleri, which was localized on the surface of N. fowleri. Preincubation of N. fowleri with the integrin antibodies significantly inhibited amoebic binding to fibronectin and cytotoxicity to the CHO cells. Additionally, protein kinase C activity was detected in the extract of N. fowleri. When N. fowleri was pretreated with protein kinase C activator or inhibitor, the abilities of amoebic adhesion to fibronectin and cytotoxicity to the host cells were markedly affected compared to untreated amoebae. These results suggest that an amoebic integrin-like receptor and protein kinase C play important roles in amoebic cellular processes in response to fibronectin.

  12. Dynamic localization of two tobamovirus ORF6 proteins involves distinct organellar compartments.

    PubMed

    Gushchin, Vladimir A; Lukhovitskaya, Nina I; Andreev, Dmitri E; Wright, Kathryn M; Taliansky, Michael E; Solovyev, Andrey G; Morozov, Sergey Y; MacFarlane, Stuart A

    2013-01-01

    ORF6 is a small gene that overlaps the movement and coat protein genes of subgroup 1a tobamoviruses. The ORF6 protein of tomato mosaic virus (ToMV) strain L (L-ORF6), interacts in vitro with eukaryotic elongation factor 1α, and mutation of the ORF6 gene of tobacco mosaic virus (TMV) strain U1 (U1-ORF6) reduces the pathogenicity in vivo of TMV, whereas expression of this gene from two other viruses, tobacco rattle virus (TRV) and potato virus X (PVX), increases their pathogenicity. In this work, the in vivo properties of the L-ORF6 and U1-ORF6 proteins were compared to identify sequences that direct the proteins to different subcellular locations and also influence virus pathogenicity. Site-specific mutations in the ORF6 protein were made, hybrid ORF6 proteins were created in which the N-terminal and C-terminal parts were derived from the two proteins, and different subregions of the protein were examined, using expression either from a recombinant TRV vector or as a yellow fluorescent protein fusion from a binary plasmid in Agrobacterium tumefaciens. L-ORF6 caused mild necrotic symptoms in Nicotiana benthamiana when expressed from TRV, whereas U1-ORF6 caused severe symptoms including death of the plant apex. The difference in symptoms was associated with the C-terminal region of L-ORF6, which directed the protein to the endoplasmic reticulum (ER), whereas U1-ORF6 was directed initially to the nucleolus and later to the mitochondria. Positively charged residues at the N terminus allowed nucleolar entry of both U1-ORF6 and L-ORF6, but hydrophobic residues at the C terminus of L-ORF6 directed this protein to the ER.

  13. High-Affinity Binding of the Staphylococcal HarA Protein to Haptoglobin and Hemoglobin Involves a Domain with an Antiparallel Eight-Stranded β-Barrel Fold▿

    PubMed Central

    Dryla, Agnieszka; Hoffmann, Bernd; Gelbmann, Dieter; Giefing, Carmen; Hanner, Markus; Meinke, Andreas; Anderson, Annaliesa S.; Koppensteiner, Walter; Konrat, Robert; von Gabain, Alexander; Nagy, Eszter

    2007-01-01

    Iron scavenging from the host is essential for the growth of pathogenic bacteria. In this study, we further characterized two staphylococcal cell wall proteins previously shown to bind hemoproteins. HarA and IsdB harbor homologous ligand binding domains, the so called NEAT domain (for “near transporter”) present in several surface proteins of gram-positive pathogens. Surface plasmon resonance measurements using glutathione S-transferase (GST)-tagged HarAD1, one of the ligand binding domains of HarA, and GST-tagged full-length IsdB proteins confirmed high-affinity binding to hemoglobin and haptoglobin-hemoglobin complexes with equilibrium dissociation constants (KD) of 5 to 50 nM. Haptoglobin binding could be detected only with HarA and was in the low micromolar range. In order to determine the fold of this evolutionarily conserved ligand binding domain, the untagged HarAD1 protein was subjected to nuclear magnetic resonance spectroscopy, which revealed an eight-stranded, purely antiparallel β-barrel with the strand order (-β1↓-β2↑-β3↓-β6↑-β5↓-β4↑-β7↓-β8↑), forming two Greek key motifs. Based on structural-homology searches, the topology of the HarAD1 domain resembles that of the immunoglobulin (Ig) fold family, whose members are involved in protein-protein interactions, but with distinct structural features. Therefore, we consider that the HarAD1/NEAT domain fold is a novel variant of the Ig fold that has not yet been observed in other proteins. PMID:17041047

  14. Classical Swine Fever Virus p7 protein is a viroporin involved in virulence in swine

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The non-structural protein p7 of Classical Swine Fever Virus (CSFV) is a hydrophobic polypeptide with an apparent molecular mass of 7 kDa. The protein contains two hydrophobic stretches of amino acids interrupted by a short charged segment that are predicted to form transmembrane helices and a cytos...

  15. Involvement of C-Terminal Histidines in Soybean PM1 Protein Oligomerization and Cu2+ Binding.

    PubMed

    Liu, Guobao; Liu, Ke; Gao, Yang; Zheng, Yizhi

    2017-04-06

    Late embryogenesis abundant (LEA) proteins are widely distributed among plant species, where they contribute to abiotic stress tolerance. LEA proteins can be classified into seven groups according to conserved sequence motifs. The PM1 protein from soybean, which belongs to the Pfam LEA_1 group, has been shown previously to be at least partially natively unfolded, to bind metal ions and potentially to stabilize proteins and membranes. Here, we investigated the role of the PM1 C-terminal domain and in particular the multiple histidine residues in this half of the protein. We constructed recombinant plasmids expressing full-length PM1 and two truncated forms, PM1-N and PM1-C, which represent the N- and C-terminal halves of the protein, respectively. Immunoblotting and cross-linking experiments showed that full-length PM1 forms oligomers and high molecular weight (HMW) complexes in vitro and in vivo, while PM1-C, but not PM1-N, also formed oligomers and HMW complexes in vitro. When the histidine residues in PM1 and PM1-C were chemically modified, oligomerization was abolished, suggesting that histidines play a key role in this process. Furthermore, we demonstrated that high Cu2+ concentrations promote oligomerization and induce PM1 and PM1-C to form HMW complexes. Therefore, we speculate that PM1 proteins not only maintain ion homeostasis in the cytoplasm, but also potentially stabilize and protect other proteins during abiotic stress by forming a large, oligomeric molecular shield around biological targets.

  16. Quantitative Proteomic Analysis of Differentially Expressed Protein Profiles Involved in Pancreatic Ductal Adenocarcinoma

    PubMed Central

    Kuo, Kung-Kai; Kuo, Chao-Jen; Chiu, Chiang-Yen; Liang, Shih-Shin; Huang, Chun-Hao; Chi, Shu-Wen; Tsai, Kun-Bow; Chen, Chiao-Yun; Hsi, Edward; Cheng, Kuang-Hung; Chiou, Shyh-Horng

    2016-01-01

    Objectives The aim of this study was to identify differentially expressed proteins among various stages of pancreatic ductal adenocarcinoma (PDAC) by shotgun proteomics using nano-liquid chromatography coupled tandem mass spectrometry and stable isotope dimethyl labeling. Methods Differentially expressed proteins were identified and compared based on the mass spectral differences of their isotope-labeled peptide fragments generated from protease digestion. Results Our quantitative proteomic analysis of the differentially expressed proteins with stable isotope (deuterium/hydrogen ratio, ≥2) identified a total of 353 proteins, with at least 5 protein biomarker proteins that were significantly differentially expressed between cancer and normal mice by at least a 2-fold alteration. These 5 protein biomarker candidates include α-enolase, α-catenin, 14-3-3 β, VDAC1, and calmodulin with high confidence levels. The expression levels were also found to be in agreement with those examined by Western blot and histochemical staining. Conclusions The systematic decrease or increase of these identified marker proteins may potentially reflect the morphological aberrations and diseased stages of pancreas carcinoma throughout progressive developments leading to PDAC. The results would form a firm foundation for future work concerning validation and clinical translation of some identified biomarkers into targeted diagnosis and therapy for various stages of PDAC. PMID:26262590

  17. Identification of candidate effector proteins potentially involved in Fusarium graminearum-wheat interactions

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Pathogen-derived small secreted cysteine-rich proteins (SSCPs) are known to be a common source of fungal effectors that trigger resistance or susceptibility in specific host plants. This group of proteins has not been well studied in Fusarium graminearum, the primary cause of Fusarium head blight (...

