Structural characteristics of novel protein folds

Folds are the basic building blocks of protein structures. Understanding the emergence of novel protein folds is an important step towards understanding the rules governing the evolution of protein structure and function and for developing tools for protein structure modeling and design. We explored the frequency of occurrences of an exhaustively classified library of supersecondary structural elements (Smotifs), in protein structures, in order to identify features that would define a fold as novel compared to previously known structures. We found that a surprisingly small set of Smotifs is sufficient to describe all known folds. Furthermore, novel folds do not require novel Smotifs, but rather are a new combination of existing ones. Novel folds can be typified by the inclusion of a relatively higher number of rarely occurring Smotifs in their structures and, to a lesser extent, by a novel topological combination of commonly occurring Smotifs. When investigating the structural features of Smotifs, we found that the top 10% of most frequent ones have a higher fraction of internal contacts, while some of the most rare motifs are larger, and contain a longer loop region

The HSV-1 ubiquitin ligase ICP0: modifying the cellular proteome to promote infection

Herpes simplex virus 1 (HSV-1) hijacks ubiquitination machinery to modify the cellular proteome to create an environment permissive for virus replication. HSV-1 encodes its own RING-finger E3 ubiquitin (Ub) ligase, Infected Cell Protein 0 (ICP0), that directly interfaces with component proteins of the Ub pathway to inactivate host immune defences and cellular processes that restrict the progression of HSV-1 infection. Consequently, ICP0 plays a critical role in the infectious cycle of HSV-1 that is required to promote the efficient onset of lytic infection and productive reactivation of viral genomes from latency. This review will describe the current knowledge regarding the biochemical properties and known substrates of ICP0 during HSV-1 infection. We will highlight the gaps in the characterization of ICP0 function and propose future areas of research required to understand fully the biological properties of this important HSV-1 regulatory protein

EPIC-DB: a proteomics database for studying Apicomplexan organisms

<p>Abstract</p> <p>Background</p> <p>High throughput proteomics experiments are useful for analyzing the protein expression of an organism, identifying the correct gene structure of a genome, or locating possible post-translational modifications within proteins. High throughput methods necessitate publicly accessible and easily queried databases for efficiently and logically storing, displaying, and analyzing the large volume of data.</p> <p>Description</p> <p>EPICDB is a publicly accessible, queryable, relational database that organizes and displays experimental, high throughput proteomics data for <it>Toxoplasma gondii </it>and <it>Cryptosporidium parvum</it>. Along with detailed information on mass spectrometry experiments, the database also provides antibody experimental results and analysis of functional annotations, comparative genomics, and aligned expressed sequence tag (EST) and genomic open reading frame (ORF) sequences. The database contains all available alternative gene datasets for each organism, which comprises a complete theoretical proteome for the respective organism, and all data is referenced to these sequences. The database is structured around clusters of protein sequences, which allows for the evaluation of redundancy, protein prediction discrepancies, and possible splice variants. The database can be expanded to include genomes of other organisms for which proteome-wide experimental data are available.</p> <p>Conclusion</p> <p>EPICDB is a comprehensive database of genome-wide <it>T. gondii </it>and <it>C. parvum </it>proteomics data and incorporates many features that allow for the analysis of the entire proteomes and/or annotation of specific protein sequences. EPICDB is complementary to other -genomics- databases of these organisms by offering complete mass spectrometry analysis on a comprehensive set of all available protein sequences.</p

Comparative proteomics identifies Schlafen 5 (SLFN5) as a herpes simplex virus restriction factor that suppresses viral transcription

Intrinsic antiviral host factors confer cellular defence by limiting virus replication and are often counteracted by viral countermeasures. We reasoned that host factors that inhibit viral gene expression could be identified by determining proteins bound to viral DNA (vDNA) in the absence of key viral antagonists. Herpes simplex virus 1 (HSV-1) expresses E3 ubiquitin-protein ligase ICP0 (ICP0), which functions as an E3 ubiquitin ligase required to promote infection. Cellular substrates of ICP0 have been discovered as host barriers to infection but the mechanisms for inhibition of viral gene expression are not fully understood. To identify restriction factors antagonized by ICP0, we compared proteomes associated with vDNA during HSV-1 infection with wild-type virus and a mutant lacking functional ICP0 (ΔICP0). We identified the cellular protein Schlafen family member 5 (SLFN5) as an ICP0 target that binds vDNA during HSV-1 ΔICP0 infection. We demonstrated that ICP0 mediates ubiquitination of SLFN5, which leads to its proteasomal degradation. In the absence of ICP0, SLFN5 binds vDNA to repress HSV-1 transcription by limiting accessibility of RNA polymerase II to viral promoters. These results highlight how comparative proteomics of proteins associated with viral genomes can identify host restriction factors and reveal that viral countermeasures can overcome SLFN antiviral activity

Computational Analysis and Experimental Validation of Gene Predictions in Toxoplasma gondii

