The study of a prokaryotic glycolytic enzyme

The overall objective of this project is to generate novel carbohydrate binding proteins for use in glycoprotein analysis which are amenable to large scale production. The approach used here is the modification of prokaryotic glycolytic enzymes. Their enzymatic activity will be eliminated while hoping they still retain their binding capabilities. These proteins will be immobilized onto different surfaces to generate advanced bioanalytical platforms which will have huge commercial potential in the field of glycoanalysis

DNA Binding in High Salt: Analysing the Salt Dependence of Replication Protein A3 from the Halophile Haloferax volcanii

Halophilic archaea maintain intracellular salt concentrations close to saturation to survive in high-salt environments and their cellular processes have adapted to function under these conditions. Little is known regarding halophilic adaptation of the DNA processing machinery, particularly intriguing since protein-DNA interactions are classically salt sensitive. To investigate such adaptation, we characterised the DNA-binding capabilities of recombinant RPA3 from Haloferax volcanii (HvRPA3). Under physiological salt conditions (3M KCl), HvRPA3 is monomeric, binding 18 nucleotide ssDNA with nanomolar affinity, demonstrating that RPAs containing the single OB-fold/zinc finger architecture bind with broadly comparable affinity to two OB-fold/zinc finger RPAs. Reducing the salt concentration to 1M KCl induces dimerisation of the protein, which retains its ability to bind DNA. On circular ssDNA, two concentration-dependent binding modes are observed. Conventionally, increased salt concentration adversely affects DNA binding but HvRPA3 does not bind DNA in 0.2M KCl, although multimerisation may occlude the binding site. The single N-terminal OB-fold is competent to bind DNA in the absence of the C-terminal zinc finger, albeit with reduced affinity. This study represents the first quantitative characterisation of DNA binding in a halophilic protein in extreme salt concentrations

Beads, boats and switches: making things happen with molecular photoswitches

In this paper we present recent results obtained with a stimulus-responsive materials based on the photo-switchable behaviour exhibited by spiro-cyclic derivatives. Our results suggest that these highly novel materials offer unique capabilities hitherto inaccessible using conventional materials. In particular, we will focus on photocontrolled guest binding and release, inherent signalling of status, photo-actuation and solvent driven motion of small structures as examples of the fascinating behaviour of these exceptional materials

Connectionist Inference Models

The performance of symbolic inference tasks has long been a challenge to connectionists. In this paper, we present an extended survey of this area. Existing connectionist inference systems are reviewed, with particular reference to how they perform variable binding and rule-based reasoning, and whether they involve distributed or localist representations. The benefits and disadvantages of different representations and systems are outlined, and conclusions drawn regarding the capabilities of connectionist inference systems when compared with symbolic inference systems or when used for cognitive modeling

The neck region of the C-type lectin DC-SIGN regulates its surface spatiotemporal organization and virus-binding capacity on antigen presenting cells

The C-type lectin DC-SIGN expressed on dendritic cells (DCs) facilitates capture and internalization of a plethora of different pathogens. Although it is known that DC-SIGN organizes in nanoclusters at the surface of DCs, the molecular mechanisms responsible for this well defined nanopatterning and role in viral binding remain enigmatic. By combining biochemical and advanced biophysical techniques, including optical superresolution and single particle tracking, we demonstrate that DC-SIGN intrinsic nanoclustering strictly depends on its molecular structure. DC-SIGN nanoclusters exhibited free, Brownian diffusion on the cell membrane. Truncation of the extracellular neck region, known to abrogate tetramerization, significantly reduced nanoclustering and concomitantly increased lateral diffusion. Importantly, DC-SIGN nanocluster dissolution exclusively compromised binding to nanoscale size pathogens. Monte Carlo simulations revealed that heterogeneity on nanocluster density and spatial distribution confers broader binding capabilities to DC-SIGN. As such, our results underscore a direct relationship between spatial nanopatterning, driven by intermolecular interactions between the neck regions, and receptor diffusion to provide DC-SIGN with the exquisite ability to dock pathogens at the virus length scale. Insight into how virus receptors are organized prior to virus binding and how they assemble into functional platforms for virus docking is helpful to develop novel strategies to prevent virus entry and infectio