  18. Different glycosyltransferases are involved in lipid glycosylation and protein N-glycosylation in the halophilic archaeon Haloferax volcanii.

    PubMed

    Naparstek, Shai; Vinagradov, Evguenii; Eichler, Jerry

    2010-07-01

    Both the lipid and the protein components of biological membranes can be modified by the covalent addition of polysaccharides. Whereas eukaryal and bacterial pathways of lipid and protein glycosylation are relatively well defined, considerably less is known of the parallel processes in Archaea. Recent efforts have identified glycosyltransferases involved in N-glycosylation of the surface-layer glycoprotein of the halophilic archaeon Haloferax volcanii. In the present study, the involvement of these same glycosyltransferases in the biosynthesis of Hfx. volcanii glycolipids was considered by performing nuclear magnetic resonance analysis of the glycolipid fraction of Hfx. volcanii cells deleted of genes encoding those glycosyltransferases, as well as the oligosaccharyltransferase, AglB. The results reveal that different glycosyltransferases are involved in the biosynthesis of N-linked glycoproteins and glycolipids in Archaea.

  19. Comparison of Algorithms for Prediction of Protein Structural Features from Evolutionary Data.

    PubMed

    Bywater, Robert P

    2016-01-01

    Proteins have many functions and predicting these is still one of the major challenges in theoretical biophysics and bioinformatics. Foremost amongst these functions is the need to fold correctly thereby allowing the other genetically dictated tasks that the protein has to carry out to proceed efficiently. In this work, some earlier algorithms for predicting protein domain folds are revisited and they are compared with more recently developed methods. In dealing with intractable problems such as fold prediction, when different algorithms show convergence onto the same result there is every reason to take all algorithms into account such that a consensus result can be arrived at. In this work it is shown that the application of different algorithms in protein structure prediction leads to results that do not converge as such but rather they collude in a striking and useful way that has never been considered before.

  20. Leucine alleviates dexamethasone-induced suppression of muscle protein synthesis via synergy involvement of mTOR and AMPK pathways

    PubMed Central

    Wang, Xiao J.; Yang, Xin; Wang, Ru X.; Jiao, Hong C.; Zhao, Jing P.; Song, Zhi G.; Lin, Hai

    2016-01-01

    Glucocorticoids (GCs) are negative muscle protein regulators that contribute to the whole-body catabolic state during stress. Mammalian target of rapamycin (mTOR)-signalling pathway, which acts as a central regulator of protein metabolism, can be activated by branched-chain amino acids (BCAA). In the present study, the effect of leucine on the suppression of protein synthesis induced by GCs and the pathway involved were investigated. In vitro experiments were conducted using cultured C2C12 myoblasts to study the effect of GCs on protein synthesis, and the involvement of mTOR pathway was investigated as well. After exposure to dexamethasone (DEX, 100 μmol/l) for 24 h, protein synthesis in muscle cells was significantly suppressed (P<0.05), the phosphorylations of mTOR, ribosomal protein S6 protein kinase 1 (p70s6k1) and eukaryotic initiation factor 4E binding protein 1 (4EBP1) were significantly reduced (P<0.05). Leucine supplementation (5 mmol/l, 10 mmol/l and 15 mmol/l) for 1 h alleviated the suppression of protein synthesis induced by DEX (P<0.05) and was accompanied with the increased phosphorylation of mTOR and decreased phosphorylation of AMPK (P<0.05). Branched-chain amino transferase 2 (BCAT2) mRNA level was not influenced by DEX (P>0.05) but was increased by leucine supplementation at a dose of 5 mmol/l (P<0.05). PMID:27129299

  1. The involvement of FANCM, FANCI, and checkpoint proteins in the interstrand DNA crosslink repair pathway is conserved in C. elegans.

    PubMed

    Lee, Kyong Yun; Chung, Kee Yang; Koo, Hyeon-Sook

    2010-04-04

    Fanconi anemia (FA) patients are specifically defective in the repair of interstrand DNA crosslinks (ICLs), a complex process involving at least 13 FA proteins and other repair/checkpoint proteins. Of the 13 FA proteins, FANCD1/BRCA2, FANCD2, and FANCJ were previously found to be functionally conserved in C. elegans. We have also identified C. elegans homologs of FANCM and FANCI, and determined their epistatic relationships with homologs of FANCD2, checkpoint proteins, and RAD51 upon DNA crosslinking. The counterparts of FANCM, FANCI, and three checkpoint proteins (RPA, ATR and CHK1) are required for focus formation and ubiquitination associated with FANCD2 in C. elegans. However, C. elegans FANCM affects neither RPA focus formation nor CHK1 phosphorylation induced by ICLs, unlike the reported role of human FANCM, which influences ATR-CHK1 signaling at stalled replication forks. Although focus formation by both FANCD2 and RAD51 requires ATR-CHK1 signaling, FANCD2 and RAD51 acted independently in the formation of their respective foci. Thus, the FANCD2 activation pathway involving FANCM, FANCI, and the checkpoint proteins is conserved in C. elegans but with distinct differences.

  2. Mapping the H+ (V)-ATPase interactome: identification of proteins involved in trafficking, folding, assembly and phosphorylation

    PubMed Central

    Merkulova, Maria; Păunescu, Teodor G.; Azroyan, Anie; Marshansky, Vladimir; Breton, Sylvie; Brown, Dennis

    2015-01-01

    V-ATPases (H+ ATPases) are multisubunit, ATP-dependent proton pumps that regulate pH homeostasis in virtually all eukaryotes. They are involved in key cell biological processes including vesicle trafficking, endosomal pH sensing, membrane fusion and intracellular signaling. They also have critical systemic roles in renal acid excretion and blood pH balance, male fertility, bone remodeling, synaptic transmission, olfaction and hearing. Furthermore, V-ATPase dysfunction either results in or aggravates various other diseases, but little is known about the complex protein interactions that regulate these varied V-ATPase functions. Therefore, we performed a proteomic analysis to identify V-ATPase associated proteins and construct a V-ATPase interactome. Our analysis using kidney tissue revealed V-ATPase-associated protein clusters involved in protein quality control, complex assembly and intracellular trafficking. ARHGEF7, DMXL1, EZR, NCOA7, OXR1, RPS6KA3, SNX27 and 9 subunits of the chaperonin containing TCP1 complex (CCT) were found to interact with V-ATPase for the first time in this study. Knockdown of two interacting proteins, DMXL1 and WDR7, inhibited V-ATPase-mediated intracellular vesicle acidification in a kidney cell line, providing validation for the utility of our interactome as a screen for functionally important novel V-ATPase-regulating proteins. Our data, therefore, provide new insights and directions for the analysis of V-ATPase cell biology and (patho)physiology. PMID:26442671

  3. The glue protein of ribbed mussels (Geukensia demissa): a natural adhesive with some features of collagen.

    PubMed

    Waite, J H; Hansen, D C; Little, K T

    1989-01-01

    The Atlantic ribbed mussel Geukensia (Modiolus) demissa attaches itself to the roots of cord grass and other hard objects in tidal salt marshes by spinning adhesive byssal threads. The precursor of a protein apparently present in the adhesive plaques of the threads was isolated in quantity from the foot of the mussel. The protein has an apparent molecular weight of 130,000, a pI of 8.1, and contains a high proportion of Gly, Glu/Gln, Lys and 3,4-dihydroxyphenyl-L-alanine (DOPA). Sequence of tryptic peptides suggests a pattern of repeated motifs, such as: Gly--DOPA--Lys, and X--Gly--DOPA--Y--Z--Gly--DOPA/Tyr--Lys, where X is Thr or Ala in octapeptides and Gln--Thr in nonapeptides. Y is variable, but more often than not hydrophobic; and Z is frequently Pro or 4-trans-hydroxyproline (Hyp). The presence of Pro--Gly and Hyp--Gly sequences of delta-hydroxylysine in the protein is reminiscent of typical collagens; however, the protein is not labile to clostridial collagenase, nor does collagen cross-react with antibodies raised against the mussel protein. Unlike typical collagens, Gly probably occurs only at every 4th or 5th residue in this unusual mussel protein.

