New and continuing developments at PROSITE.

PROSITE (http://prosite.expasy.org/) consists of documentation entries describing protein domains, families and functional sites, as well as associated patterns and profiles to identify them. It is complemented by ProRule a collection of rules, which increases the discriminatory power of these profiles and patterns by providing additional information about functionally and/or structurally critical amino acids. PROSITE signatures, together with ProRule, are used for the annotation of domains and features of UniProtKB/Swiss-Prot entries. Here, we describe recent developments that allow users to perform whole-proteome annotation as well as a number of filtering options that can be combined to perform powerful targeted searches for biological discovery. The latest version of PROSITE (release 20.85, of 30 August 2012) contains 1308 patterns, 1039 profiles and 1041 ProRules

Method validation of nanoparticle tracking analysis to measure pulmonary nanoparticle content: the size distribution in exhaled breath condensate depends on occupational exposure

A particle exposure assessment based on the dose deposited in the lungs would be the gold standard for the evaluation of any resulting health effects. Measuring particles in exhaled breath condensate (EBC)-a matrix containing water and airway lining fluid-could help to evaluate particle retention in the lungs. This study aimed to (1) validate a nanoparticle tracking analysis (NTA) method for determining the particle number concentration and their hydrodynamic size distribution in EBC, and (2) apply this method to EBC collected from workers exposed to soapstone (n = 55) or quartz dust (n = 12) and controls (n = 11). A standard latex bead solution was used to determine the linear range, limit of detection (LOD), repeatability (coefficient of variation, CV), and bias in spiked EBC. An LM10 NanoSight instrument with NTA version 3.1 software was used for measurement. RTubes(®) were used for field collection of EBC. The repeatability obtained for a D50 size distribution in EBC showed less than 8% variability, with a bias <7%. The particle concentration was linear in the range ≤2.5 × 10(8) particles ml(-1) with a LOD of 4 × 10(6) particles ml(-1). A recovery of 117 ± 20% at 6.2 × 10(7) particles ml(-1) was obtained with a CV <10% and a bias <20%. EBC from workers exposed to quartz, who experienced the largest exposure to silica particles, consistently exhibited a statistically significant (p < 0.01) higher concentration of particles in their EBC, with a size distribution shift towards larger values than the other groups. Results showed that the NTA technique performed well for characterizing the size distribution and concentrations of particles in EBC. The technique needs to be corroborated with a larger population of workers

Does exposure to inflammatory particles modify the pattern of anion in exhaled breath condensate?

Exposure to environmental and occupational particulate matter (PM) induces health effects on the cardio-pulmonary system. In addition, associations between exposure to PM and metabolic syndromes like diabetes mellitus or obesity are now emerging in the literature. Collection of exhaled breath condensate (EBC) is an appealing non-invasive technique to sample pulmonary fluids. This hypothesis-generating study aims to (1) validate an ion chromatography method allowing the robust determination of different metabolism-related molecules (lactate, formate, acetate, propionate, butyrate, pyruvate, nitrite, nitrate) in EBC; (2) apply this method to EBC samples collected from workers exposed to quartz (a known inflammatory particle), to soapstone (a less inflammatory particle than quartz), as well as to controls. A multi-compound standard solution was used to determine the linearity range, detection limit, repeatability and bias from spiked EBC. The biological samples were injected without further treatment into an ion chromatograph with a conductivity detector. RTube <sup>®</sup> were used for field collection of EBC from 11 controls, 55 workers exposed to soapstone and 12 volunteers exposed to quartz dust. The analytical method used proved to be adequate for quantifying eight anions in EBC samples. Its sub-micromolar detection limits and repeatability, combined with a very simple sample preparation, allowed an easy and fast quantification of different glycolysis or nitrosative stress metabolites. Using multivariate discriminant analysis to maximize differences between groups, we observed a different pattern of anions with a higher formate/acetate ratio in the EBC samples for quartz exposed workers compared to the two other groups. We hypothesize that a modification of the metabolic signature could be induced by exposure to inflammatory particles like quartz and might be observed in the EBC via a change in the formate/acetate ratio

PeroxiBase: a database with new tools for peroxidase family classification

Peroxidases (EC 1.11.1.x), which are encoded by small or large multigenic families, are involved in several important physiological and developmental processes. They use various peroxides as electron acceptors to catalyse a number of oxidative reactions and are present in almost all living organisms. We have created a peroxidase database (http://peroxibase.isb-sib.ch) that contains all identified peroxidase-encoding sequences (about 6000 sequences in 940 organisms). They are distributed between 11 superfamilies and about 60 subfamilies. All the sequences have been individually annotated and checked. PeroxiBase can be consulted using six major interlink sections ‘Classes’, ‘Organisms’, ‘Cellular localisations’, ‘Inducers’, ‘Repressors’ and ‘Tissue types’. General documentation on peroxidases and PeroxiBase is accessible in the ‘Documents’ section containing ‘Introduction’, ‘Class description’, ‘Publications’ and ‘Links’. In addition to the database, we have developed a tool to classify peroxidases based on the PROSITE profile methodology. To improve their specificity and to prevent overlaps between closely related subfamilies the profiles were built using a new strategy based on the silencing of residues. This new profile construction method and its discriminatory capacity have been tested and validated using the different peroxidase families and subfamilies present in the database. The peroxidase classification tool called PeroxiScan is accessible at the following address: http://peroxibase.isb-sib.ch/peroxiscan.php

