Combinatorial impact of Sec signal peptides from Bacillus subtilis and bioprocess conditions on heterologous cutinase secretion by Corynebacterium glutamicum

The impact of Sec signal peptides (SPs) from Bacillus subtilis in combination with isopropyl-β- d -1-thiogalactopyranoside concentration and feeding profile was investigated for heterologous protein secretion performance by Corynebacterium glutamicum using cutinase as model enzyme. Based on a comprehensive data set of about 150 bench-scale bioreactor cultivations in fed-batch mode and choosing the cutinase yield as objective, it was shown that relative secretion performance for bioprocesses remains very similar, irrespective of the applied SP enabling Sec-mediated cutinase secretion. However, to achieve the maximal absolute cutinase yield, careful adjustment of bioprocess conditions was found to be necessary. A model-based, two-step multiple regression approach resembled the collected data in a comprehensive way. The corresponding results suggest that the choice of the heterologous Sec SP and its interaction with the adjusted exponential feeding profile is highly relevant to maximize absolute cutinase yield in this study. For example, the impact of Sec SP is high at low growth rates and low at high growth rates. However, promising Sec SPs could be inferred from less complex batch cultivations. The extensive data were also evaluated in terms of cutinase productivity, highlighting the well-known trade-off between yield and productivity in bioprocess development in detail. Conclusively, only the right combination of target protein, Sec SP, and bioprocess conditions is the key to success.     

Optimizing MPI collective communication by orthogonal structures

MPI collective communication operations to distribute or gather data are used for many parallel applications from scientific computing, but they may lead to scalability problems since their execution times increase with the number of participating processors. In this article, we show how the execution time of collective communication operations can be improved significantly by an internal restructuring based on orthogonal processor structures with two or more levels. The execution time of operations like MPI_Bcast() or MPI_Allgather() can be reduced by 40% and 70% on a dual Xeon cluster and a Beowulf cluster with single-processor nodes. But also on a Cray T3E a significant performance improvement can be obtained by a careful selection of the processor structure. The use of these optimized communication operations can reduce the execution time of data parallel implementations of complex application programs significantly without requiring any other change of the computation and communication structure. We present runtime functions for the modeling of two-phase realizations and verify that these runtime functions can predict the execution time both for communication operations in isolation and in the context of application programs.     

Inferring modules from human protein interactome classes

Background  

The integration of protein-protein interaction networks derived from high-throughput screening approaches and complementary sources is a key topic in systems biology. Although integration of protein interaction data is conventionally performed, the effects of this procedure on the result of network analyses has not been examined yet. In particular, in order to optimize the fusion of heterogeneous interaction datasets, it is crucial to consider not only their degree of coverage and accuracy, but also their mutual dependencies and additional salient features.     

Targeting antibody responses to the membrane proximal external region of the envelope glycoprotein of human immunodeficiency virus

Although human immunodeficiency type 1 (HIV-1) infection induces strong antibody responses to the viral envelope glycoprotein (Env) only a few of these antibodies possess the capacity to neutralize a broad range of strains. The induction of such antibodies represents an important goal in the development of a preventive vaccine against the infection. Among the broadly neutralizing monoclonal antibodies discovered so far, three (2F5, Z13 and 4E10) target the short and hidden membrane proximal external region (MPER) of the gp41 transmembrane protein. Antibody responses to MPER are rarely observed in HIV-infected individuals or after immunization with Env immunogens. To initiate antibody responses to MPER in its membrane-embedded native conformation, we generated expression plasmids encoding the membrane-anchored ectodomain of gp41 with N-terminal deletions of various sizes. Following transfection of these plasmids, the MPER domains are displayed on the cell surface and incorporated into HIV virus like particles (VLP). Transfected cells displaying MPER mutants bound as efficiently to both 2F5 and 4E10 as cells transfected with a plasmid encoding full-length Env. Mice immunized with VLPs containing the MPER mutants produced MPER-specific antibodies, the levels of which could be increased by the trimerization of the displayed proteins as well as by a DNA prime-VLP boost immunization strategy. Although 2F5 competed for binding to MPER with antibodies in sera of some of the immunized mice, neutralizing activity could not be detected. Whether this is due to inefficient binding of the induced antibodies to MPER in the context of wild type Env or whether the overall MPER-specific antibody response induced by the MPER display mutants is too low to reveal neutralizing activity, remains to be determined.     

DOGS: reaction-driven de novo design of bioactive compounds

We present a computational method for the reaction-based de novo design of drug-like molecules. The software DOGS (Design of Genuine Structures) features a ligand-based strategy for automated ‘in silico’ assembly of potentially novel bioactive compounds. The quality of the designed compounds is assessed by a graph kernel method measuring their similarity to known bioactive reference ligands in terms of structural and pharmacophoric features. We implemented a deterministic compound construction procedure that explicitly considers compound synthesizability, based on a compilation of 25''144 readily available synthetic building blocks and 58 established reaction principles. This enables the software to suggest a synthesis route for each designed compound. Two prospective case studies are presented together with details on the algorithm and its implementation. De novo designed ligand candidates for the human histamine H₄ receptor and γ-secretase were synthesized as suggested by the software. The computational approach proved to be suitable for scaffold-hopping from known ligands to novel chemotypes, and for generating bioactive molecules with drug-like properties.     