  4. Functional dissection of protein domains involved in the immunomodulatory properties of PE_PGRS33 of Mycobacterium tuberculosis.

    PubMed

    Zumbo, Antonella; Palucci, Ivana; Cascioferro, Alessandro; Sali, Michela; Ventura, Marcello; D'Alfonso, Pamela; Iantomasi, Raffaella; Di Sante, Gabriele; Ria, Francesco; Sanguinetti, Maurizio; Fadda, Giovanni; Manganelli, Riccardo; Delogu, Giovanni

    2013-12-01

    PE_PGRSs are a large family of proteins identified in Mycobacterium tuberculosis complex and in few other pathogenic mycobacteria. The PE domain of PE_PGRS33 mediates localization of the protein on the mycobacterial cell surface, where the PGRS domain is available to interact with host components. In this study, PE_PGRS33 and its functional deletion mutants were expressed in M. smegmatis, and in vitro and in vivo assays were used to dissect the protein domains involved in the immunomodulatory properties of the protein. We demonstrate that PE_PGRS33-mediated secretion of TNF-α by macrophages occurs by extracellular interaction with TLR2. Our results also show that while the PGRS domain of the protein is required for triggering TNF-α secretion, mutation in the PE domain affects the pro-inflammatory properties of the protein. These results indicate that PE_PGRS33 is a protein with immunomodulatory activity and that protein stability and localization on the mycobacterial surface can affect these properties.

  5. STP1, a gene involved in pre-tRNA processing, encodes a nuclear protein containing zinc finger motifs.

    PubMed Central

    Wang, S S; Stanford, D R; Silvers, C D; Hopper, A K

    1992-01-01

    STP1 is an unessential yeast gene involved in the removal of intervening sequences from some, but not all, families of intervening sequence-containing pre-tRNAs. Previously, we proposed that STP1 might encode a product that generates pre-tRNA conformations efficiently recognized by tRNA-splicing endonuclease. To test the predictions of this model, we have undertaken a molecular analysis of the STP1 gene and its products. The STP1 locus is located on chromosome IV close to at least two other genes involved in RNA splicing: PRP3 and SPP41. The STP1 open reading frame (ORF) could encode a peptide of 64,827 Da; however, inspection of putative transcriptional and translational regulatory signals and mapping of the 5' ends of mRNA provide evidence that translation of the STP1 ORF usually initiates at a second AUG to generate a protein of 58,081 Da. The STP1 ORF contains three putative zinc fingers. The first of these closely resembles both the DNA transcription factor consensus and the Xenopus laevis p43 RNA-binding protein consensus. The third motif more closely resembles the fingers found in spliceosomal proteins. Employing antisera to the endogenous STP1 protein and to STP1-LacZ fusion proteins, we show that the STP1 protein is localized to nuclei. The presence of zinc finger motifs and the nuclear location of the STP1 protein support the model that this gene product is involved directly in pre-tRNA splicing. Images PMID:1588961

  6. Prediction of subcellular location of apoptosis proteins combining tri-gram encoding based on PSSM and recursive feature elimination.

    PubMed

    Liu, Taigang; Tao, Peiying; Li, Xiaowei; Qin, Yufang; Wang, Chunhua

    2015-02-07

    Knowledge of apoptosis proteins plays an important role in understanding the mechanism of programmed cell death. Obtaining information on subcellular location of apoptosis proteins is very helpful to reveal the apoptosis mechanism and understand the function of apoptosis proteins. Because of the cost in time and labor associated with large-scale wet-bench experiments, computational prediction of apoptosis proteins subcellular location becomes very important and many computational tools have been developed in the recent decades. Existing methods differ in the protein sequence representation techniques and classification algorithms adopted. In this study, we firstly introduce a sequence encoding scheme based on tri-grams computed directly from position-specific score matrices, which incorporates evolution information represented in the PSI-BLAST profile and sequence-order information. Then SVM-RFE algorithm is applied for feature selection and reduced vectors are input to a support vector machine classifier to predict subcellular location of apoptosis proteins. Jackknife tests on three widely used datasets show that our method provides the state-of-the-art performance in comparison with other existing methods.

  7. Neutrophil extracellular traps contain calprotectin, a cytosolic protein complex involved in host defense against Candida albicans.

    PubMed

    Urban, Constantin F; Ermert, David; Schmid, Monika; Abu-Abed, Ulrike; Goosmann, Christian; Nacken, Wolfgang; Brinkmann, Volker; Jungblut, Peter R; Zychlinsky, Arturo

    2009-10-01

    Neutrophils are the first line of defense at the site of an infection. They encounter and kill microbes intracellularly upon phagocytosis or extracellularly by degranulation of antimicrobial proteins and the release of Neutrophil Extracellular Traps (NETs). NETs were shown to ensnare and kill microbes. However, their complete protein composition and the antimicrobial mechanism are not well understood. Using a proteomic approach, we identified 24 NET-associated proteins. Quantitative analysis of these proteins and high resolution electron microscopy showed that NETs consist of modified nucleosomes and a stringent selection of other proteins. In contrast to previous results, we found several NET proteins that are cytoplasmic in unstimulated neutrophils. We demonstrated that of those proteins, the antimicrobial heterodimer calprotectin is released in NETs as the major antifungal component. Absence of calprotectin in NETs resulted in complete loss of antifungal activity in vitro. Analysis of three different Candida albicans in vivo infection models indicated that NET formation is a hitherto unrecognized route of calprotectin release. By comparing wild-type and calprotectin-deficient animals we found that calprotectin is crucial for the clearance of infection. Taken together, the present investigations confirmed the antifungal activity of calprotectin in vitro and, moreover, demonstrated that it contributes to effective host defense against C. albicans in vivo. We showed for the first time that a proportion of calprotectin is bound to NETs in vitro and in vivo.

  8. A Universal Stress Protein Involved in Oxidative Stress Is a Phosphorylation Target for Protein Kinase CIPK6.

    PubMed

    Gutiérrez-Beltrán, Emilio; Personat, José María; de la Torre, Fernando; Del Pozo, Olga

    2017-01-01

    Calcineurin B-like interacting protein kinases (CIPKs) decode calcium signals upon interaction with the calcium sensors calcineurin B like proteins into phosphorylation events that result into adaptation to environmental stresses. Few phosphorylation targets of CIPKs are known and therefore the molecular mechanisms underlying their downstream output responses are not fully understood. Tomato (Solanum lycopersicum) Cipk6 regulates immune and susceptible Programmed cell death in immunity transforming Ca(2+) signals into reactive oxygen species (ROS) signaling. To investigate SlCipk6-induced molecular mechanisms and identify putative substrates, a yeast two-hybrid approach was carried on and a protein was identified that contained a Universal stress protein (Usp) domain present in bacteria, protozoa and plants, which we named "SlRd2". SlRd2 was an ATP-binding protein that formed homodimers in planta. SlCipk6 and SlRd2 interacted using coimmunoprecipitation and bimolecular fluorescence complementation (BiFC) assays in Nicotiana benthamiana leaves and the complex localized in the cytosol. SlCipk6 phosphorylated SlRd2 in vitro, thus defining, to our knowledge, a novel target for CIPKs. Heterologous SlRd2 overexpression in yeast conferred resistance to highly toxic LiCl, whereas SlRd2 expression in Escherichia coli UspA mutant restored bacterial viability in response to H2O2 treatment. Finally, transient expression of SlCipk6 in transgenic N benthamiana SlRd2 overexpressors resulted in reduced ROS accumulation as compared to wild-type plants. Taken together, our results establish that SlRd2, a tomato UspA, is, to our knowledge, a novel interactor and phosphorylation target of a member of the CIPK family, SlCipk6, and functionally regulates SlCipk6-mediated ROS generation.