Java GUI for InterProScan (JIPS): A tool to help process multiple InterProScans and perform ortholog analysis

BACKGROUND: Recent, rapid growth in the quantity of available genomic data has generated many protein sequences that are not yet biochemically classified. Thus, the prediction of biochemical function based on structural motifs is an important task in post-genomic analysis. The InterPro databases are a major resource for protein function information. For optimal results, these databases should be searched at regular intervals, since they are frequently updated. RESULTS: We describe here a new program JIPS (Java GUI for InterProScan), a tool for tracking and viewing results obtained from repeated InterProScan searches. JIPS stores matches (in a local database) obtained from InterProScan searches performed with multiple versions of the InterPro database and highlights hits that have been added since the last search of the InterPro database. Results are displayed in an easy-to-use tabular format. JIPS also contains tools to assist with ortholog-based comparative studies of protein signatures. CONCLUSION: JIPS is an efficient tool for performing repeated InterProScans on large batches of protein sequences, tracking and viewing search results, and mining the collected data

ORENZA: a web resource for studying ORphan ENZyme activities

BACKGROUND: Despite the current availability of several hundreds of thousands of amino acid sequences, more than 36% of the enzyme activities (EC numbers) defined by the Nomenclature Committee of the International Union of Biochemistry and Molecular Biology (NC-IUBMB) are not associated with any amino acid sequence in major public databases. This wide gap separating knowledge of biochemical function and sequence information is found for nearly all classes of enzymes. Thus, there is an urgent need to explore these sequence-less EC numbers, in order to progressively close this gap. DESCRIPTION: We designed ORENZA, a PostgreSQL database of ORphan ENZyme Activities, to collate information about the EC numbers defined by the NC-IUBMB with specific emphasis on orphan enzyme activities. Complete lists of all EC numbers and of orphan EC numbers are available and will be periodically updated. ORENZA allows one to browse the complete list of EC numbers or the subset associated with orphan enzymes or to query a specific EC number, an enzyme name or a species name for those interested in particular organisms. It is possible to search ORENZA for the different biochemical properties of the defined enzymes, the metabolic pathways in which they participate, the taxonomic data of the organisms whose genomes encode them, and many other features. The association of an enzyme activity with an amino acid sequence is clearly underlined, making it easy to identify at once the orphan enzyme activities. Interactive publishing of suggestions by the community would provide expert evidence for re-annotation of orphan EC numbers in public databases. CONCLUSION: ORENZA is a Web resource designed to progressively bridge the unwanted gap between function (enzyme activities) and sequence (dataset present in public databases). ORENZA should increase interactions between communities of biochemists and of genomicists. This is expected to reduce the number of orphan enzyme activities by allocating gene sequences to the relevant enzymes

Rampant Adaptive Evolution in Regions of Proteins with Unknown Function in Drosophila simulans

Adaptive protein evolution is pervasive in Drosophila. Genomic studies, thus far, have analyzed each protein as a single entity. However, the targets of adaptive events may be localized to particular parts of proteins, such as protein domains or regions involved in protein folding. We compared the population genetic mechanisms driving sequence polymorphism and divergence in defined protein domains and non-domain regions. Interestingly, we find that non-domain regions of proteins are more frequent targets of directional selection. Protein domains are also evolving under directional selection, but appear to be under stronger purifying selection than non-domain regions. Non-domain regions of proteins clearly play a major role in adaptive protein evolution on a genomic scale and merit future investigations of their functional properties

The Molecular Clockwork of the Fire Ant Solenopsis invicta

This is an open-access article, free of all copyright, and may be freely reproduced, distributed, transmitted, modified, built upon, or otherwise used by anyone for any lawful purpose. The work is made available under the Creative Commons CC0 public domain dedication

Local comparison of protein structures highlights cases of convergent evolution in analogous functional sites

We performed an exhaustive search for local structural similarities in an ensemble of non-redundant protein functional sites. With the purpose of finding new examples of convergent evolution, we selected only those matching sites composed of structural regions whose residue order is inverted in the relative protein sequences

The Pseudouridine Synthase RPUSD4 Is an Essential Component of Mitochondrial RNA Granules

Mitochondrial gene expression is a fundamental process that is largely dependent on nuclear-encoded proteins. Several steps of mitochondrial RNA processing and maturation, including RNA post-transcriptional modification, appear to be spatially organized into distinct foci, which we have previously termed mitochondrial RNA granules (MRGs). Although an increasing number of proteins have been localized to MRGs, a comprehensive analysis of the proteome of these structures is still lacking. Here, we have applied a microscopy-based approach that has allowed us to identify novel components of the MRG proteome. Among these, we have focused our attention on RPUSD4, an uncharacterized mitochondrial putative pseudouridine synthase. We show that RPUSD4 depletion leads to a severe reduction of the steady-state level of the 16S mitochondrial (mt) rRNA with defects in the biogenesis of the mitoribosome large subunit and consequently in mitochondrial translation. We report that RPUSD4 binds 16S mt-rRNA, mt-tRNA(Met), and mt-tRNA(Phe), and we demonstrate that it is responsible for pseudouridylation of the latter. These data provide new insights into the relevance of RNA pseudouridylation in mitochondrial gene expression.This work was supported by the Swiss National Science Foundation Grant SNF310030B_160257/1, Wellcome Trust Grant 096919/Z/11/Z, and core funding of Medical Research Council, UK