IHMCIF: An Extension of the PDBx/mmCIF Data Standard for Integrative Structure Determination Methods.

IHMCIF (github.com/ihmwg/IHMCIF) is a data information framework that supports archiving and disseminating macromolecular structures determined by integrative or hybrid modeling (IHM), and making them Findable, Accessible, Interoperable, and Reusable (FAIR). IHMCIF is an extension of the Protein Data Bank Exchange/macromolecular Crystallographic Information Framework (PDBx/mmCIF) that serves as the framework for the Protein Data Bank (PDB) to archive experimentally determined atomic structures of biological macromolecules and their complexes with one another and small molecule ligands (e.g., enzyme cofactors and drugs). IHMCIF serves as the foundational data standard for the PDB-Dev prototype system, developed for archiving and disseminating integrative structures. It utilizes a flexible data representation to describe integrative structures that span multiple spatiotemporal scales and structural states with definitions for restraints from a variety of experimental methods contributing to integrative structural biology. The IHMCIF extension was created with the benefit of considerable community input and recommendations gathered by the Worldwide Protein Data Bank (wwPDB) Task Force for Integrative or Hybrid Methods (wwpdb.org/task/hybrid). Herein, we describe the development of IHMCIF to support evolving methodologies and ongoing advancements in integrative structural biology. Ultimately, IHMCIF will facilitate the unification of PDB-Dev data and tools with the PDB archive so that integrative structures can be archived and disseminated through PDB.

Structural highlights of macromolecular complexes and assemblies.

The structures of macromolecular assemblies have given us deep insights into cellular processes and have profoundly impacted biological research and drug discovery. We highlight the structures of macromolecular assemblies that have been modeled using integrative and computational methods and describe how open access to these structures from structural archives has empowered the research community. The arsenal of experimental and computational methods for structure determination ensures a future where whole organelles and cells can be modeled.

Kidney tissue regeneration using bioactive scaffolds incorporated with differentiating extracellular vesicles and intermediate mesoderm cells.

BACKGROUND: To overcome the limitations of current alternative therapies for chronic kidney disease (CKD), tissue engineering-mediated regeneration strategies have demonstrated the possibilities for complete kidney tissue regeneration. Given the challenges associated with the reproducibility of renal basal cells, the incorporation of intermediate mesoderm (IM) cells and bioactive materials to control bioactivities of cells with supported scaffolds should be considered as a viable approach to enable the regeneration of the complex kidney structure via renal differentiation. METHODS: We developed PMEZ scaffolds by combining crucial bioactive components, such as ricinoleic acid-grafted Mg(OH)2 (M), extracellular matrix (E), and alpha lipoic acid-conjugated ZnO (Z) integrated into biodegradable porous PLGA (P) platform. Additionally, we utilized differentiating extracellular vesicles (dEV) isolated during intermediate mesoderm differentiation into kidney progenitor cells, and IM cells were serially incorporated to facilitate kidney tissue regeneration through their differentiation into kidney progenitor cells in the 3/4 nephrectomy mouse model. RESULTS: The use of differentiating extracellular vesicles facilitated IM differentiation into kidney progenitor cells without additional differentiation factors. This led to improvements in various regeneration-related bioactivities including tubule and podocyte regeneration, anti-fibrosis, angiogenesis, and anti-inflammation. Finally, implanting PMEZ/dEV/IM scaffolds in mouse injury model resulted in the restoration of kidney function. CONCLUSIONS: Our study has demonstrated that utilizing biodegradable PLGA-based scaffolds, which include multipotent cells capable of differentiating into various kidney progenitor cells along with supporting components, can facilitate kidney tissue regeneration in the mouse model that simulates CKD through 3/4 nephrectomy.

Research on ecological restoration assessment and eco-economic development of sea area by introducing the K-means clustering algorithm.