  9. The Arabidopsis Tellurite resistance C protein together with ALB3 is involved in photosystem II protein synthesis.

    PubMed

    Schneider, Anja; Steinberger, Iris; Strissel, Henning; Kunz, Hans-Henning; Manavski, Nikolay; Meurer, Jörg; Burkhard, Gabi; Jarzombski, Sabine; Schünemann, Danja; Geimer, Stefan; Flügge, Ulf-Ingo; Leister, Dario

    2014-04-01

    Assembly of photosystem II (PSII) occurs sequentially and requires several auxiliary proteins, such as ALB3 (ALBINO3). Here, we describe the role of the Arabidopsis thaliana thylakoid membrane protein Tellurite resistance C (AtTerC) in this process. Knockout of AtTerC was previously shown to be seedling-lethal. This phenotype was rescued by expressing TerC fused C-terminally to GFP in the terc-1 background, and the resulting terc-1TerC- GFP line and an artificial miRNA-based knockdown allele (amiR-TerC) were used to analyze the TerC function. The alterations in chlorophyll fluorescence and thylakoid ultrastructure observed in amiR-TerC plants and terc-1TerC- GFP were attributed to defects in PSII. We show that this phenotype resulted from a reduction in the rate of de novo synthesis of PSII core proteins, but later steps in PSII biogenesis appeared to be less affected. Yeast two-hybrid assays showed that TerC interacts with PSII proteins. In particular, its interaction with the PSII assembly factor ALB3 has been demonstrated by co-immunoprecipitation. ALB3 is thought to assist in incorporation of CP43 into PSII via interaction with Low PSII Accumulation2 (LPA2) Low PSII Accumulation3 (LPA3). Homozygous lpa2 mutants expressing amiR-TerC displayed markedly exacerbated phenotypes, leading to seedling lethality, indicating an additive effect. We propose a model in which TerC, together with ALB3, facilitates de novo synthesis of thylakoid membrane proteins, for instance CP43, at the membrane insertion step.

  10. Identification of surface proteins involved in the adhesion of a probiotic Bacillus cereus strain to mucin and fibronectin.

    PubMed

    Sánchez, B; Arias, S; Chaignepain, S; Denayrolles, M; Schmitter, J M; Bressollier, P; Urdaci, M C

    2009-05-01

    Several Bacillus strains isolated from commercial probiotic preparations were identified at the species level, and their adhesion capabilities to three different model intestinal surfaces (mucin, Matrigel and Caco-2 cells) were assessed. In general, adhesion of spores was higher than that of vegetative cells to the three matrices, and overall strain Bacillus cereus(CH) displayed the best adhesion. Different biochemical treatments revealed that surface proteins of B. cereus(CH) were involved in the adhesion properties of the strain. Surface-associated proteins from vegetative cells and spores of B. cereus(CH) were extracted and identified, and some proteins such as S-layer components, flagellin and cell-bound proteases were found to bind to mucin or fibronectin. These facts suggest that those proteins might play important roles in the interaction of this probiotic Bacillus strain within the human gastrointestinal tract.

  11. HotPatch: A Statistical Approach to Finding Biologically Relevant Features on Protein Surfaces

    PubMed Central

    Pettit, Frank K.; Bare, Emiko; Tsai, Albert; Bowie, James U.

    2007-01-01

    We describe a fully automated algorithm for finding functional sites on protein structures. Our method finds surface patches of unusual physicochemical properties on protein structures, and estimates the patches’ probability of overlapping functional sites. Other methods for predicting the locations of specific types of functional sites exist, but in previous analyses, it has been difficult to compare methods when they are applied to different types of sites. Thus, we introduce a new statistical framework that enables rigorous comparisons of the usefulness of different physicochemical properties for predicting virtually any kind of functional site. The program’s statistical models were trained for 11 individual properties (electrostatics, concavity, hydrophobicity, etc.) and for 15 neural network combination properties, all optimized and tested on 15 diverse protein functions. To simulate what to expect if the program were run on proteins of unknown function, as might arise from structural genomics, we tested it on 618 proteins of diverse mixed functions. In the higher-scoring top half of all predictions, a functional residue could typically be found within the first 1.7 residues chosen at random. The program may or may not use partial information about the protein’s function type as an input, depending on which statistical model the user chooses to employ. If function type is used as an additional constraint, prediction accuracy usually increases, and is particularly good for enzymes, DNA-interacting sites, and oligomeric interfaces. The program can be accessed online at http://hotpatch.mbi.ucla.edu. PMID:17451744

  12. The LspB protein is involved in the secretion of the LspA1 and LspA2 proteins by Haemophilus ducreyi.

    PubMed

    Ward, Christine K; Mock, Jason R; Hansen, Eric J

    2004-04-01

    The LspA1 and LspA2 proteins of Haemophilus ducreyi 35000 are two very large macromolecules that can be detected in concentrated culture supernatant fluid. Both of these proteins exhibit homology with the N-terminal region of the Bordetella pertussis filamentous hemagglutinin (FHA), which is involved in secretion of the latter macromolecule. The lspA2 open reading frame is flanked upstream by a gene, lspB, that encodes a predicted protein with homology to the B. pertussis FhaC outer membrane protein that is involved in secretion of FHA across the outer membrane. The H. ducreyi lspB gene encodes a protein with a predicted molecular mass of 66,573 Da. Reverse transcription-PCR analysis suggested that the lspB gene was transcribed together with the lspA2 gene on a single mRNA transcript. Polyclonal H. ducreyi LspB antiserum reacted with a 64-kDa antigen present in the Sarkosyl-insoluble cell envelope fraction of H. ducreyi 35000, which indicated that the LspB protein is likely an outer membrane protein. Concentrated culture supernatant fluids from H. ducreyi lspB and lspA1 lspB mutants did not contain detectable LspA1 and detectable LspA2, respectively. However, complementation of the lspB mutant with the wild-type lspB gene on a plasmid restored LspB protein expression and resulted in release of detectable amounts of the LspA1 protein into culture supernatant fluid. When evaluated in the temperature-dependent rabbit model of infection, the lspB mutant was attenuated in the ability to cause lesions and was never recovered in a viable form from lesions. These results indicated that the H. ducreyi LspB protein is involved in secretion of the LspA1 and LspA2 proteins across the outer membrane.

  13. Dietary fish protein alters blood lipid concentrations and hepatic genes involved in cholesterol homeostasis in the rat model.

    PubMed

    Shukla, Anjali; Bettzieche, Anja; Hirche, Frank; Brandsch, Corinna; Stangl, Gabriele I; Eder, Klaus

    2006-10-01

    It is known that various dietary plant proteins are capable of influencing the lipid metabolism of human subjects and animals when compared with casein. Less, however, is known about the effects of fish protein on the cholesterol and triacylglycerol metabolism. Therefore, two experiments were conducted in which rats were fed diets containing 200 g of either fish protein, prepared from Alaska pollack fillets, or casein, which served as control, per kilogram, over 20 and 22 d, respectively. As parameters of lipid metabolism, the concentrations of cholesterol and triacylglycerols in the plasma and liver, the faecal excretion of bile acids and the hepatic expression of genes encoding proteins involved in lipid homeostasis were determined. In both experiments, rats fed fish protein had higher concentrations of cholesteryl esters in the liver, a lower concentration of cholesterol in the HDL fraction (rho > 1.063 kg/l) and lower plasma triacylglycerol concentrations than rats fed casein (P < 0.05). The gene expression analysis performed in experiment 2 showed that rats fed fish protein had higher relative mRNA concentrations of sterol regulatory element-binding protein (SREBP)-2, 3-hydroxy-3-methylglutaryl coenzyme A reductase, LDL receptor, apo AI, scavenger receptor B1 and lecithin-cholesterol-acyltransferase in their liver than did rats fed casein (P < 0.05). The faecal excretion of bile acids and the mRNA concentrations of cholesterol 7alpha-hydroxylase, SREBP-1c and corresponding target genes were not altered. These findings show that fish protein had multiple effects on plasma and liver lipids that were at least in part caused by an altered expression of the hepatic genes involved in lipid homeostasis.

