We present a model of opinion dynamics in which agents adjust continuous opinions as a result of random binary encounters whenever their difference in opinion is below a given threshold. High thresholds yield convergence of opinions towards an average opinion, whereas low thresholds result in several opinion clusters: members of the same cluster share the same opinion but are no longer influenced by members of other clusters.
The synthesis of both pro-inflammatory leukotrienes and anti-inflammatory lipoxins requires the enzyme 5-lipoxygenase (5-LOX). 5-LOX activity is short-lived, apparently in part due to an intrinsic instability of the enzyme. We identified a 5-LOX-specific destabilizing sequence that is involved in orienting the carboxy-terminus which binds the catalytic iron. Herein we report the crystal structure at 2.4 Å resolution of human 5-LOX stabilized by replacement of this sequence.
Neuropilin (Nrp) is a cell surface receptor with essential roles in angiogenesis and axon guidance. Interactions between Nrp and the positively charged C termini of its ligands, VEGF and semaphorin, are mediated by Nrp domains b1 and b2, which share homology to coagulation factor domains. We report here the crystal structure of the tandem b1 and b2 domains of Nrp-1 (N1b1b2) and show that they form a single structural unit. Cocrystallization of N1b1b2 with Tuftsin, a peptide mimic of the VEGF C terminus, reveals the site of interaction with the basic tail of VEGF on the b1 domain. We also show that heparin promotes N1b1b2 dimerization and map the heparin binding site on N1b1b2. These results provide a detailed picture of interactions at the core of the Nrp signaling complex and establish a molecular basis for the synergistic effects of heparin on Nrp-mediated signaling.semaphorin ͉ Tuftsin ͉ VEGF N europilins (Nrps) are essential cell surface receptors with central roles in both angiogenesis and axon guidance (1-3). During angiogenesis, Nrp directly binds VEGF and functions as a coreceptor for VEGF along with VEGF receptor (VEGFR)-2, one of the three VEGFR tyrosine kinases (3, 4). During neural development, Nrp directly binds semaphorin and functions as a semaphorin coreceptor with members of the plexin family (5). Additionally, interactions with both neural adhesion protein L1 and Nrp-interacting protein (NIP), have been shown to be involved in a variety of other cellular processes (6-8).Nrp plays a stimulatory role in angiogenesis, a process critical for growth of solid tumors (reviewed in refs. 4 and 9-11). Nrp expression is observed in tumor vasculature, and overexpression promotes tumorigenesis in vivo for a variety of solid tumors including pituitary, prostate, breast, and colon cancers (12-15). In contrast, a soluble splice form containing only part of the extracellular domain of Nrp inhibits tumorigenesis (16) as do a number of peptides that block VEGF binding to Nrp (17,18). Recent evidence has also demonstrated a role for Nrp in hematological malignancies. Nrp overexpression is observed in both multiple myeloma and acute myeloid leukemia and, in the latter case, is associated with significantly reduced survival (19,20). Strategies to inhibit Nrp activity are thus being developed as potential antitumor therapies (reviewed in ref. 11).Higher eukaryotes possess two Nrp homologs, Nrp-1 and Nrp-2, which share 44% amino acid sequence identity (1). Nrp extracellular regions are composed of two complement binding CUB domains (a1 and a2) followed by two coagulation factor domains (b1 and b2), a MAM (meprin, A5, ) domain (c1), a single membrane-spanning region, and a short cytoplasmic tail (Fig. 1A) (21, 22). The a1 and a2 domains of Nrp are essential for binding to the core seven-bladed Sema domain of semaphorin as well as contributing to interactions with VEGF (23, 24). The coagulation factor domains b1 and b2 contain the high-affinity binding site for the basic heparin binding domain (HBD) of VEGF165 as well a...
Background: 15-Lipoxygenase-2 is linked to atherosclerotic plaque formation; the homologous enzyme 5-lipoxygenase initiates the synthesis of proinflammatory leukotrienes. Results: We determined the crystal structure of 15-LOX-2 in the presence of a substrate mimic. Conclusion: 15-Lipoxygenase-2 and 5-lipoxgenase display active site variations that confer distinct product specificities. Significance: These differences can be exploited for the design of isoform-specific anti-inflammatories.