The research uses the K-means clustering algorithm to examine the correlation between ecological restoration assessment, economic development, and investment in sea areas. The study intends to shed light on the dynamics and relationships between these variables and offer insightful information for managing the coastal and marine environments. We have made important discoveries on the effects of ecological restoration and eco-economic development on research investment by thoroughly analyzing data from various indicators, including the ecological index, eco-economic variables, and geographical index. Our findings underscore the significance of protecting marine ecosystems while promoting economic growth by demonstrating the beneficial effects of ecological restoration initiatives on research investment in maritime regions. The report also emphasizes the critical role of eco-economic development in influencing research investment and advancing sustainable practices that balance economic growth and environmental preservation. The results highlight the need for coordinated efforts among local communities, government agencies, academic institutions, business stakeholders, and industry to create comprehensive programs prioritizing ecological restoration, economic development, and research spending. This holistic strategy can support technical advancement, economic diversification, and sustainable development in maritime regions. The study's findings emphasize the importance of coordinating research funding with ecological restoration and eco-economic growth for coastal communities and residents' long-term well-being. Policymakers are urged to prioritize investments in research infrastructure, programs that increase capability, and encouraging regulations that support an environment significant to research and innovation. Stakeholders can use the advantages of research investment, utilize the potential of sea areas, and support the accomplishment of sustainable development goals by implementing these ideas. This study contributes to the existing literature by providing empirical data and insights into the complex relationships between ecological restoration, economic growth, and research expenditures in maritime regions. This paper highlights the study's limitations to encourage further research on new variables and comparisons across various industries and areas. This study provides insights into the importance of incorporating ecological restoration, eco-economic development, and research investment for sustainable coastal and marine management.

The effect of exhalation before the inhalation of dry powder aerosol drugs on the breathing parameters, emitted doses and aerosol size distributions.

Airway deposition of aerosol drugs is highly dependent on the breathing manoeuvre of the patients. Though incorrect exhalation before the inhalation of the drug is one of the most common mistakes, its effect on the rest of the manoeuvre and on the airway deposition distribution of aerosol drugs is not explored in the open literature. The aim of the present work was to conduct inhalation experiments using six dry powder inhalers in order to quantify the effect of the degree of lung emptying on the inhalation time, inhaled volume and peak inhalation flow. Another goal of the research was to determine the effect of the exhalation on the aerodynamic properties of the drugs emitted by the same inhalers. According to the measurements, deep exhalation before drug inhalation increased the volume of the inhaled air and the average and maximum values of the inhalation flow rate, but the extent of the increase was patient and inhaler specific. For different inhalers, the mean value of the relative increase in peak inhalation flow due to forceful exhalation was between 15.3 and 38.4% (min: Easyhaler®, max: Breezhaler®), compared to the case of normal (tidal) exhalation before the drug inhalation. The relative increase in the inhaled volume was between 36.4 and 57.1% (min: NEXThaler®, max: Turbuhaler®). By the same token, forceful exhalation resulted in higher emitted doses and smaller emitted particles, depending on the individual breathing ability of the patient, the inhalation device and the drug metered in it. The relative increase in the emitted dose varied between 0.2 and 8.0% (min: Foster® NEXThaler®, max: Bufomix® Easyhaler®), while the relative enhancement of fine particle dose ranged between 1.9 and 30.8% (min: Foster® NEXThaler®, max: Symbicort® Turbuhaler®), depending on the inhaler. All these effects and parameter values point toward higher airway doses due to forceful exhalation before the inhalation of the drug. At the same time, the present findings highlight the necessity of proper patient education on the importance of lung emptying, but also the importance of patient-specific inhaler-drug pair choice in the future.

Role of Corneal Epithelial Measurements in Differentiating Eyes with Stable Keratoconus from Eyes that Are Progressing.