  14. Proteomics, metabolomics, and protein interactomics in the characterization of the molecular features of major depressive disorder.

    PubMed

    Martins-de-Souza, Daniel

    2014-03-01

    Omics technologies emerged as complementary strategies to genomics in the attempt to understand human illnesses. In general, proteomics technologies emerged earlier than those of metabolomics for major depressive disorder (MDD) research, but both are driven by the identification of proteins and/or metabolites that can delineate a comprehensive characterization of MDD's molecular mechanisms, as well as lead to the identification of biomarker candidates of all types-prognosis, diagnosis, treatment, and patient stratification. Also, one can explore protein and metabolite interactomes in order to pinpoint additional molecules associated with the disease that had not been picked up initially. Here, results and methodological aspects of MDD research using proteomics, metabolomics, and protein interactomics are reviewed, focusing on human samples.

  15. The Shwachman-Bodian-Diamond Syndrome Protein Family Is Involved in RNA Metabolism

    SciTech Connect

    Savchenko, A; Krogan, Nevan; Cort, John R.; Evdokimova, Elena; Lew, Jocelyne M.; Yee, Adelinda; Sanchez-Pulido, Luis; Andrade, Miguel; Bochkarev, Alexey; Watson, James D.; Kennedy, Michael A.; Greenblatt, Jack; Hughes, Timothy; Arrowsmith, Cheryl H.; Rommens, Johanna M.; Edwards, Aled M.

    2005-05-13

    A combination of structural, biochemical, and genetic studies in model organisms was used to infer a cellular role for the human protein (SBDS) responsible for Shwachman-Bodian-Diamond syndrome. The crystal structure of the SBDS homologue in Archaeoglobus fulgidus, AF0491, revealed a three domain protein. The N-terminal domain, which harbors the majority of disease-linked mutations, has a novel three-dimensional fold.

  16. Bioinformatics and functional analyses of coronavirus nonstructural proteins involved in the formation of replicative organelles.

    PubMed

    Neuman, Benjamin W

    2016-11-01

    Replication of eukaryotic positive-stranded RNA viruses is usually linked to the presence of membrane-associated replicative organelles. The purpose of this review is to discuss the function of proteins responsible for formation of the coronavirus replicative organelle. This will be done by identifying domains that are conserved across the order Nidovirales, and by summarizing what is known about function and structure at the level of protein domains.

  17. iTRAQ-Based Quantitative Proteomics Identifies Potential Regulatory Proteins Involved in Chicken Eggshell Brownness

    PubMed Central

    Wu, Guiqin; Shi, Fengying; Liu, Aiqiao; Yang, Ning

    2016-01-01

    Brown eggs are popular in many countries and consumers regard eggshell brownness as an important indicator of egg quality. However, the potential regulatory proteins and detailed molecular mechanisms regulating eggshell brownness have yet to be clearly defined. In the present study, we performed quantitative proteomics analysis with iTRAQ technology in the shell gland epithelium of hens laying dark and light brown eggs to investigate the candidate proteins and molecular mechanisms underlying variation in chicken eggshell brownness. The results indicated 147 differentially expressed proteins between these two groups, among which 65 and 82 proteins were significantly up-regulated in the light and dark groups, respectively. Functional analysis indicated that in the light group, the down-regulated iron-sulfur cluster assembly protein (Iba57) would decrease the synthesis of protoporphyrin IX; furthermore, the up-regulated protein solute carrier family 25 (mitochondrial carrier; adenine nucleotide translocator), member 5 (SLC25A5) and down-regulated translocator protein (TSPO) would lead to increased amounts of protoporphyrin IX transported into the mitochondria matrix to form heme with iron, which is supplied by ovotransferrin protein (TF). In other words, chickens from the light group produce less protoporphyrin IX, which is mainly used for heme synthesis. Therefore, the exported protoporphyrin IX available for eggshell deposition and brownness is reduced in the light group. The current study provides valuable information to elucidate variation of chicken eggshell brownness, and demonstrates the feasibility and sensitivity of iTRAQ-based quantitative proteomics analysis in providing useful insights into the molecular mechanisms underlying brown eggshell pigmentation. PMID:28006025

  18. A conserved motif in S-layer proteins is involved in peptidoglycan binding in Thermus thermophilus.

    PubMed Central

    Olabarría, G; Carrascosa, J L; de Pedro, M A; Berenguer, J

    1996-01-01

    There is experimental evidence to suggest that the 100-kDa S-layer protein from Thermus thermophilus HB8 binds to the peptidoglycan cell wall. This property could be related to the presence of a region (SLH) of homology with other S-layer proteins and extracellular enzymes (A. Lupas, H. Engelhardt, J. Peters, U. Santarius, S. Volker, and W. Baumeister, J. Bacteriol. 176:1224-1233, 1994). By using specific monoclonal antibodies, we show that similar regions are present in different members of the Deinococcus-Thermus phylogenetic group. To analyze the role that the SLH domain plays in vivo and in vitro in T. thermophilus, we have obtained a mutant form (slpA.X) of the S-layer gene (slpA) in which the SLH domain was deleted. The slpA.X gene was inserted into the chromosome of the thermophile by gene replacement, resulting in a mutant which expressed a major membrane protein with the size expected from the construction (90 kDa). This protein was identified as the product of slpA.X by its differential reaction with monoclonal antibodies. Mutants expressing the SlpA.X protein grow as groups of cells, surrounded by a common external envelope of trigonal symmetry that contains the SlpA.X protein as a main component, thus showing the inability of the SLH-defective protein to attach to the underlying material in vivo. In addition, averaged images of SlpA.X-rich fractions showed a regular arrangement, identical to that built up by the wild-type (SlpA) protein in the absence of peptidoglycan. Finally, we demonstrate by Western blotting (immunoblotting) the direct role of the SLH domain in the binding of the S-layer of T. thermophilus HB8 to the peptidoglycan layer. PMID:8759836

  19. Plasma membrane associated membranes (PAM) from Jurkat cells contain STIM1 protein is PAM involved in the capacitative calcium entry?

    PubMed

    Kozieł, Katarzyna; Lebiedzinska, Magdalena; Szabadkai, Gyorgy; Onopiuk, Marta; Brutkowski, Wojciech; Wierzbicka, Katarzyna; Wilczyński, Grzegorz; Pinton, Paolo; Duszyński, Jerzy; Zabłocki, Krzysztof; Wieckowski, Mariusz R

    2009-12-01

    A proper cooperation between the plasma membrane, the endoplasmic reticulum and the mitochondria seems to be essential for numerous cellular processes involved in Ca(2+) signalling and maintenance of Ca(2+) homeostasis. A presence of microsomal and mitochondrial proteins together with those characteristic for the plasma membrane in the fraction of the plasma membrane associated membranes (PAM) indicates a formation of stabile interactions between these three structures. We isolated the plasma membrane associated membranes from Jurkat cells and found its significant enrichment in the plasma membrane markers including plasma membrane Ca(2+)-ATPase, Na(+), K(+)-ATPase and CD3 as well as sarco/endoplasmic reticulum Ca(2+) ATPase as a marker of the endoplasmic reticulum membranes. In addition, two proteins involved in the store-operated Ca(2+) entry, Orai1 located in the plasma membrane and an endoplasmic reticulum protein STIM1 were found in this fraction. Furthermore, we observed a rearrangement of STIM1-containing protein complexes isolated from Jurkat cells undergoing stimulation by thapsigargin. We suggest that the inter-membrane compartment composed of the plasma membrane and the endoplasmic reticulum, and isolated as a stabile plasma membrane associated membranes fraction, might be involved in the store-operated Ca(2+) entry, and their formation and rebuilding have an important regulatory role in cellular Ca(2+) homeostasis.

