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

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 ProRule

pfsearchV3: a code acceleration and heuristic to search PROSITE profiles

Summary: The PROSITE resource provides a rich and well annotated source of signatures in the form of generalized profiles that allow protein domain detection and functional annotation. One of the major limiting factors in the application of PROSITE in genome and metagenome annotation pipelines is the time required to search protein sequence databases for putative matches. We describe an improved and optimized implementation of the PROSITE search tool pfsearch that, combined with a newly developed heuristic, addresses this limitation. On a modern x86_64 hyper-threaded quad-core desktop computer, the new pfsearchV3 is two orders of magnitude faster than the original algorithm. Availability and implementation: Source code and binaries of pfsearchV3 are freely available for download at http://web.expasy.org/pftools/#pfsearchV3, implemented in C and supported on Linux. PROSITE generalized profiles including the heuristic cut-off scores are available at the same address. Contact: [email protected]

DmCatD, a cathepsin D-like peptidase of the hematophagous insect Dipetalogaster maxima (Hemiptera: Reduviidae): Purification, bioinformatic analyses and the significance of its interaction with lipophorin in the internalization by developing oocytes

DmCatD, a cathepsin D-like peptidase of the hematophagous insect Dipetalogaster maxima, is synthesized by the fat body and the ovary and functions as yolk protein precursor. Functionally, DmCatD is involved in vitellin proteolysis. In this work, we purified and sequenced DmCatD, performed bioinformatic analyses and investigated the events involved in its targeting and storage in developing oocytes. By ion exchange and gel filtration chromatography, DmCatD was purified from egg homogenates and its identity was confirmed by mass spectrometry. Approximately 73% of the full-length transcript was sequenced. The phylogeny indicated that DmCatD has features which suggest its distancing from “classical” cathepsins D. Bioinformatic analyses using a chimeric construct were employed to predict post-translational modifications. Structural modeling showed that DmCatD exhibited the expected folding for this type of enzyme, and an active site with conserved architecture. The interaction between DmCatD and lipophorin in the hemolymph was demonstrated by co-immunoprecipitation. Colocalization of both proteins in developing oocyte membranes and yolk bodies was detected by immunofluorescence. Docking assays favoring the interaction DmCatD-lipophorin were carried out after modeling lipophorin of a related triatomine species. Our results suggest that lipophorin acts as a carrier for DmCatD to facilitate its further internalization by the oocytes. The mechanisms involved in the uptake of peptidases within the oocytes of insects have not been reported. This is the first experimental work supporting the interaction between cathepsin D and lipophorin in an insect species, enabling us to propose a pathway for its targeting and storage in developing oocytes.Fil: Leyria, Jimena. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Córdoba. Centro de Investigaciones en Bioquímica Clínica e Inmunología; Argentina. Universidad Nacional de Córdoba. Facultad de Ciencias Químicas. Departamento de Bioquímica Clínica; ArgentinaFil: Fruttero, Leonardo Luis. Universidad Nacional de Córdoba. Facultad de Ciencias Químicas. Departamento de Bioquímica Clínica; Argentina. Pontificia Universidade Católica do Rio Grande do Sul; Brasil. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Córdoba. Centro de Investigaciones en Bioquímica Clínica e Inmunología; ArgentinaFil: Ligabue Braun, Rodrigo. Universidade Federal do Rio Grande do Sul; BrasilFil: Defferrari, Marina S.. University of Toronto; CanadáFil: Arrese, Estela L.. Oklahoma State University; Estados UnidosFil: Soulages, José L.. Oklahoma State University; Estados UnidosFil: Settembrini, Beatriz Patricia. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Museo Argentino de Ciencias Naturales “Bernardino Rivadavia”; ArgentinaFil: Carlini, Célia Regina R S. Pontificia Universidade Católica do Rio Grande do Sul; BrasilFil: Canavoso, Lilian Etelvina. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Córdoba. Centro de Investigaciones en Bioquímica Clínica e Inmunología; Argentina. Universidad Nacional de Córdoba. Facultad de Ciencias Químicas. Departamento de Bioquímica Clínica; Argentin