Toxoplasma gondii is an obligate intracellular protozoan that infects 20 to 90% of the population. It can cause both acute and chronic infections, many of which are asymptomatic, and, in immunocompromised hosts, can cause fatal infection due to reactivation from an asymptomatic chronic infection. An essential step towards understanding molecular mechanisms controlling transitions between the various life stages and identifying candidate drug targets is to accurately characterize the T. gondii proteome.We have explored the proteome of T. gondii tachyzoites with high throughput proteomics experiments and by comparison to publicly available cDNA sequence data. Mass spectrometry analysis validated 2,477 gene coding regions with 6,438 possible alternative gene predictions; approximately one third of the T. gondii proteome. The proteomics survey identified 609 proteins that are unique to Toxoplasma as compared to any known species including other Apicomplexan. Computational analysis identified 787 cases of possible gene duplication events and located at least 6,089 gene coding regions. Commonly used gene prediction algorithms produce very disparate sets of protein sequences, with pairwise overlaps ranging from 1.4% to 12%. Through this experimental and computational exercise we benchmarked gene prediction methods and observed false negative rates of 31 to 43%.This study not only provides the largest proteomics exploration of the T. gondii proteome, but illustrates how high throughput proteomics experiments can elucidate correct gene structures in genomes

The evolution of protein folds from the perspective of structure motifs

Understanding how protein structures evolve is essential for deciphering relationships between homologous proteins, which can inform structure classification and function annotation and aid in protein modeling and design methods. The observation that structure is more conserved than sequence, over the course of evolution, implies a model of evolution where sequences diverge within a discrete set of well-defined folds, which suggests that homology does not exist across fold definitions. However, as more structures have been experimentally solved and the coverage of the universe of folds has increased, the original view of a discrete fold space has been revised to include a more nuanced view of a continuous space defined by regions in which structural similarities can connect globally disparate topologies. Structural, functional and evolutionary relationships are known to, in some cases, span fold definitions. A hallmark of relationships connecting disparate topologies is the conservation of local structure motifs within globally different folds. In order to systematically identify and analyze these relationships, a new approach to structure comparisons and structure classification is required. The goal of this work is to systematically identify evolutionary relationships between folds and to generate a classification of the fold universe that can accurately represent even the relationships that exist across disparate topologies. An exhaustive library of supersecondary-structure motifs (Smotif), defined as two secondary structures connected by a loop, is established and characterized. A novel Smotif-based, superposition-independent structure comparison method (SmotifCOMP) is developed that quantitatively measures the Smotif-based similarity of compared structures in order to identify evolutionary relationships. SmotifCOMP is able to provide a quantitative and robust measure of similarity between disparate topologies since it does not rely on a global superposition. The comparison method is used to perform a systematic analysis of the SCOP Superfamilies and generate a non-hierarchical, network-based representation of the fold universe. This thesis describes the development of a novel method of comparing structures and an improved representation of the relationships within the fold space. This work provides insight into the existence of evolutionary relationships between folds and strengthens the view of a connected and continuous fold universe

The E3 ubiquitin ligase Cul4b promotes CD4+ T cell expansion by aiding the repair of damaged DNA.

The capacity for T cells to become activated and clonally expand during pathogen invasion is pivotal for protective immunity. Our understanding of how T cell receptor (TCR) signaling prepares cells for this rapid expansion remains limited. Here we provide evidence that the E3 ubiquitin ligase Cullin-4b (Cul4b) regulates this process. The abundance of total and neddylated Cul4b increased following TCR stimulation. Disruption of Cul4b resulted in impaired proliferation and survival of activated T cells. Additionally, Cul4b-deficient CD4+ T cells accumulated DNA damage. In T cells, Cul4b preferentially associated with the substrate receptor DCAF1, and Cul4b and DCAF1 were found to interact with proteins that promote the sensing or repair of damaged DNA. While Cul4b-deficient CD4+ T cells showed evidence of DNA damage sensing, downstream phosphorylation of SMC1A did not occur. These findings reveal an essential role for Cul4b in promoting the repair of damaged DNA to allow survival and expansion of activated T cells

Fertilization competence and sperm size variation in sperm-heteromorphic insects

Between species, variation in sperm size has been related to male-female coevolution and male-male competition. In contrast, variation within species is poorly understood. A particular case of intraspecific sperm-size variation occurs in sperm-heteromorphic species, where males produce distinct sperm morphotypes, usually only one of which is fertile. This allows to investigate sperm size variation under different selection regimes. Nonfertile morphotypes, whose role is aside from fertilization, may have other functions, and this may be reflected by changes in developmental processes and a different phenotype compared to fertile sperm. We show that the intraspecific coefficient of variation in sperm length is up to four times lower for fertile than nonfertile morphotypes across 150 sperm-heteromorphic species (70 butterfly, 71 moth, 9 diopsid fly species). This is in agreement with a previous study on 11 species in the Drosophila obscura group. Significantly lower variation in fertile than nonfertile sperm morphometry may result from fertilization-related selection for optimal sperm size, novel functions of nonfertile sperm, or from tighter control of fertile sperm development. More data are needed to clarify the consequences and adaptive significance of within-morph variation, and its consistent pattern across sperm-heteromorphic insect