Enriching the annotation of Mycobacterium tuberculosis H37Rv proteome using remote homology detection approaches: Insights into structure and function

The availability of the genome sequence of Mycobacterium tuberculosis H37Rv has encouraged determination of large numbers of protein structures and detailed definition of the biological information encoded therein; yet, the functions of many proteins in M. tuberculosis remain unknown. The emergence of multidrug resistant strains makes it a priority to exploit recent advances in homology recognition and structure prediction to re-analyse its gene products. Here we report the structural and functional characterization of gene products encoded in the M. tuberculosis genome, with the help of sensitive profile-based remote homology search and fold recognition algorithms resulting in an enhanced annotation of the proteome where 95% of the M. tuberculosis proteins were identified wholly or partly with information on structure or function. New information includes association of 244 proteins with 205 domain families and a separate set of new association of folds to 64 proteins. Extending structural information across uncharacterized protein families represented in the M. tuberculosis proteome, by determining superfamily relationships between families of known and unknown structures, has contributed to an enhancement in the knowledge of structural content. In retrospect, such superfamily relationships have facilitated recognition of probable structure and/or function for several uncharacterized protein families, eventually aiding recognition of probable functions for homologous proteins corresponding to such families. Gene products unique to mycobacteria for which no functions could be identified are 183. Of these 18 were determined to be M. tuberculosis specific. Such pathogen-specific proteins are speculated to harbour virulence factors required for pathogenesis. A re-annotated proteome of M. tuberculosis, with greater completeness of annotated proteins and domain assigned regions, provides a valuable basis for experimental endeavours designed to obtain a better understanding of pathogenesis and to accelerate the process of drug target discovery. (C) 2014 Elsevier Ltd. All rights reserved.

NrichD database: sequence databases enriched with computationally designed protein-like sequences aid in remote homology detection

NrichD ( ext-link-type=''uri'' xlink:href=''http://proline.biochem.iisc.ernet.in/NRICHD/'' xlink:type=''simple''>http://proline.biochem.iisc.ernet.in/NRICHD/)< /named-content> is a database of computationally designed protein-like sequences, augmented into natural sequence databases that can perform hops in protein sequence space to assist in the detection of remote relationships. Establishing protein relationships in the absence of structural evidence or natural `intermediately related sequences' is a challenging task. Recently, we have demonstrated that the computational design of artificial intermediary sequences/linkers is an effective approach to fill naturally occurring voids in protein sequence space. Through a large-scale assessment we have demonstrated that such sequences can be plugged into commonly employed search databases to improve the performance of routinely used sequence search methods in detecting remote relationships. Since it is anticipated that such data sets will be employed to establish protein relationships, two databases that have already captured these relationships at the structural and functional domain level, namely, the SCOP database and the Pfam database, have been `enriched' with these artificial intermediary sequences. NrichD database currently contains 3 611 010 artificial sequences that have been generated between 27 882 pairs of families from 374 SCOP folds. The data sets are freely available for download. Additional features include the design of artificial sequences between any two protein families of interest to the user.

Homology and enzymatic requirements of microhomology-dependent alternative end joining

Nonhomologous DNA end joining (NHEJ) is one of the major double-strand break (DSB) repair pathways in higher eukaryotes. Recently, it has been shown that alternative NHEJ (A-NHEJ) occurs in the absence of classical NHEJ and is implicated in chromosomal translocations leading to cancer. In the present study, we have developed a novel biochemical assay system utilizing DSBs flanked by varying lengths of microhomology to study microhomology-mediated alternative end joining (MMEJ). We show that MMEJ can operate in normal cells, when microhomology is present, irrespective of occurrence of robust classical NHEJ. Length of the microhomology determines the efficiency of MMEJ, 5 nt being obligatory. Using this biochemical approach, we show that products obtained are due to MMEJ, which is dependent on MRE11, NBS1, LIGASE III, XRCC1, FEN1 and PARP1. Thus, we define the enzymatic machinery and microhomology requirements of alternative NHEJ using a well-defined biochemical system.

De-DUFing the DUFs: Deciphering distant evolutionary relationships of Domains of Unknown Function using sensitive homology detection methods

Background: In the post-genomic era where sequences are being determined at a rapid rate, we are highly reliant on computational methods for their tentative biochemical characterization. The Pfam database currently contains 3,786 families corresponding to ``Domains of Unknown Function'' (DUF) or ``Uncharacterized Protein Family'' (UPF), of which 3,087 families have no reported three-dimensional structure, constituting almost one-fourth of the known protein families in search for both structure and function. Results: We applied a `computational structural genomics' approach using five state-of-the-art remote similarity detection methods to detect the relationship between uncharacterized DUFs and domain families of known structures. The association with a structural domain family could serve as a start point in elucidating the function of a DUF. Amongst these five methods, searches in SCOP-NrichD database have been applied for the first time. Predictions were classified into high, medium and low-confidence based on the consensus of results from various approaches and also annotated with enzyme and Gene ontology terms. 614 uncharacterized DUFs could be associated with a known structural domain, of which high confidence predictions, involving at least four methods, were made for 54 families. These structure-function relationships for the 614 DUF families can be accessed on-line at http://proline.biochem.iisc.ernet.in/RHD_DUFS/. For potential enzymes in this set, we assessed their compatibility with the associated fold and performed detailed structural and functional annotation by examining alignments and extent of conservation of functional residues. Detailed discussion is provided for interesting assignments for DUF3050, DUF1636, DUF1572, DUF2092 and DUF659. Conclusions: This study provides insights into the structure and potential function for nearly 20 % of the DUFs. Use of different computational approaches enables us to reliably recognize distant relationships, especially when they converge to a common assignment because the methods are often complementary. We observe that while pointers to the structural domain can offer the right clues to the function of a protein, recognition of its precise functional role is still `non-trivial' with many DUF domains conserving only some of the critical residues. It is not clear whether these are functional vestiges or instances involving alternate substrates and interacting partners. Reviewers: This article was reviewed by Drs Eugene Koonin, Frank Eisenhaber and Srikrishna Subramanian.

