Interactions between flagellar and type III secretion proteins in <Emphasis Type="Italic">Chlamydia pneumoniae</Emphasis>

Background  

Flagellar secretion systems are utilized by a wide variety of bacteria to construct the flagellum, a conserved apparatus that allows for migration towards non-hostile, nutrient rich environments. Chlamydia pneumoniae is an obligate, intracellular pathogen whose genome contains at least three orthologs of flagellar proteins, namely FliI, FlhA and FliF, but the role of these proteins remains unknown.     

Development of an improved <Emphasis Type="Italic">Pseudoalteromonas haloplanktis</Emphasis> TAC125 strain for recombinant protein secretion at low temperature

Background  

In a previous paper, we reported the accomplishment of a cold gene-expression system for the recombinant secretion of heterologous proteins in Pseudoalteromonas haloplanktis TAC125. This system makes use of the psychrophilic α-amylase from P. haloplanktis TAB23 as secretion carrier, and allows an effective extra-cellular addressing of recombinant proteins. However, Pseudoalteromonales are reported to secrete a wide range of extra-cellular proteases. This feature works against the efficiency of the cold-adapted secretion system, because of the proteolytic degradation of recombinant products. The aim of this study is the construction of a P. haloplanktis TAC125 mutant strain with reduced extra-cellular proteolytic activity.     

AMYPdb: A database dedicated to amyloid precursor proteins

Background  

Misfolding and aggregation of proteins into ordered fibrillar structures is associated with a number of severe pathologies, including Alzheimer's disease, prion diseases, and type II diabetes. The rapid accumulation of knowledge about the sequences and structures of these proteins allows using of in silico methods to investigate the molecular mechanisms of their abnormal conformational changes and assembly. However, such an approach requires the collection of accurate data, which are inconveniently dispersed among several generalist databases.     

A novel Ecotin-Ubiquitin-Tag (ECUT) for efficient, soluble peptide production in the periplasm of Escherichia coli

Background  

Many protocols for recombinant production of peptides and proteins include secretion into the periplasmic space of Escherichia coli, as they may not properly fold in the cytoplasm. If a signal peptide is not sufficient for translocation, a larger secretion moiety can instead be fused to the gene of interest. However, due to the covalent linkage of the proteins, a protease recognition site needs to be introduced in between, altering the N-terminus of the product. In the current study, we combined the ubiquitin fusion technology, which allows production of authentic peptides and proteins, with secretion by the perpiplasmic protease inhibitor ecotin.     

Implementation of force distribution analysis for molecular dynamics simulations

Background  

The way mechanical stress is distributed inside and propagated by proteins and other biopolymers largely defines their function. Yet, determining the network of interactions propagating internal strain remains a challenge for both, experiment and theory. Based on molecular dynamics simulations, we developed force distribution analysis (FDA), a method that allows visualizing strain propagation in macromolecules.     

ProCKSI: a decision support system for Protein (Structure) Comparison,Knowledge, Similarity and Information

Background  

We introduce the decision support system for Protein (Structure) Comparison, Knowledge, Similarity and Information (ProCKSI). ProCKSI integrates various protein similarity measures through an easy to use interface that allows the comparison of multiple proteins simultaneously. It employs the Universal Similarity Metric (USM), the Maximum Contact Map Overlap (MaxCMO) of protein structures and other external methods such as the DaliLite and the TM-align methods, the Combinatorial Extension (CE) of the optimal path, and the FAST Align and Search Tool (FAST). Additionally, ProCKSI allows the user to upload a user-defined similarity matrix supplementing the methods mentioned, and computes a similarity consensus in order to provide a rich, integrated, multicriteria view of large datasets of protein structures.     

Bioinformatic identification of novel regulatory DNA sequence motifs in <Emphasis Type="Italic">Streptomyces coelicolor</Emphasis>

Background  

Streptomyces coelicolor is a bacterium with a vast repertoire of metabolic functions and complex systems of cellular development. Its genome sequence is rich in genes that encode regulatory proteins to control these processes in response to its changing environment. We wished to apply a recently published bioinformatic method for identifying novel regulatory sequence signals to gain new insights into regulation in S. coelicolor.     

<Emphasis Type="Italic">Yersinia enterocolitica</Emphasis> type III secretion: Evidence for the ability to transport proteins that are folded prior to secretion

Background  

Pathogenic Yersinia species (Y. enterocolitica, Y. pestis, Y. pseudotuberculosis) share a type three secretion system (TTSS) which allows translocation of effector proteins (called Yops) into host cells. It is believed that proteins are delivered through a hollow needle with an inner diameter of 2–3 nm. Thus transport seems to require substrates which are essentially unfolded. Recent work from different groups suggests that the Yersinia TTSS cannot accommodate substrates which are folded prior to secretion. It was suggested that folding is prevented either by co-translational secretion or by the assistance of specific Yop chaperones (called Sycs).     