  20. Immunological identification of candidate proteins involved in regulating active shape changes of outer hair cells.

    PubMed

    Knipper, M; Zimmermann, U; Köpschall, I; Rohbock, K; Jüngling, S; Zenner, H P

    1995-06-01

    By employing immunological methods, it has been demonstrated that myosin, myosin light chain (MLC) and myosin light chain kinase (MLCK) proteins in outer hair cells (OHC) are immunologically different from isoforms in platelets, smooth muscle and heart muscle, and are probably more related to isoforms found in red blood cells (RBC). Moreover, proteins related to band 3 protein (b3p) and protein 4.1 (p 4.1), ankyrin as well as fodrin and spectrin, but not glycophorin, have been identified in isolated OHCs. Both OHCs and RBC differ from other motile non-muscle cells in their lack of smooth muscle isoforms of actin, their common high levels of spectrin-, ankyrin- and band 3-like proteins, as well as the expression of the 80 kDa protein 4.1 isoform. The data support the notion that motility of OHC may be based upon regulation of the b3p/p 4.1/ankyrin complex, and thus may be reminiscent to the active shape changes in RBC.

  1. The pupylation machinery is involved in iron homeostasis by targeting the iron storage protein ferritin.

    PubMed

    Küberl, Andreas; Polen, Tino; Bott, Michael

    2016-04-26

    The balance of sufficient iron supply and avoidance of iron toxicity by iron homeostasis is a prerequisite for cellular metabolism and growth. Here we provide evidence that, in Actinobacteria, pupylation plays a crucial role in this process. Pupylation is a posttranslational modification in which the prokaryotic ubiquitin-like protein Pup is covalently attached to a lysine residue in target proteins, thus resembling ubiquitination in eukaryotes. Pupylated proteins are recognized and unfolded by a dedicated AAA+ ATPase (Mycobacterium proteasomal AAA+ ATPase; ATPase forming ring-shaped complexes). In Mycobacteria, degradation of pupylated proteins by the proteasome serves as a protection mechanism against several stress conditions. Other bacterial genera capable of pupylation such as Corynebacterium lack a proteasome, and the fate of pupylated proteins is unknown. We discovered that Corynebacterium glutamicum mutants lacking components of the pupylation machinery show a strong growth defect under iron limitation, which was caused by the absence of pupylation and unfolding of the iron storage protein ferritin. Genetic and biochemical data support a model in which the pupylation machinery is responsible for iron release from ferritin independent of degradation.

  2. Prion Protein and Shadoo Are Involved in Overlapping Embryonic Pathways and Trophoblastic Development

    PubMed Central

    Makhzami, Samira; Vilotte, Marthe; Jaffrezic, Florence; Halliez, Sophie; Bouet, Stéphan; Marthey, Sylvain; Khalifé, Manal; Kanellopoulos-Langevin, Colette; Béringue, Vincent; Le Provost, Fabienne; Laude, Hubert; Vilotte, Jean-Luc

    2012-01-01

    The potential requirement of either the Prion or Shadoo protein for early mouse embryogenesis was recently suggested. However, the current data did not allow to precise the developmental process that was affected in the absence of both proteins and that led to the observed early lethal phenotype. In the present study, using various Prnp transgenic mouse lines and lentiviral vectors expressing shRNAs that target the Shadoo-encoding mRNA, we further demonstrate the specific requirement of at least one of these two PrP-related proteins at early developmental stages. Histological analysis reveals developmental defect of the ectoplacental cone and important hemorrhage surrounding the Prnp-knockout-Sprn-knockdown E7.5 embryos. By restricting the RNA interference to the trophoblastic cell lineages, the observed lethal phenotype could be attributed to the sole role of these proteins in this trophectoderm-derived compartment. RNAseq analysis performed on early embryos of various Prnp and Sprn genotypes indicated that the simultaneous down-regulation of these two proteins affects cell-adhesion and inflammatory pathways as well as the expression of ectoplacental-specific genes. Overall, our data provide biological clues in favor of a crucial and complementary embryonic role of the prion protein family in Eutherians and emphasizes the need to further evaluate its implication in normal and pathological human placenta biology. PMID:22860039

  3. The pupylation machinery is involved in iron homeostasis by targeting the iron storage protein ferritin

    PubMed Central

    Küberl, Andreas; Polen, Tino; Bott, Michael

    2016-01-01

    The balance of sufficient iron supply and avoidance of iron toxicity by iron homeostasis is a prerequisite for cellular metabolism and growth. Here we provide evidence that, in Actinobacteria, pupylation plays a crucial role in this process. Pupylation is a posttranslational modification in which the prokaryotic ubiquitin-like protein Pup is covalently attached to a lysine residue in target proteins, thus resembling ubiquitination in eukaryotes. Pupylated proteins are recognized and unfolded by a dedicated AAA+ ATPase (Mycobacterium proteasomal AAA+ ATPase; ATPase forming ring-shaped complexes). In Mycobacteria, degradation of pupylated proteins by the proteasome serves as a protection mechanism against several stress conditions. Other bacterial genera capable of pupylation such as Corynebacterium lack a proteasome, and the fate of pupylated proteins is unknown. We discovered that Corynebacterium glutamicum mutants lacking components of the pupylation machinery show a strong growth defect under iron limitation, which was caused by the absence of pupylation and unfolding of the iron storage protein ferritin. Genetic and biochemical data support a model in which the pupylation machinery is responsible for iron release from ferritin independent of degradation. PMID:27078093

  4. Prediction of protein structure classes using hybrid space of multi-profile Bayes and bi-gram probability feature spaces.

    PubMed

    Hayat, Maqsood; Tahir, Muhammad; Khan, Sher Afzal

    2014-04-07

    Proteins are the executants of biological functions in living organisms. Comprehension of protein structure is a challenging problem in the era of proteomics, computational biology, and bioinformatics because of its pivotal role in protein folding patterns. Owing to the large exploration of protein sequences in protein databanks and intricacy of protein structures, experimental and theoretical methods are insufficient for prediction of protein structure classes. Therefore, it is highly desirable to develop an accurate, reliable, and high throughput computational model to predict protein structure classes correctly from polygenetic sequences. In this regard, we propose a promising model employing hybrid descriptor space in conjunction with optimized evidence-theoretic K-nearest neighbor algorithm. Hybrid space is the composition of two descriptor spaces including Multi-profile Bayes and bi-gram probability. In order to enhance the generalization power of the classifier, we have selected high discriminative descriptors from the hybrid space using particle swarm optimization, a well-known evolutionary feature selection technique. Performance evaluation of the proposed model is performed using the jackknife test on three low similarity benchmark datasets including 25PDB, 1189, and 640. The success rates of the proposed model are 87.0%, 86.6%, and 88.4%, respectively on the three benchmark datasets. The comparative analysis exhibits that our proposed model has yielded promising results compared to the existing methods in the literature. In addition, our proposed prediction system might be helpful in future research particularly in cases where the major focus of research is on low similarity datasets.

  5. Conserved hypothetical protein Rv1977 in Mycobacterium tuberculosis strains contains sequence polymorphisms and might be involved in ongoing immune evasion.