Purpose: To evaluate measures of corneal epithelium in eyes that showed documented signs of keratoconus (KC) progression and compare with stable eyes and healthy controls. Also, to determine the correlation of these epithelial parameters with maximum keratometry (K max) and pachymetry. Design: Prospective, observational, comparative study. Participants: One-hundred and fifty eyes from 150 patients. The study included 50 eyes from patients with documented KC progression, 50 eyes with stable KC, and 50 clinically normal eyes to serve as controls. Methods: A spectral-domain (SD)-OCT imaging was obtained in all eyes, and mean values were compared between the groups. The correlation of epithelial parameters with K max and thinnest pachymetry was also investigated. Main Outcome Measures: For the purposes of this study, the epithelial measures maximum, minimum, superior, and inferior values as well as the difference between the minimum and maximum (min-max) and epithelial standard deviation were considered, obtained from SD-OCT and compared between groups. Measurements of the thinnest point and min-max in pachymetry were also recorded. Results: The only epithelial parameter that presented a statistically significant difference between stable and progressive KC was epithelium min-max. Although stable KC presented epithelium min-max mean values of -18.2 ± 6.6, progressive KC eyes presented mean values of -23.4 ± 10.3 (P < 0.0001). Epithelial maximum (P = 0.16), minimum (P = 0.25), superior (P = 0.28), inferior (P = 0.23), and standard deviation (P = 0.25) values were not significantly different between stable and progressive eyes. Difference min-max pachymetry points in stable (-108.3 ± 33.5) and progressive KC (-115.2 ± 56.0) were not significantly different (P = 0.723). There was no significant correlation between epithelium min-max with corneal thinning (P = 0.39) or K max (P = 0.09) regardless of disease progression. Conclusions: Epithelial measures are useful to identify KC eyes that are progressing; the parameters that measure the difference between min-max epithelium points were significantly different between stable and progressive groups, unlike this difference in pachymetry. Finally, this epithelial parameter seems to be independent of corneal thinning and K max. Financial Disclosures: Proprietary or commercial disclosure may be found after the references.

Electron microscopy holdings of the Protein Data Bank: the impact of the resolution revolution, new validation tools, and implications for the future.

As a discipline, structural biology has been transformed by the three-dimensional electron microscopy (3DEM) "Resolution Revolution" made possible by convergence of robust cryo-preservation of vitrified biological materials, sample handling systems, and measurement stages operating a liquid nitrogen temperature, improvements in electron optics that preserve phase information at the atomic level, direct electron detectors (DEDs), high-speed computing with graphics processing units, and rapid advances in data acquisition and processing software. 3DEM structure information (atomic coordinates and related metadata) are archived in the open-access Protein Data Bank (PDB), which currently holds more than 11,000 3DEM structures of proteins and nucleic acids, and their complexes with one another and small-molecule ligands (~ 6% of the archive). Underlying experimental data (3DEM density maps and related metadata) are stored in the Electron Microscopy Data Bank (EMDB), which currently holds more than 21,000 3DEM density maps. After describing the history of the PDB and the Worldwide Protein Data Bank (wwPDB) partnership, which jointly manages both the PDB and EMDB archives, this review examines the origins of the resolution revolution and analyzes its impact on structural biology viewed through the lens of PDB holdings. Six areas of focus exemplifying the impact of 3DEM across the biosciences are discussed in detail (icosahedral viruses, ribosomes, integral membrane proteins, SARS-CoV-2 spike proteins, cryogenic electron tomography, and integrative structure determination combining 3DEM with complementary biophysical measurement techniques), followed by a review of 3DEM structure validation by the wwPDB that underscores the importance of community engagement.

New system for archiving integrative structures.

Structures of many complex biological assemblies are increasingly determined using integrative approaches, in which data from multiple experimental methods are combined. A standalone system, called PDB-Dev, has been developed for archiving integrative structures and making them publicly available. Here, the data standards and software tools that support PDB-Dev are described along with the new and updated components of the PDB-Dev data-collection, processing and archiving infrastructure. Following the FAIR (Findable, Accessible, Interoperable and Reusable) principles, PDB-Dev ensures that the results of integrative structure determinations are freely accessible to everyone.