Chemically defined medium and <Emphasis Type="Italic">Caenorhabditis elegans</Emphasis>

Background  

C. elegans has been established as a powerful genetic system. Use of a chemically defined medium (C. elegans Maintenance Medium (CeMM)) now allows standardization and systematic manipulation of the nutrients that animals receive. Liquid cultivation allows automated culturing and experimentation and should be of use in large-scale growth and screening of animals.     

Functional characterization of <Emphasis Type="Italic">Arabidopsis thaliana</Emphasis> transthyretin-like protein

Background  

Arabidopsis thaliana transthyretin-like (TTL) protein is a potential substrate in the brassinosteroid signalling cascade, having a role that moderates plant growth. Moreover, sequence homology revealed two sequence domains similar to 2-oxo-4-hydroxy-4-carboxy-5-ureidoimidazoline (OHCU) decarboxylase (N-terminal domain) and 5-hydroxyisourate (5-HIU) hydrolase (C-terminal domain). TTL is a member of the transthyretin-related protein family (TRP), which comprises a number of proteins with sequence homology to transthyretin (TTR) and the characteristic C-terminal sequence motif Tyr-Arg-Gly-Ser. TRPs are single domain proteins that form tetrameric structures with 5-HIU hydrolase activity. Experimental evidence is fundamental for knowing if TTL is a tetrameric protein, formed by the association of the 5-HIU hydrolase domains and, in this case, if the structural arrangement allows for OHCU decarboxylase activity. This work reports about the biochemical and functional characterization of TTL.     

Improved mycobacterial protein production using a <Emphasis Type="Italic">Mycobacterium smegmatis groEL1ΔC</Emphasis>expression strain

Background  

The non-pathogenic bacterium Mycobacterium smegmatis is widely used as a near-native expression host for the purification of Mycobacterium tuberculosis proteins. Unfortunately, the Hsp60 chaperone GroEL1, which is relatively highly expressed, is often co-purified with polyhistidine-tagged recombinant proteins as a major contaminant when using this expression system. This is likely due to a histidine-rich C-terminus in GroEL1.     

Natural transformation of <Emphasis Type="Italic">Vibrio cholerae</Emphasis> as a tool - Optimizing the procedure

Background  

Vibrio cholerae gains natural competence upon growth on chitin. This allows the organism to take up free DNA from the environment and to incorporate it into its genome by homologous recombination.     

Rapid detection of viable Legionella pneumophila in tap water by a qPCR and RT‐PCR‐based method

Aims

A molecular method for a rapid detection of viable Legionella pneumophila of all serogroups in tap water samples was developed as an alternative to the reference method (ISO). Legionellae are responsible for Legionnaires’ disease, a severe pneumonia in humans with high lethality.

Methods and Results

The developed method is based on a nutritional stimulation and detection of an increase in precursor 16S rRNA as an indicator for viability. For quantification, DNA was detected by qPCR. This method was compared to the ISO method using water samples obtained from public sports facilities in Switzerland. The sensitivity and specificity were 91 and 97%, respectively, when testing samples for compliance with a microbiological criterion of 1000 cell equivalents per l.

Conclusion

The new method is sensitive and specific for Leg. pneumophila and allows results to be obtained within 8 h upon arrival, compared to one week or more by the ISO method.

Significance and Impact of the Study

The method represents a useful tool for a rapid detection of viable Leg. pneumophila of all serogroups in water by molecular biology. It can be used as an alternative to the ISO method for official water analysis for legionellae and particularly when a short test time is required.     

Evolution of the multifaceted eukaryotic akirin gene family

Background  

Akirins are nuclear proteins that form part of an innate immune response pathway conserved in Drosophila and mice. This studies aim was to characterise the evolution of akirin gene structure and protein function in the eukaryotes.     

Biotinylated-sortase self-cleavage purification (BISOP) method for cell-free produced proteins

Background  

Technology used for the purification of recombinant proteins is a key issue for the biochemical and structural analyses of proteins. In general, affinity tags, such as glutathione-S-transferase or six-histidines, are used to purify recombinant proteins. Since such affinity tags often interfere negatively with the structural and functional analyses of proteins, they are usually removed by treatment with proteases. Previously, Dr. H. Mao reported self-cleavage purification of a target protein by fusing the sortase protein to its N-terminal end, and subsequently obtained tag-free recombinant protein following expression in Escherichia coli. This method, however, is yet to be applied to the cell-free based protein production.     

Toward a solution of allocation in life cycle inventories: the use of least-squares techniques

Purpose  

The matrix method for the solution of the so-called inventory problem in LCA generally determines the inventory vector related to a specific system of processes by solving a system of linear equations. The paper proposes a new approach to deal with systems characterized by a rectangular (and thus non-invertible) coefficients matrix. The approach, based on the application of regression techniques, allows solving the system without using computational expedients such as the allocation procedure.     

Mapping sequences by parts

Background:  

We present the N-map method, a pairwise and asymmetrical approach which allows us to compare sequences by taking into account evolutionary events that produce shuffled, reversed or repeated elements. Basically, the optimal N-map of a sequence s over a sequence t is the best way of partitioning the first sequence into N parts and placing them, possibly complementary reversed, over the second sequence in order to maximize the sum of their gapless alignment scores.