Estimation of Density-Dependent Mortality of Juvenile Bivalves in the Wadden Sea

We investigated density-dependent mortality within the early months of life of the bivalves Macoma balthica (Baltic tellin) and Cerastoderma edule (common cockle) in the Wadden Sea. Mortality is thought to be density-dependent in juvenile bivalves, because there is no proportional relationship between the size of the reproductive adult stocks and the numbers of recruits for both species. It is not known however, when exactly density dependence in the pre-recruitment phase occurs and how prevalent it is. The magnitude of recruitment determines year class strength in bivalves. Thus, understanding pre-recruit mortality will improve the understanding of population dynamics. We analyzed count data from three years of temporal sampling during the first months after bivalve settlement at ten transects in the Sylt-Rømø-Bay in the northern German Wadden Sea. Analyses of density dependence are sensitive to bias through measurement error. Measurement error was estimated by bootstrapping, and residual deviances were adjusted by adding process error. With simulations the effect of these two types of error on the estimate of the density-dependent mortality coefficient was investigated. In three out of eight time intervals density dependence was detected for M. balthica, and in zero out of six time intervals for C. edule. Biological or environmental stochastic processes dominated over density dependence at the investigated scale.     

Mycorrhizal suppression and phosphorus addition influence the stability of plant community composition and function in a temperate steppe

Nutrient enrichment can reduce ecosystem stability, typically measured as temporal stability of a single function, e.g. plant productivity. Moreover, nutrient enrichment can alter plant–soil interactions (e.g. mycorrhizal symbiosis) that determine plant community composition and productivity. Thus, it is likely that nutrient enrichment and interactions between plants and their soil communities co-determine the stability in plant community composition and productivity. Yet our understanding as to how nutrient enrichment affects multiple facets of ecosystem stability, such as functional and compositional stability, and the role of above–belowground interactions are still lacking. We tested how mycorrhizal suppression and phosphorus (P) addition influenced multiple facets of ecosystem stability in a three-year field study in a temperate steppe. Here we focused on the functional and compositional stability of plant community; functional stability is the temporal community variance in primary productivity; compositional stability is represented by compositional resistance, turnover, species extinction and invasion. Community variance was partitioned into population variance defined as community productivity weighted average of the species temporal variance in performance, and species synchrony defined as the degree of temporal positive covariation among species. Compared to treatments with mycorrhizal suppression, the intact AM fungal communities reduced community variance in primary productivity by reducing species synchrony at high levels of P addition. Species synchrony and population variance were linearly associated with community variance with the intact AM fungal communities, while these relationships were decoupled or weakened by mycorrhizal suppression. The intact AM fungal communities promoted the compositional resistance of plant communities by reducing compositional turnover, but this effect was suppressed by P addition. P addition increased the number of species extinctions and thus promoted compositional turnover. Our study shows P addition and AM fungal communities can jointly and independently modify the various components of ecosystem stability in terms of plant community productivity and composition.     

Partially-deuterated samples of HET-s(218–289) fibrils: assignment and deuterium isotope effect

Fast magic-angle spinning and partial sample deuteration allows direct detection of ¹H in solid-state NMR, yielding significant gains in mass sensitivity. In order to further analyze the spectra, ¹H detection requires assignment of the ¹H resonances. In this work, resonance assignments of backbone H^N and Hα are presented for HET-s(218–289) fibrils, based on the existing assignment of Cα, Cβ, C’, and N resonances. The samples used are partially deuterated for higher spectral resolution, and the shifts in resonance frequencies of Cα and Cβ due to the deuterium isotope effect are investigated. It is shown that the deuterium isotope effect can be estimated and used for assigning resonances of deuterated samples in solid-state NMR, based on known resonances of the protonated protein.     

Between-species correlations in the number of migrants at Ulmeth?chi (Switzerland)

We analysed the number of autumn migrants at a bird ringing station over 41 years in the Jura mountains of Switzerland. For 12 irruptive or potentially irruptive bird species, the correlations between their numbers per year were calculated and the species were clustered accordingly. We found high correlations in the number of migrants between the Coal Tit Periparus ater, Great Tit Parus major and Blue Tit Cyanistes caeruleus. Most correlations of passage number between species pairs changed dramatically over time. Only Blue Tit and Coal Tit showed continuously high correlation in this respect. The variation and changes over time in between-species correlations in the number of migrants needs more attention.     

T cell responses against microsatellite instability-induced frameshift peptides and influence of regulatory T cells in colorectal cancer

High-level microsatellite-unstable (MSI-H) colorectal carcinomas (CRC) represent a distinct subtype of tumors commonly characterized by dense infiltration with cytotoxic T cells, most likely due to expression of MSI-H-related frameshift peptides (FSP). The contribution of FSP and classical antigens like MUC1 and CEA to the cellular immune response against MSI-H CRC had not been analyzed so far. We analyzed tumor-infiltrating and peripheral T cells from MSI-H (n = 4 and n = 14, respectively) and microsatellite-stable (MSS) tumor patients (n = 26 and n = 17) using interferon gamma ELISpot assays. Responses against 4 FSP antigens and peptides derived from MUC1 to CEA were compared with and without depletion of regulatory T cells, and the results were related to the presence of the respective antigens in tumor tissue. Preexisting FSP-specific T cell responses were detected in all (4 out of 4) tumor-infiltrating and in the majority (10 out of 14) of peripheral T cell samples from MSI-H CRC patients, but rarely observed in MSS CRC patients. Preexisting T cell responses in MSI-H CRC patients were significantly more frequently directed against FSP tested in the present study than against peptides derived from classical antigens MUC1 or CEA (p = 0.049). Depletion of regulatory T cells increased the frequency of effector T cell responses specific for MUC1/CEA-derived peptides and, to a lesser extent, T cell responses specific for FSP. Our data suggest that the analyzed FSP may represent an immunologically relevant pool of antigens capable of eliciting antitumoral effector T cell responses.