Lucha epidemiológica contra pandemia COVID-19: Impacto de las medidas sociales en el SVI / Epidemiological fight against the COVID-19 pandemic: Impact of social measures on IVS

La lucha epidemiológica contra la pandemia COVID-19 ha incluido medidas sociales como el confinamiento y el cierre de actividades laborales y académicas. A consecuencia, tareas que se llevaban a cabo en el contexto presencial debieron ser ejecutadas desde los hogares, adoptando estrategias como el teletrabajo y la educación virtual, haciendo necesario el uso de herramientas como ordenadores y laptops. De allí que las personas han incrementado la exposición a las pantallas de dispositivos electrónicos, como son computadoras y laptops, trayendo como consecuencia afectaciones en la salud visual de las personas tales como el síndrome visual informático (SVI). Se realizó un estudio transversal con el objetivo de determinar la afectación por SVI en la población peruana y su relación con la exposición prolongada a DEV, generada a raíz de las medidas sociales de enfrentamiento a la pandemia de COVID-19. El tiempo promedio en que los sujetos de nuestro estudio usaron DEV dentro de los hogares se acrecentó un 120%, es decir 4,26±2,36 horas diarias adicionales al comparar con el año anterior a la pandemia COVID-19. La adopción masiva de actividades como el teletrabajo y la educación virtual podrían explicar el hecho que los grupos de empleados/patronos y estudiantes fueron los mayores usuarios de DEV, con 10,41 y 9,32 horas diarias. De acuerdo a los hallazgos obtenidos, es estadísticamente válido afirmar que las medidas sociales para enfrentar la pandemia COVID-19 indujeron al aumento en la prevalencia de SVI en los pobladores peruanos (p<0,001). En la actual investigación, la proporción de individuos que manifestaron SVI pasó de 38 a 64%, antes y durante la aplicación de las medidas, respectivamente(AU)

The epidemiological fight against the COVID-19 pandemic has included social measures such as confinement and the closure of work and academic activities. As a consequence, tasks that were carried out in the face-to-face context had to be carried out from homes, adopting strategies such as teleworking and virtual education, requiring the use of tools such as computers and laptops. Hence, people have increased exposure to the screens of electronic devices, such as computers and laptops, resulting in effects on people's visual health such as computer vision syndrome (SVI). A cross-sectional study was carried out in order to determine the affectation by SVI in the Peruvian population and its relationship with prolonged exposure to DEV, generated as a result of social measures to cope with the COVID-19 pandemic. The average time in which the subjects in our study used DEV within homes increased by 120%, that is, 4.26 ± 2.36 additional daily hours when compared to the year before the COVID-19 pandemic. The massive adoption of activities such as teleworking and virtual education could explain the fact that groups of employees / employers and students were the largest users of DEV, with 10.41 and 9.32 hours per day. According to the findings obtained, it is statistically valid to affirm that the social measures to face the COVID-19 pandemic induced an increase in the prevalence of SVI in the Peruvian population (p <0.001). In the current investigation, the proportion of individuals who manifested SVI went from 38 to 64%, before and during the application of the measures, respectively(AU)

Structural and functional analysis of the C-terminal region of Streptococcus gordonii SspB.

Streptococcus gordonii is a member of the viridans streptococci and is an early colonizer of the tooth surface. Adherence to the tooth surface is enabled by proteins present on the S. gordonii cell surface, among which SspB belongs to one of the most well studied cell-wall-anchored adhesin families: the antigen I/II (AgI/II) family. The C-terminal region of SspB consists of three tandemly connected individual domains that display the DEv-IgG fold. These C-terminal domains contain a conserved Ca2+-binding site and isopeptide bonds, and they adhere to glycoprotein 340 (Gp340; also known as salivary agglutinin, SAG). Here, the structural and functional characterization of the C123SspB domain at 2.7 Šresolution is reported. Although the individual C-terminal domains of Streptococcus mutans AgI/II and S. gordonii SspB show a high degree of both sequence and structural homology, superposition of these structures highlights substantial differences in their electrostatic surface plots, and this can be attributed to the relative orientation of the individual domains (C1, C2 and C3) with respect to each other and could reflect their specificity in binding to extracellular matrix molecules. Studies further confirmed that affinity for Gp340 or its scavenger receptor cysteine-rich (SRCR) domains requires two of the three domains of C123SspB, namely C12 or C23, which is different from AgI/II. Using protein-protein docking studies, models for this observed functional difference between C123SspB and C123AgI/II in their binding to SRCR1 are presented.