Designing Topological Bands in Reciprocal Space

Motivated by new capabilities to realise artificial gauge fields in ultracold atomic systems, and by their potential to access correlated topological phases in lattice systems, we present a new strategy for designing topologically non-trivial band structures. Our approach is simple and direct: it amounts to considering tight-binding models directly in reciprocal space. These models naturally cause atoms to experience highly uniform magnetic flux density and lead to topological bands with very narrow dispersion, without fine-tuning of parameters. Further, our construction immediately yields instances of optical Chern lattices, as well as band structures of higher Chern number, |C|>1

Beyond state-centrism: international law and non-state actors in cyberspace

Classically, States and non-State actors were differentiated not only by disparities in legal status but also by significant imbalances in resources and capabilities. Not surprisingly, international law developed a State-centric bias to account for these imbalances. Cyberspace and cyber operations, however, have closed a number of formerly significant gaps between States’ and non-State actors’ abilities to compromise international peace and security. In fact, some non-State actors now match, if not exceed, the cyber capabilities of many States in this respect. Where public international law had long proved chiefly relevant to States’ interactions with other States, cyber operations by non-State actors increase the frequency with which public international law provides relevant and binding legal rules. This article surveys existing public international law for norms relevant to the cyber interactions of cyber-empowered States and non-State actors. Specifically, the article illustrates how the principles of sovereignty, State responsibility and the jus ad bellum are particularly relevant to States engaged in struggles with non-State actors for security and supremacy in cyberspace

Environment sensing and response mediated by ABC transporters

<p>Abstract</p> <p>Background</p> <p>Transporter proteins are one of an organism’s primary interfaces with the environment. The expressed set of transporters mediates cellular metabolic capabilities and influences signal transduction pathways and regulatory networks. The functional annotation of most transporters is currently limited to general classification into families. The development of capabilities to map ligands with specific transporters would improve our knowledge of the function of these proteins, improve the annotation of related genomes, and facilitate predictions for their role in cellular responses to environmental changes.</p> <p>Results</p> <p>To improve the utility of the functional annotation for ABC transporters, we expressed and purified the set of solute binding proteins from <it>Rhodopseudomonas palustris</it> and characterized their ligand-binding specificity. Our approach utilized ligand libraries consisting of environmental and cellular metabolic compounds, and fluorescence thermal shift based high throughput ligand binding screens. This process resulted in the identification of specific binding ligands for approximately 64% of the purified and screened proteins. The collection of binding ligands is representative of common functionalities associated with many bacterial organisms as well as specific capabilities linked to the ecological niche occupied by <it>R. palustris</it>.</p> <p>Conclusion</p> <p>The functional screen identified specific ligands that bound to ABC transporter periplasmic binding subunits from <it>R. palustris</it>. These assignments provide unique insight for the metabolic capabilities of this organism and are consistent with the ecological niche of strain isolation. This functional insight can be used to improve the annotation of related organisms and provides a route to evaluate the evolution of this important and diverse group of transporter proteins.</p

Mapping of the Epstein-Barr virus and C3dg binding sites to a common domain on complement receptor type 2.

Complement receptor type 2 (CR2;CD21), a member of the superfamily of proteins containing short consensus repeats (SCRs), is the B cell receptor for both the gp350/220 envelope protein of Epstein-Barr virus (EBV), and for the C3dg protein of complement. By analysis of CR2 deletion mutants and chimeras formed with CR1 (CD35) we determined that of the 15 SCRs in CR2, the NH2-terminal two SCRs are necessary and sufficient to bind both gp350/220 and C3dg with affinities equivalent to those of the wild-type receptor. The epitope for OKB-7, a mAb that blocks binding of both EBV and C3dg and shares with these ligands B cell-activating capabilities, also requires both SCR-1 and SCR-2, whereas mAbs lacking these functions bind to other SCRs. Thus, EBV, a polyclonal activator of B cells, has selected a site that is proximate or identical to the natural ligand binding site in CR2, perhaps reflecting the relative immutability of that site as well as its signal transducing function