Crystal Structure of the 6-Hydroxymethyl-7,8-Dihydropterin Pyrophosphokinase•Dihydropteroate Synthase Bifunctional Enzyme from Francisella tularensis

The 6-hydroxymethyl-7,8-dihydropterin pyrophosphokinase (HPPK) and dihydropteroate synthase (DHPS) enzymes catalyze sequential metabolic reactions in the folate biosynthetic pathway of bacteria and lower eukaryotes. Both enzymes represent validated targets for the development of novel anti-microbial therapies. We report herein that the genes which encode FtHPPK and FtDHPS from the biowarfare agent Francisella tularensis are fused into a single polypeptide. The potential of simultaneously targeting both modules with pterin binding inhibitors prompted us to characterize the molecular details of the multifunctional complex. Our high resolution crystallographic analyses reveal the structural organization between FtHPPK and FtDHPS which are tethered together by a short linker. Additional structural analyses of substrate complexes reveal that the active sites of each module are virtually indistinguishable from those of the monofunctional enzymes. The fused bifunctional enzyme therefore represents an excellent vehicle for finding inhibitors that engage the pterin binding pockets of both modules that have entirely different architectures. To demonstrate that this approach has the potential of producing novel two-hit inhibitors of the folate pathway, we identify and structurally characterize a fragment-like molecule that simultaneously engages both active sites. Our study provides a molecular framework to study the enzyme mechanisms of HPPK and DHPS, and to design novel and much needed therapeutic compounds to treat infectious diseases

DiVenn: An Interactive and Integrated Web-Based Visualization Tool for Comparing Gene Lists

Gene expression data generated from multiple biological samples (mutant, double mutant, and wild-type) are often compared via Venn diagram tools. It is of great interest to know the expression pattern between overlapping genes and their associated gene pathways or gene ontology (GO) terms. We developed DiVenn (Dive into the Venn diagram and create a force directed graph)—a novel web-based tool that compares gene lists from multiple RNA-Seq experiments in a force-directed graph, which shows the gene regulation levels for each gene and integrated KEGG pathway and gene ontology knowledge for the data visualization. DiVenn has four key features: (1) informative force-directed graph with gene expression levels to compare multiple data sets; (2) interactive visualization with biological annotations and integrated pathway and GO databases, which can be used to subset or highlight gene nodes to pathway or GO terms of interest in the graph; (3) Pathway and GO enrichment analysis of all or selected genes in the graph; and (4) high resolution image and gene-associated information export. DiVenn is freely available at http://divenn.noble.org/

BGEM: An In Situ Hybridization Database of Gene Expression in the Embryonic and Adult Mouse Nervous System

This article describes an open-access gene expression database analyzed for more than 2,000 genes on mouse nervous system tissue in the coronal, sagittal, and transverse orientation representing multiple developmental ages

The OneOklahoma Cyberinfrastructure Initiative: A Model for Multi-institutional Collaboration

The OneOklahoma Cyberinfrastructure Initiative: A Model for Multi-institutional Collaboration A White Paper submitted by the OneOklahoma Cyberinfrastructure Initiative membership to the National Science Foundation Workshop on The Role of Regional Organizations in Improving Access to the National Computational Infrastructure The OneOklahoma Cyberinfrastructure Initiative (OneOCII) is a statewide all-inclusive advanced digital services collaboration that has been providing access to Cyberinfrastructure (CI) resources, as well as expertise and education, so far to over 100 institutions and organizations statewide (over 50 academic and almost 50 non-academic), including PhD-granting universities, primarily undergraduate institutions, community colleges, career techs and high schools, among them 10 Minority Serving Institutions (MSIs).The OneOklahoma Cyberinfrastructure Initiative OneOCIIN

Interaction of the ^FtDHPS module with Compound 1.

<p>(A) Schematic comparison between the scaffolds of Compound 1 and DHP-PP. Compound 1 comprises a pterin-like core and is missing half of the B-ring as highlighted in orange. (B) Stereo view of Compound 1 (orange) bound within the pterin pocket of the TIM-barrel. Residues that make van der Waals and hydrogen-bond contacts are labeled and shown as pink sticks. The <i>F</i>o-<i>F</i>c simulated-annealing omit electron density for Compound 1 is shown as a blue mesh contoured at 3.5 σ.</p

The overall structure of the HPPK-DHPS bifunctional enzyme from <i>Francisella tularensis</i>.

<p>(A) A stereo view of the overall fold and domain organization showing the secondary structure elements within each module. Each element is labeled with the prefixes ‘H’ and ‘D’ to reflect their locations in the HPPK (blue) and DHPS (purple) domains, respectively. The N- and C-termini and the linker region (green) are labeled. Note that helix Dα8 in the canonical DHPS TIM-barrel is missing. (B) A surface representation of the view shown in (A) that highlights the position of the domain linker and the cleft within the DHPS module corresponding to the missing Dα8 TIM-barrel α-helix.</p

Transcript profiling of hairy vetch (Vicia villosa Roth) identified interesting genes for seed dormancy

Abstract Hairy vetch, a diploid annual legume species, has a robust growth habit, high biomass yield, and winter hardy characteristics. Seed hardness is a major constraint for growing hairy vetch commercially. Hard seeded cultivars are valuable as forages, whereas soft seeded and shatter resistant cultivars have advantages for their use as a cover crop. Transcript analysis of hairy vetch was performed to understand the genetic mechanisms associated with important hairy vetch traits. RNA was extracted from leaves, flowers, immature pods, seed coats, and cotyledons of contrasting soft and hard seeded “AU Merit” plants. A range of 31.22–79.18 Gb RNA sequence data per tissue sample were generated with estimated coverage of 1040–2639×. RNA sequence assembly and mapping of the contigs against the Medicago truncatula (V4.0) genome identified 76,422 gene transcripts. A total of 24,254 transcripts were constitutively expressed in hairy vetch tissues. Key genes, such as KNOX4 (a class II KNOTTED‐like homeobox KNOXII gene), qHs1 (endo‐1,4‐β‐glucanase), GmHs1‐1 (calcineurin‐like metallophosphoesterase), chitinase, shatterproof 1 and 2 (SHP1, SHP2), shatter resistant 1–5 (SHAT1–5)(NAC transcription factor), PDH1 (prephenate dehydrogenase 1), and pectin methylesterases with a potential role in seed hardness and pod shattering, were further explored based on genes involved in seed hardness from other species to query the hairy vetch transcriptome data. Identification of interesting candidate genes in hairy vetch can facilitate the development of improved cultivars with desirable seed characteristics for use as a forage and as a cover crop

Data Collection and Refinement Statistics.

<p>*Data were collected from a single crystal. Values in parentheses are for the highest-resolution shell.</p>a<p>R_free was calculated using 5% of the reflections.</p