Structural interactions define assembly adapter function of a type II secretion system pseudopilin.

The type IV filament superfamily comprises widespread membrane-associated polymers in prokaryotes. The type II secretion system (T2SS), a virulence pathway in many pathogens, belongs to this superfamily. A knowledge gap in understanding of the T2SS is the molecular role of a small "pseudopilin" protein. Using multiple biophysical techniques, we have deciphered how this missing component of the Xcp T2SS architecture is structurally integrated, and thereby unlocked its function. We demonstrate that low-abundance XcpH is the adapter that bridges a trimeric initiating tip complex, XcpIJK, with a periplasmic filament of XcpG subunits. Each pseudopilin protein caps an XcpG protofilament in an overall pseudopilus compatible with dimensions of the periplasm and the outer membrane-spanning secretin through which substrates pass. Unexpectedly, to fulfill its adapter function, the XcpH N-terminal helix must be unwound, a property shared with XcpG subunits. We provide an experimentally validated three-dimensional structural model of a complete type IV filament.

Electrochemical investigations for COVID-19 detection-A comparison with other viral detection methods.

Virus-induced infection such as SARS-CoV-2 is a serious threat to human health and the economic setback of the world. Continued advances in the development of technologies are required before the viruses undergo mutation. The low concentration of viruses in environmental samples makes the detection extremely challenging; simple, accurate and rapid detection methods are in urgent need. Of all the analytical techniques, electrochemical methods have the established capabilities to address the issues. Particularly, the integration of nanotechnology would allow miniature devices to be made available at the point-of-care. This review outlines the capabilities of electrochemical methods in conjunction with nanotechnology for the detection of SARS-CoV-2. Future directions and challenges of the electrochemical biosensors for pathogen detection are covered including wearable and conformal biosensors, detection of plant pathogens, multiplexed detection, and reusable biosensors for on-site monitoring, thereby providing low-cost and disposable biosensors.

Duck enteritis virus pUL47, as a late structural protein localized in the nucleus, mainly depends on residues 40 to 50 and 768 to 777 and inhibits IFN-ß signalling by interacting with STAT1.

Duck enteritis virus (DEV) is a member of the Alphaherpesvirinae subfamily. The characteristics of some DEV genes have been reported. However, information regarding the DEV UL47 gene is limited. In this study, we identified the DEV UL47 gene encoding a late structural protein located in the nucleus of infected cells. We further found that two domains of DEV pUL47, amino acids (aa) 40 to 50 and 768 to 777, could function as nuclear localization sequence (NLS) to guide the nuclear localization of pUL47 and nuclear translocation of heterologous proteins, including enhanced green fluorescent protein (EGFP) and beta-galactosidase (ß-Gal). Moreover, pUL47 significantly inhibited polyriboinosinic:polyribocytidylic acid [poly(I:C)]-induced interferon beta (IFN-ß) production and downregulated interferon-stimulated gene (ISG) expression, such as Mx and oligoadenylate synthetase-like (OASL), by interacting with signal transducer and activator of transcription-1 (STAT1).

Neural oscillatory dynamics serving abstract reasoning reveal robust sex differences in typically-developing children and adolescents.

Fluid intelligence, the ability to problem-solve in novel situations, is linked to higher-order cognitive abilities, and to academic achievement in youth. Previous research has demonstrated that fluid intelligence and the underlying neural circuitry continues to develop throughout adolescence. Neuroimaging studies have predominantly focused on identifying the spatial distribution of brain regions associated with fluid intelligence, with only a few studies examining the temporally-sensitive cortical oscillatory dynamics underlying reasoning abilities. The present study collected magnetoencephalography (MEG) during an abstract reasoning task to examine these spatiotemporal dynamics in a sample of 10-to-16 year-old youth. We found increased cortical activity across a distributed frontoparietal network. Specifically, our key results showed: (1) age was associated with increased theta activity in occipital and cerebellar regions, (2) robust sex differences were distributed across frontoparietal regions, and (3) that specific frontoparietal regions differentially predicted abstract reasoning performance among males versus females despite similar mean performance. Among males, increased theta activity mediated the relationship between age and faster reaction times; conversely, among females, decreased theta mediated the relationship between age and improved accuracy. These findings may suggest that males and females engage in distinct neurocognitive strategies across development to achieve similar behavioral outcomes during fluid reasoning tasks.