On the link Floer homology of L-space links

We will prove that, for a 2 or 3 component L-space link, HFL^- is completely determined by the multi-variable Alexander polynomial of all the sub-links of L, as well as the pairwise linking numbers of all the components of L. We will also give some restrictions on the multi-variable Alexander polynomial of an L-space link. Finally, we use the methods in this paper to prove a conjecture of Yajing Liu classifying all 2-bridge L-space links.

Topological strings, double affine Hecke algebras, and exceptional knot homology

In this thesis, we consider two main subjects: refined, composite invariants and exceptional knot homologies of torus knots. The main technical tools are double affine Hecke algebras ("DAHA") and various insights from topological string theory.

In particular, we define and study the composite DAHA-superpolynomials of torus knots, which depend on pairs of Young diagrams and generalize the composite HOMFLY-PT polynomials from the full HOMFLY-PT skein of the annulus. We also describe a rich structure of differentials that act on homological knot invariants for exceptional groups. These follow from the physics of BPS states and the adjacencies/spectra of singularities associated with Landau-Ginzburg potentials. At the end, we construct two DAHA-hyperpolynomials which are closely related to the Deligne-Gross exceptional series of root systems.

In addition to these main themes, we also provide new results connecting DAHA-Jones polynomials to quantum torus knot invariants for Cartan types A and D, as well as the first appearance of quantum E6 knot invariants in the literature.

The Homology and Relationship of Human (Homo sapiens) Chromosomes 1, 19 and Dusky Langur (Trachypithacus obscurus) Chromosomes 6, 8 Demonstrated with Chromosome Painting

http://www.jstage.jst.go.jp/

De novo RNA synthesis and homology modeling of the classical swine fever virus RNA polymerase

Classical swine fever virus (CSFV) non-structural protein 5B (NS5B) encodes an RNA-dependent RNA polymerase (RdRp), a key enzyme which initiates RNA replication by a de novo mechanism without a primer and is a potential target for anti-virus therapy. We expressed the NS5B protein in Escherichia coli. The rGTP can stimulate de novo initiation of RNA synthesis and mutation of the GDD motif to Gly-Asp-Asp (GAA) abolishes the RNA synthesis. To better understand the mechanism of viral RNA synthesis in CSFV, a three-dimensional model was built by homology modeling based on the alignment with several virus RdRps. The model contains 605 residues folded in the characteristic fingers, palm and thumb domains. The fingers domain contains an N-terminal region that plays an important role in conformational change. We propose that the experimentally observed promotion of polymerase efficiency by rGTP is probably due to the conformational changes of the polymerase caused by binding the rGTP. Mutation of the GDD to GAA interferes with the interaction between the residues at the polymerase active site and metal ions, and thus renders the polymerase inactive. (c) 2005 Elsevier B.V. All rights reserved.

Homology Induction: the use of machine learning to improve sequence similarity searches

Karwath, A. King, R. Homology induction: the use of machine learning to improve sequence similarity searches. BMC Bioinformatics. 23rd April 2002. 3:11 Additional File Describes the title organims species declaration in one string [http://www.biomedcentral.com/content/supplementary/1471- 2105-3-11-S1.doc] Sponsorship: Andreas Karwath and Ross D. King were supported by the EPSRC grant GR/L62849.

Persistent Homology Analysis of Brain Artery Trees.

New representations of tree-structured data objects, using ideas from topological data analysis, enable improved statistical analyses of a population of brain artery trees. A number of representations of each data tree arise from persistence diagrams that quantify branching and looping of vessels at multiple scales. Novel approaches to the statistical analysis, through various summaries of the persistence diagrams, lead to heightened correlations with covariates such as age and sex, relative to earlier analyses of this data set. The correlation with age continues to be significant even after controlling for correlations from earlier significant summaries.

Regulation of 3-phosphoinositide-dependent protein kinase-1 (PDK1) by src involves tyrosine phosphorylation of PDK1 and Src homology 2 domain binding

3-Phosphoinositide-dependent protein kinase-1 (PDK1) appears to play a central regulatory role in many cell signalings between phosphoinositide-3 kinase and various intracellular serine/threonine kinases. In resting cells, PDK1 is known to be constitutively active and is further activated by tyrosine phosphorylation (Tyr(9) and Tyr(373/376)) following the treatment of the cell with insulin or pervanadate. However, little is known about the mechanisms for this additional activation of PDK1. Here, we report that the SH2 domain of Src, Crk, and GAP recognized tyrosine-phosphorylated PDK1 in vitro. Destabilization of PDK1 induced by geldanamycin (a Hsp90 inhibitor) was partially blocked in HEK 293 cells expressing PDK1- Y9F. Co-expression of Hsp90 enhanced PDK1-Src complex formation and led to further increased PDK1 activity toward PKB and SGK. Immunohistochemical analysis with anti- phospho-Tyr9 antibodies showed that the level of Tyr9 phosphorylation was markedly increased in tumor samples compared with normal. Taken together, these data suggest that phosphorylation of PDK1 on Tyr9, distinct from Tyr373/376, is important for PDK1/Src complex formation, leading to PDK1 activation. Furthermore, Tyr9 phosphorylation is critical for the stabilization of both PDK1 and the PDK1/Src complex via Hsp90-mediated protection of PDK1 degradation.