Leukotrienes (LT) are lipid mediators of the inflammatory response that are linked to asthma and atherosclerosis. LT biosynthesis is initiated by 5-lipoxygenase (5-LOX) with the assistance of the substrate-binding 5-LOX-activating protein at the nuclear membrane. Here, we contrast the structural and functional consequences of the binding of two natural product inhibitors of 5-LOX. The redox-type inhibitor nordihydroguaiaretic acid (NDGA) is lodged in the 5-LOX active site, now fully exposed by disordering of the helix that caps it in the apo-enzyme. In contrast, the allosteric inhibitor 3-acetyl-11-keto-beta-boswellic acid (AKBA) from frankincense wedges between the membrane-binding and catalytic domains of 5-LOX, some 30 Å from the catalytic iron. While enzyme inhibition by NDGA is robust, AKBA promotes a shift in the regiospecificity, evident in human embryonic kidney 293 cells and in primary immune cells expressing 5-LOX.
The hammerhead ribozyme is a self-cleaving RNA broadly dispersed across all kingdoms of life. Although it was the first of the small, nucleolytic ribozymes discovered, the mechanism by which it catalyzes its reaction remains elusive. The nucleobase of G12 is well positioned to be a general base, but it is unclear if or how this guanine base becomes activated for proton transfer. Metal ions have been implicated in the chemical mechanism, but no interactions between divalent metal ions and the cleavage site have been observed crystallographically. To better understand how this ribozyme functions, we have solved crystal structures of wild-type and G12A mutant ribozymes. We observe a pH-dependent conformational change centered around G12, consistent with this nucleotide becoming deprotonated. Crystallographic and kinetic analysis of the G12A mutant reveals a Zn2+ specificity switch suggesting a direct interaction between a divalent metal ion and the purine at position 12. The metal ion specificity switch and the pH–rate profile of the G12A mutant suggest that the minor imino tautomer of A12 serves as the general base in the mutant ribozyme. We propose a model in which the hammerhead ribozyme rearranges prior to the cleavage reaction, positioning two divalent metal ions in the process. The first metal ion, positioned near G12, becomes directly coordinated to the O6 keto oxygen, to lower the pKa of the general base and organize the active site. The second metal ion, positioned near G10.1, bridges the N7 of G10.1 and the scissile phosphate and may participate directly in the cleavage reaction.
The enzyme 5-lipoxygenase (5-LOX) initiates biosynthesis of the proinflammatory leukotriene lipid mediators and, together with 15-LOX, is also required for synthesis of the anti-inflammatory lipoxins. The catalytic activity of 5-LOX is regulated through multiple mechanisms, including Ca(2+)-targeted membrane binding and phosphorylation at specific serine residues. To investigate the consequences of phosphorylation at S663, we mutated the residue to the phosphorylation mimic Asp, providing a homogenous preparation suitable for catalytic and structural studies. The S663D enzyme exhibits robust 15-LOX activity, as determined by spectrophotometric and HPLC analyses, with only traces of 5-LOX activity remaining; synthesis of the anti-inflammatory lipoxin A(4) from arachidonic acid is also detected. The crystal structure of the S663D mutant in the absence and presence of arachidonic acid (in the context of the previously reported Stable-5-LOX) reveals substantial remodeling of helices that define the active site so that the once fully encapsulated catalytic machinery is solvent accessible. Our results suggest that phosphorylation of 5-LOX at S663 could not only down-regulate leukotriene synthesis but also stimulate lipoxin production in inflammatory cells that do not express 15-LOX, thus redirecting lipid mediator biosynthesis to the production of proresolving mediators of inflammation.
Access to experimental X-ray diffraction image data is fundamental for validation and reproduction of macromolecular models and indispensable for development of structural biology processing methods. Here, we established a diffraction data publication and dissemination system, Structural Biology Data Grid (SBDG; data.sbgrid.org), to preserve primary experimental data sets that support scientific publications. Data sets are accessible to researchers through a community driven data grid, which facilitates global data access. Our analysis of a pilot collection of crystallographic data sets demonstrates that the information archived by SBDG is sufficient to reprocess data to statistics that meet or exceed the quality of the original published structures. SBDG has extended its services to the entire community and is used to develop support for other types of biomedical data sets. It is anticipated that access to the experimental data sets will enhance the paradigm shift in the community towards a much more dynamic body of continuously improving data analysis.
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