Neural dynamics of verbal working memory processing in children and adolescents.

Development of cognitive functions and the underlying neurophysiology is evident throughout childhood and adolescence, with higher order processes such as working memory (WM) being some of the last cognitive faculties to fully mature. Previous functional neuroimaging studies of the neurodevelopment of WM have largely focused on overall regional activity levels rather than the temporal dynamics of neural component recruitment. In this study, we used magnetoencephalography (MEG) to examine the neural dynamics of WM in a large cohort of children and adolescents who were performing a high-load, modified verbal Sternberg WM task. Consistent with previous studies in adults, our findings indicated left-lateralized activity throughout the task period, beginning in the occipital cortices and spreading anterior to include temporal and prefrontal cortices during later encoding and into maintenance. During maintenance, the occipital alpha increase that has been widely reported in adults was found to be relatively weak in this developmental sample, suggesting continuing development of this component of neural processing, which was supported by correlational analyses. Intriguingly, we also found sex-specific developmental effects in alpha responses in the right inferior frontal region during encoding and in parietal and occipital cortices during maintenance. These findings suggested a developmental divergence between males and females in the maturation of neural circuitry serving WM during the transition from childhood to adolescence.

Duplicate US1 Genes of Duck Enteritis Virus Encode a Non-essential Immediate Early Protein Localized to the Nucleus.

The duplicate US1 genes of duck enteritis virus (DEV) encode a protein with a conserved Herpes_IE68 domain, which was found to be closely related to the herpes virus immediate early regulatory protein family and is highly conserved among counterparts encoded by Herpes_IE68 genes. Previous studies found the homologous proteins HSV-1 ICP22 and VZV ORF63/ORF70 to be critical for virus transcription and replication. However, little is known about the DEV ICP22 protein. In this paper, we describe the characteristics of this protein based on pharmacological experiments, real-time quantitative Polymerase Chain Reaction, Western blot, and immunofluorescence assays. We also investigate the role of the protein in DEV replication via mutation of US1. As a result, we found that the DEV ICP22 protein is a non-essential immediate early protein predominantly located in the nucleus of infected DEF cells and that DEV replication is impaired by US1 deletion. We also found that ICP22 contains a classical nuclear localization signal (NLS) at 305-312AA, and ICP22 cannot enter the nucleus by itself after mutating residue 309.

Co-localization of and interaction between duck enteritis virus glycoprotein H and L.

BACKGROUND: Duck Enteritis Virus (DEV), belonging to the α-herpesvirus subfamily, is a linear double-stranded DNA virus. Glycoprotein H and L (gH and gL), encoded by UL22 and UL1, are conserved in the family of herpesviruses. They play important roles as gH/gL dimers during viral entry into host cells through cell-cell fusion. The interaction between gH and gL has been confirmed in several human herpesviruses, such as Herpes Simplex Virus (HSV), Epstein-Barr virus (EBV) and Human Cytomegalovirus (HCMV). In this paper, we studied the interaction between DEV gH and gL. RESULTS: Recombinant plasmids pEGFP-N-gH and pDsRED-N-gL were constructed successfully. Expressions of both DEV gH and gL were observed after incubation of COS-7 cells transfected with pEGFP-N-gH and pDsRED-N-gL plasmids after 12 h, respectively. Also, the co-localization of a proportion of the gH and gL was detected in the cytoplasm of COS-7 cells after co-transfection for 24 h. Then, pCMV-Flag-gL and pCMV-Myc-gH recombinant plasmids were constructed and co-transfected into COS-7 cells. It was showed that both gH and gL were tested with positive results through co-immunoprecipitation and Western-blotting. CONCLUSIONS: Our results demonstrated not only the co-localization of DEV gH and gL in COS-7 cells, but also the interaction between them. It will provide an insight for the further studies in terms of protein-protein interaction in DEV.