    PubMed

    Jiang, Yi; Liu, Haican; Wang, Xuezhi; Li, Guilian; Qiu, Yan; Dou, Xiangfeng; Wan, Kanglin

    2015-01-01

    Host immune pressure and associated parasite immune evasion are key features of host-pathogen co-evolution. A previous study showed that human T cell epitopes of Mycobacterium tuberculosis are evolutionarily hyperconserved and thus it was deduced that M. tuberculosis lacks antigenic variation and immune evasion. Here, we selected 151 clinical Mycobacterium tuberculosis isolates from China, amplified gene encoding Rv1977 and compared the sequences. The results showed that Rv1977, a conserved hypothetical protein, is not conserved in M. tuberculosis strains and there are polymorphisms existed in the protein. Some mutations, especially one frameshift mutation, occurred in the antigen Rv1977, which is uncommon in M.tb strains and may lead to the protein function altering. Mutations and deletion in the gene all affect one of three T cell epitopes and the changed T cell epitope contained more than one variable position, which may suggest ongoing immune evasion.

  6. The SeqFEATURE library of 3D functional site models: comparison to existing methods and applications to protein function annotation.

    PubMed

    Wu, Shirley; Liang, Mike P; Altman, Russ B

    2008-01-16

    Structural genomics efforts have led to increasing numbers of novel, uncharacterized protein structures with low sequence identity to known proteins, resulting in a growing need for structure-based function recognition tools. Our method, SeqFEATURE, robustly models protein functions described by sequence motifs using a structural representation. We built a library of models that shows good performance compared to other methods. In particular, SeqFEATURE demonstrates significant improvement over other methods when sequence and structural similarity are low.

  7. Identification and characterization of a Streptococcus equi ssp. zooepidemicus immunogenic GroEL protein involved in biofilm formation.

    PubMed

    Yi, Li; Wang, Yang; Ma, Zhe; Lin, Hui-Xing; Xu, Bin; Grenier, Daniel; Fan, Hong-Jie; Lu, Cheng-Ping

    2016-04-18

    Streptococcus equi ssp. zooepidemicus (S. equi spp. zooepidemicus) is an opportunistic pathogen that causes major economic losses in the swine industry in China and is also a threat for human health. Biofilm formation by this bacterium has been previously reported. In this study, we used an immunoproteomic approach to search for immunogenic proteins expressed by biofilm-grown S. equi spp. zooepidemicus. Seventeen immunoreactive proteins were found, of which nine common immunoreactive proteins were identified in planktonic and biofilm-grown bacteria. The immunogenicity and protective efficacy of the S. equi spp. zooepidemicus immunoreactive GroEL chaperone protein was further investigated in mice. The protein was expressed in vivo and elicited high antibody titers following S. equi spp. zooepidemicus infections of mice. An animal challenge experiment with S. equi spp. zooepidemicus showed that 75% of mice immunized with the GroEL protein were protected. Using in vitro biofilm inhibition assays, evidence was obtained that the chaperonin GroEL may represent a promising target for the prevention and treatment of persistent S. equi spp. zooepidemicus biofilm infections. In summary, our results suggest that the recombinant GroEL protein, which is involved in biofilm formation, may efficiently stimulate an immune response, which protects against S. equi spp. zooepidemicus infections. It may therefore be a candidate of interest to be included in vaccines against S. equi spp. zooepidemicus infections.

  8. Lung involvement in systemic sclerosis: role of high resolution computed tomography and its relationship with other pulmonary and clinico-serological features.

    PubMed

    Colaci, M; Sebastiani, M; Manfredi, A; Giuggioli, D; Cassone, G; Manzini, C U; Ghizzoni, C; Cerri, S; Ferri, C

    2014-01-01

    The study investigated the characteristic of interstitial lung disease in a large series of systemic sclerosis (SSc) patients by means of HRCT and the correlations between functional lung parameters, serological features and the extent of lung involvement evaluated by high-resolution computed tomography (HRCT). One hundred and seven SSc patients, consecutively investigated by means of HRCT, standard chest X-ray, and pulmonary function tests, were retrospectively evaluated. Chest radiogram and HRCT scores were strongly associated (Pearson’'s r=0.82, p < .0001); moreover, the first significantly correlated with spirometric parameters, even if weakly. Anti-Scl70 and anti-centromere antibodies were associated with higher (p=0.01) and lower HRCT score (p=0.0002), respectively. The extension of interstitial lung involvement in SSc evaluated with HRCT is directly proportional to functional lung parameters. HRCT, spirometry and DLco should be considered essential in the core-set of non-invasive diagnostic tools for the first-line assessment of scleroderma lung involvement.

  9. Two RND proteins involved in heavy metal efflux in Caulobacter crescentus belong to separate clusters within proteobacteria

    PubMed Central

    2013-01-01

    Background Heavy metal Resistance-Nodulation-Division (HME-RND) efflux systems help Gram-negative bacteria to keep the intracellular homeostasis under high metal concentrations. These proteins constitute the cytoplasmic membrane channel of the tripartite RND transport systems. Caulobacter crescentus NA1000 possess two HME-RND proteins, and the aim of this work was to determine their involvement in the response to cadmium, zinc, cobalt and nickel, and to analyze the phylogenetic distribution and characteristic signatures of orthologs of these two proteins. Results Expression assays of the czrCBA operon showed significant induction in the presence of cadmium and zinc, and moderate induction by cobalt and nickel. The nczCBA operon is highly induced in the presence of nickel and cobalt, moderately induced by zinc and not induced by cadmium. Analysis of the resistance phenotype of mutant strains showed that the ΔczrA strain is highly sensitive to cadmium, zinc and cobalt, but resistant to nickel. The ΔnczA strain and the double mutant strain showed reduced growth in the presence of all metals tested. Phylogenetic analysis of the C. crescentus HME-RND proteins showed that CzrA-like proteins, in contrast to those similar to NczA, are almost exclusively found in the Alphaproteobacteria group, and the characteristic protein signatures of each group were highlighted. Conclusions The czrCBA efflux system is involved mainly in response to cadmium and zinc with a secondary role in response to cobalt. The nczCBA efflux system is involved mainly in response to nickel and cobalt, with a secondary role in response to cadmium and zinc. CzrA belongs to the HME2 subfamily, which is almost exclusively found in the Alphaproteobacteria group, as shown by phylogenetic analysis. NczA belongs to the HME1 subfamily which is more widespread among diverse Proteobacteria groups. Each of these subfamilies present distinctive amino acid signatures. PMID:23578014

  10. Location of the serine residue involved in the linkage between the terminal protein and the DNA of phage phi 29.

    PubMed Central

    Hermoso, J M; Méndez, E; Soriano, F; Salas, M

    1985-01-01

    B. subtilis phage phi 29 has a terminal protein, p3, covalently linked to the 5' ends of the DNA through a phosphodiester bond between a serine residue and 5'-dAMP. This protein acts as a primer in DNA replication by forming an initiation complex with the 5'-terminal nucleotide dAMP. The amino acid sequence of the terminal protein, deduced from the nucleotide sequence of gene 3, showed the presence of 18 serine residues in a total of 266 amino acids. In this paper we have identified the serine involved in the linkage with the DNA as the residue 232, located close to the C-terminus of the molecule. This result was obtained by amino acid analysis of the peptide that remains linked to the DNA after proteinase K digestion of the terminal protein-phi 29 DNA complex and automated Edman degradation of the corresponding [125I]-labeled tryptic peptide. Prediction of the secondary structure of the terminal protein suggested that the serine residue involved in the linkage with the DNA is placed in a beta-turn, probably located on the external part of the molecule, as indicated by hydropathic values. Images PMID:3934646