In silico identification, characterization and expression profile of WUSCHEL-related homeobox (WOX) gene family in Vanilla planifolia

Vanilla planifolia is an economically important orchid, which is being commercially exploited by the food industry for the highly valued secondary metabolite vanillin. WUSCHEL-related homeobox (WOX) gene family encodes for WUSCHEL-related homeobox (WOX) transcription factors that participate in embryogenesis, organogenesis and florigenesis and in diverse plant developmental processes as well. In the present study, we analysed V. planifolia transcriptome and identified 6 WOX (VpWOX) transcripts, that encode putative WOX (VpWOX) transcription factor proteins. Domain analysis was done which indicates the presence of helix-loop-helix-turn-helix which is identifying feature of WOX gene family proteins. We executed phylogenetic clustering for the VpWOX proteins with their counterpart from the model plant Arabidopsis thaliana (AtWOX) and other closely related orchid species, Phalaenopsis equestris (PeWOX), Dendrobium catenatum (DcWOX) and Apostasia shenzhenica (AsWOX) and established their clade specific grouping. Spatio-temporal expression profile for VpWOX genes was analysed for different plant developmental stages which shows that VpWOX13 is expressing uniformly in all the developmental stages whereas, other genes have tissue specific expression. Based on gene expression patterns, we selected four VpWOX proteins and carried out secondary and tertiary structural analysis which indicates the presence of alpha helix and beta turn in the protein structure. The present study provides basic understanding of the functioning of WOX gene family in V. planifolia and paves the path for functional characterization of selected VpWOX genes in planta and in heterologous system in future for commercial utilization

Genome wide characterization of WUSCHEL-related homeobox (WOX) gene family in Apostasia shenzhenica, a primeval orchid

In the present study, we report identification and characterization of the plant-specific WUSCHEL-related homeobox (WOX) gene family in Apostasia shenzhenica, a primeval orchid. WOX proteins are DNA-binding WUSCHEL-related homeobox (WOX) encoding transcription factors that play critical role in zygote patterning, embryo development, organogenesis, florigenesis, stress responses etc. Ten putative AsWOX genes were predicted in the A. shenzhenica genome and were characterized by the presence of DNA-binding helix-loop-helix-turn-helix motif. AsWOX proteins were grouped into three clades, ancient, intermediate and WUS on the basis of sequence homology with Arabidopsis thaliana (AtWOX), Oryza sativa (OsWOX), Phalaenopsis equestris (PeWOX) and Dendrobium catenatum (DcWOX) and their phylogenetic relationship was established. Gene structure analysis revealed that three AsWOX genes had two introns, six genes had a single intron, and one gene was intron-less. Expression profiling in a variety of tissue (tubers, seeds and pollens) was analysed in light of the presence of specific cis-regulatory elements in the promoter region and their role in various developmental processes was discussed. Three dimensional structures were predicted for three selected AsWOX proteins representing the three clades. The present study provides insights to the role of AsWOX gene family in various vital developmental processes, establishes phylogenetic relationships with related plant species and provides a platform for functional validation of specific AsWOX genes

Motif Discovery in Protein Sequences

Biology has become a data‐intensive research field. Coping with the flood of data from the new genome sequencing technologies is a major area of research. The exponential increase in the size of the datasets produced by “next‐generation sequencing” (NGS) poses unique computational challenges. In this context, motif discovery tools are widely used to identify important patterns in the sequences produced. Biological sequence motifs are defined as short, usually fixed length, sequence patterns that may represent important structural or functional features in nucleic acid and protein sequences such as transcription binding sites, splice junctions, active sites, or interaction interfaces. They can occur in an exact or approximate form within a family or a subfamily of sequences. Motif discovery is therefore an important field in bioinformatics, and numerous methods have been developed for the identification of motifs shared by a set of functionally related sequences. This chapter will review the existing motif discovery methods for protein sequences and their ability to discover biologically important features as well as their limitations for the discovery of new motifs. Finally, we will propose new horizons for motif discovery in order to address the short comings of the existent methods