Developing Hierarchical Schemas and Building Schema Chains Through Practice Play Behavior.

Examining the different stages of learning through play in humans during early life has been a topic of interest for various scholars. Play evolves from practice to symbolic and then later to play with rules. During practice play, infants go through a process of developing knowledge while they interact with the surrounding objects, facilitating the creation of new knowledge about objects and object related behaviors. Such knowledge is used to form schemas in which the manifestation of sensorimotor experiences is captured. Through subsequent play, certain schemas are further combined to generate chains able to achieve behaviors that require multiple steps. The chains of schemas demonstrate the formation of higher level actions in a hierarchical structure. In this work we present a schema-based play generator for artificial agents, termed Dev-PSchema. With the help of experiments in a simulated environment and with the iCub robot, we demonstrate the ability of our system to create schemas of sensorimotor experiences from playful interaction with the environment. We show the creation of schema chains consisting of a sequence of actions that allow an agent to autonomously perform complex tasks. In addition to demonstrating the ability to learn through playful behavior, we demonstrate the capability of Dev-PSchema to simulate different infants with different preferences toward novel vs. familiar objects.

Development of a Prototype System for Archiving Integrative/Hybrid Structure Models of Biological Macromolecules.

Essential processes in biology are carried out by large macromolecular assemblies, whose structures are often difficult to determine by traditional methods. Increasingly, researchers combine measured data and computed information from several complementary methods to obtain "hybrid" or "integrative" structural models of macromolecules and their assemblies. These integrative/hybrid (I/H) models are not archived in the PDB because of the absence of standard data representations and processing mechanisms. Here we present the development of data standards and a prototype system for archiving I/H models. The data standards provide the definitions required for representing I/H models that span multiple spatiotemporal scales and conformational states, as well as spatial restraints derived from different experimental techniques. Based on these data definitions, we have built a prototype system called PDB-Dev, which provides the infrastructure necessary to archive I/H structural models. PDB-Dev is now accepting structures and is open to the community for new submissions.

Molecular characterization of duck enteritis virus UL41 protein.

BACKGROUND: Duck enteritis virus (DEV) belongs to the subfamily Alphaherpesvirinae, and information on the DEV UL41 gene is limited. METHODS: The DEV UL41 gene was cloned into the pET32a(+) vector and expressed in a prokaryotic expression system. Antiserum was raised against a bacterially expressed UL41-His fusion protein for further experiments. Transcription was quantified and UL41 protein expression levels were determined in DEV-infected cells at different time points by RT-qPCR and western blotting, respectively. DEV virions were purified by sucrose gradient centrifugation and analyzed by mass spectrometry to identify protein content. We confirmed the DEV UL41 gene kinetic class using a pharmacological test. IFA was used to analyze the intracellular localization of pUL41. RESULTS: The recombinant expression plasmid, pET-32a(+)-UL41, which highly expresses a 76.0 kDa fusion protein, was constructed and expressed in E. coli BL21 (DE3) after induction with 0.2 mM IPTG at 30 °C for 10 h, generating a specific mouse anti-UL41 protein polyclonal antibody. RT-qPCR and western blot analyses revealed that the UL41 transcript number peaked at 36 hpi, and peak protein expression occurred at 48 hpi. The pharmacological test showed that UL41 was a γ2 gene. Mass spectrometry analysis showed that pUL41 was a virion component. IFA results revealed that pUL41 was localized throughout DEV-infected cells but only localized to the cytoplasm of transfected cells. DEV pUL47 translocated pUL41 to the nuclei of DEF cells; this translocation was dependent on predicted pUL47 NLS signals (40-50 aa and 768-777 aa). CONCLUSIONS: DEV UL41 is a γ2 gene that encodes a virion structural protein, pUL41 localizes throughout DEV-infected cells but only localizes to the cytoplasm of transfected cells. pUL41 cannot autonomously localize to the nucleus, as this nuclear localization is dependent on predicted DEV pUL47 NLS signals (40-50 aa and 768-777 aa).