  11. Microaggregate-associated protein involved in invasion of epithelial cells by Mycobacterium avium subsp. hominissuis

    PubMed Central

    Babrak, Lmar; Danelishvili, Lia; Rose, Sasha J; Bermudez, Luiz E

    2015-01-01

    The environmental opportunistic pathogen Mycobacterium avium subsp hominissuis (MAH), a member of the nontuberculous mycobacteria (NTM) cluster, causes respiratory as well as disseminated disease in patients such as those with chronic respiratory illnesses or AIDS. Currently, there is no effective method to prevent NTM respiratory infections. The formation of mycobacterial microaggregates comprises of phenotypic changes that lead to efficient adherence and invasion of the respiratory mucosa in vitro and in vivo. Microaggregate adhesion to the respiratory epithelium is mediated in part through the mycobacterial protein, MAV_3013 (MBP-1). Through DNA microarray analysis, the small hypothetical gene MAV_0831 (Microaggregate Invasion Protein-1, MIP-1) was identified as being upregulated during microaggregate formation. When MIP-1 was overexpressed in poorly-invasive Mycobacterium smegmatis, it provided the bacterium the ability to bind and enter epithelial cells. In addition, incubating microaggregates with recombinant MIP-1 protein enhanced the ability of microaggregates to invade HEp-2 cells, and exposure to anti-MIP-1 immune serum reduced the invasion of the host epithelium. Through protein-protein interaction assays, MIP-1 was found to bind to the host protein filamin A, a cytoskeletal actin-binding protein integral to the modulation of host cell shape and migration. As visualized by immunofluorescence, filamin A was able to co-localize with microaggregates and to a lesser extent planktonic bacteria. Invasion of HEp-2 cells by microaggregates and planktonic bacteria was also inhibited by the addition of anti-filamin A antibody suggesting that filamin A plays an important role during infection. In addition, at earlier time points binding and invasion assay results suggest that MBP-1 participates significantly during the first interactions with the host cell while MIP-1 becomes important once the bacteria adhere to the host epithelium. In summary, we have unveiled

  12. LEA proteins are involved in cyst desiccation resistance and other abiotic stresses in Azotobacter vinelandii.

    PubMed

    Rodriguez-Salazar, Julieta; Moreno, Soledad; Espín, Guadalupe

    2017-03-03

    Late embryogenesis abundant (LEA) proteins constitute a large protein family that is closely associated with resistance to abiotic stresses in multiple organisms and protect cells against drought and other stresses. Azotobacter vinelandii is a soil bacterium that forms desiccation-resistant cysts. This bacterium possesses two genes, here named lea1 and lea2, coding for avLEA1 and avLEA2 proteins, both containing 20-mer motifs characteristic of eukaryotic plant LEA proteins. In this study, we found that disruption of the lea1 gene caused a loss of the cysts' viability after 3 months of desiccation, whereas at 6 months, wild-type or lea2 mutant strain cysts remained viable. Vegetative cells of the lea1 mutant were more sensitive to osmotic stress; cysts developed by this mutant were also more sensitive to high temperatures than cysts or vegetative cells of the wild type or of the lea2 mutant. Expression of lea1 was induced several fold during encystment. In addition, the protective effects of these proteins were assessed in Escherichia coli cells. We found that E. coli cells overexpressing avLEA1 were more tolerant to salt stress than control cells; finally, in vitro analysis showed that avLEA1 protein was able to prevent the freeze thaw-induced inactivation of lactate dehydrogenase. In conclusion, avLEA1 is essential for the survival of A. vinelandii in dry conditions and for protection against hyper-osmolarity, two major stress factors that bacteria must cope with for survival in the environment. This is the first report on the role of bacterial LEA proteins on the resistance of cysts to desiccation.

  13. Antigenic features of prion proteins of sheep and of other mammalian species.

    PubMed

    Groschup, M H; Harmeyer, S; Pfaff, E

    1997-08-22

    Pathological prion protein (PrPSc) which is a conformational isoform of a host-encoded protein designated (PrPC) serves as a specific marker protein for the immunochemical diagnosis of transmissible spongiform encephalopathies (TSE). The generation of suitable antibodies to PrPSc therefore underlies the specificity and sensitivity of diagnostic assays. However, most antibodies reported to date are directed to a limited number of epitopes only. PrPC is a highly conserved cell membrane protein in all mammalian species studied to date. In an attempt to generate antibodies to further regions of PrP we raised antisera in rabbits and chicken against sixteen synthetic peptides which represent the complete aminoacid sequence of ovine PrP. By this approach immunotolerance was overcome and immunoblot-reactive antibodies were stimulated to epitopes at almost any site of ovine PrPC and PrPSc. A large number of different antibodies cross-reacted also with affinity-purified PrPCs from other mammalian species including cow, goat, pig, man, dog, cat, mink, mouse, hamster and guinea pig. No epitope, however, was recognized exclusively on the pathological or cellular isoform of PrP indicating that both isoforms occur in highly denatured conformations on the immunoblots. Antibodies to the amino-terminus are suitable for immunoprecipitation of PrP. The availability of rabbit and chicken anti-peptide antibodies to PrP will greatly improve immunochemical diagnosis and pathogenetic studies on these diseases.

  14. Correlation spectroscopy and molecular dynamics simulations to study the structural features of proteins.

    PubMed

    Varriale, Antonio; Marabotti, Anna; Mei, Giampiero; Staiano, Maria; D'Auria, Sabato

    2013-01-01

    In this work, we used a combination of fluorescence correlation spectroscopy (FCS) and molecular dynamics (MD) simulation methodologies to acquire structural information on pH-induced unfolding of the maltotriose-binding protein from Thermus thermophilus (MalE2). FCS has emerged as a powerful technique for characterizing the dynamics of molecules and it is, in fact, used to study molecular diffusion on timescale of microsecond and longer. Our results showed that keeping temperature constant, the protein diffusion coefficient decreased from 84±4 µm(2)/s to 44±3 µm(2)/s when pH was changed from 7.0 to 4.0. An even more marked decrease of the MalE2 diffusion coefficient (31±3 µm(2)/s) was registered when pH was raised from 7.0 to 10.0. According to the size of MalE2 (a monomeric protein with a molecular weight of 43 kDa) as well as of its globular native shape, the values of 44 µm(2)/s and 31 µm(2)/s could be ascribed to deformations of the protein structure, which enhances its propensity to form aggregates at extreme pH values. The obtained fluorescence correlation data, corroborated by circular dichroism, fluorescence emission and light-scattering experiments, are discussed together with the MD simulations results.

  15. Involvement of a Botulinum Toxin-Sensitive 22-kDa G Protein in Stimulated Exocytosis of Human Neutrophils

    DTIC Science & Technology

    1994-01-01

    FUNDING NUMBERS ]involvement -of -a Botulinun toxin -sesftUv&2_2At6-d -- Protein in stinulated exocytosis of humnan neutrophi1s. WICD I. AUTHOR(S) Nath...Botulinum Toxin -Sensitive 22-kDa G Protein in Stimulated Exocytosis of Human Neutrophils jaymaree Nath,’ Annette Powledge, and Daniel G. Wright2...observed. Although both peslussis toxin and ST-O Inhibited excocytosis In FlMvLP-stimvulated PMNs, the inhibitory effects o( the two toxins were found to be

  16. Identification of new protein-protein interactions involving the products of the chromosome- and plasmid-encoded type IV secretion loci of the phytopathogen Xanthomonas axonopodis pv. citri.

    PubMed

    Alegria, Marcos C; Souza, Diorge P; Andrade, Maxuel O; Docena, Cassia; Khater, Leticia; Ramos, Carlos H I; da Silva, Ana C R; Farah, Chuck S

    2005-04-01

    The recently sequenced genome of the bacterial plant pathogen Xanthomonas axonopodis pv. citri contains two virB gene clusters, one on the chromosome and one on a 64-kb plasmid, each of which codes for a previously uncharacterized type IV secretion system (T4SS). Here we used a yeast two-hybrid assay to identify protein-protein interactions in these two systems. Our results revealed interactions between known T4SS components as well as previously uncharacterized interactions involving hypothetical proteins coded by open reading frames in the two X. axonopodis pv. citri virB loci. Our results indicate that both loci may code for previously unidentified VirB7