Tandem fusion of hepatitis B core antigen allows assembly of virus-like particles in bacteria and plants with enhanced capacity to accommodate foreign proteins

The core protein of the hepatitis B virus, HBcAg, assembles into highly immunogenic viruslike particles (HBc VLPs) when expressed in a variety of heterologous systems. Specifically, the major insertion region (MIR) on the HBcAg protein allows the insertion of foreign sequences, which are then exposed on the tips of surface spike structures on the outside of the assembled particle. Here, we present a novel strategy which aids the display of whole proteins on the surface of HBc particles. This strategy, named tandem core, is based on the production of the HBcAg dimer as a single polypeptide chain by tandem fusion of two HBcAg open reading frames. This allows the insertion of large heterologous sequences in only one of the two MIRs in each spike, without compromising VLP formation. We present the use of tandem core technology in both plant and bacterial expression systems. The results show that tandem core particles can be produced with unmodified MIRs, or with one MIR in each tandem dimer modified to contain the entire sequence of GFP or of a camelid nanobody. Both inserted proteins are correctly folded and the nanobody fused to the surface of the tandem core particle (which we name tandibody) retains the ability to bind to its cognate antigen. This technology paves the way for the display of natively folded proteins on the surface of HBc particles either through direct fusion or through non-covalent attachment via a nanobody

A Roadmap for HEP Software and Computing R&D for the 2020s

Particle physics has an ambitious and broad experimental programme for the coming decades. This programme requires large investments in detector hardware, either to build new facilities and experiments, or to upgrade existing ones. Similarly, it requires commensurate investment in the R&D of software to acquire, manage, process, and analyse the shear amounts of data to be recorded. In planning for the HL-LHC in particular, it is critical that all of the collaborating stakeholders agree on the software goals and priorities, and that the efforts complement each other. In this spirit, this white paper describes the R&D activities required to prepare for this software upgrade.Peer reviewe

次世代メニーコアプロセッサのオンチップメモリアーキテクチャに対する性能評価の高速化技術

主1-参1情報知能工

Work-in-Progress Abstract: The impact of the period variation on execution time distributions of programs

International audienceDesigners of embedded real-time systems derive, in general, their time parameters such as activation periods from those of sensors or actuators. By designers, we mean the team in charge of conceiving embedded real-time systems. This team includes Control Theory designers and Computer Science designers. Within this paper we present the point of view of Computer Science designers, while the periods proposed by Control Theory designers are supposed robust with respect to the physical behavior of the system. The execution times are, then, estimated by studying statically the programs structure or dynamically the programs execution. In some cases, both activation periods and execution times depend on a sensor information. For instance, they depend on the angular speed of wheels within an automotive embedded real-time system and such systems follow a rate-dependent model. Elastic tasks is another model, where one may consider execution time variation depending on the selected period. Within this paper, we are interested in describing statistically the relationship between activation periods and execution times of programs. More precisely, we study the impact of the period variation on the distributions of the execution times. To illustrate our preliminary results, we consider, as case study, the set of programs executing the autopilot of an open-source PX4 drone

Work-in-Progress Abstract: WKS, a local unsupervised statistical algorithm for the detection of transitions in timing analysis

International audienceThe increased complexity of programs and pro-cessors is an important challenge that the embedded real-time systems community faces today, as it implies substancial timing variability. Processor features like pipelines or communication buses are not always completely described, while black-box programs integrated by third parties are hidden for IP reasons. This situation explains the use of statistical approaches to study the timing variability of programs. Most existing work is concentrated on the guarantees provided by positive answers to statistical tests, while our current work concerns potential algorithms based on the negative answers to these tests and their impact on the timing analysis. We introduce here one such algorithm, the Walking Kolmogorov-Smirnov test (WKS)

Work in Progress: KDBench - towards open source benchmarks for measurement-based multicore WCET estimators

International audienceThe real-time systems community is facing the lack of benchmarks adapted to measurement-based worst-case execution time (WCET) estimators. We provide in this paper first steps towards such benchmarks by proposing them for single core microcontrollers, while we leave as future work the migration to multicore microcontrollers. The considered benchmarks are the programs of an open source drone autopilot. We conclude the paper by underlining the main difficulties of such migration

Cryo-EM analysis of plant-produced CoHe-GFPL VLPs.

<p>a) Particles were flash-frozen in vitreous ice, then subjected to cryo-electron microscopy. Class averages were obtained from 441 individual particles using EMAN software. The expanded view (lower right) is of an average of all images used. b) 3D reconstruction of the particles using icosahedral symmetry, superimposed on the He map as shown in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0120751#pone.0120751.g003" target="_blank">Fig. 3</a>. The CoHe-GFPL map is coloured red-to-blue from the centre of the volume towards its edge; the He map is shown in grey.</p

Cryo-EM of plant-produced τGFP bound with GFP.

<p>a) Class averages computed using Relion of the τGFP particles. b) A 3D reconstruction (resolution estimate 25Å using the “gold standard” cross-FSC at cutoff 0.143) coloured by distance from the centre of the particle (red to blue). The map is shown viewed down a 5-fold axis with the He reconstruction on which the construct was based fitted within (grey surface). The projecting spikes represent density arising from the bound nanobody and GFP but do not occupy every position expected, instead appearing as an average of the density present with the highest intensity at the 2-fold (pseudo- 6-fold) axes and also at the 5-fold axis. These spikes are to some extent artefacts of the icosahedral symmetry imposed on the maps, but are reflected in the spikes also shown in the unaveraged class averages shown in a).</p

τGFP expressed in plants forms VLPs.

<p>a) Predicted structure of the τGFP tandibody protein (Swiss-Prot model): green: core 1, yellow: core 2, pink: anti-GFP nanobody, red: linkers. b) Western blot of crude plant extracts. C: empty vector control, τGFP: tandem HBcAg construct with anti-GFP VHH in the core 2 MIR, μGFP: monomeric HBcAg containing anti-GFP VHH in the MIR. The 39 kDa band found in all plant extracts is non-specific. c) Electron micrograph of plant-produced τGFP particles purified by sucrose cushion. Scale bar 100 nm.</p