The GTPase function of YvcJ and its subcellular relocalization are dependent on growth conditions in Bacillus subtilis

We have recently shown that the Bacillus subtilis GTPase YvcJ is involved in the phosphorylation of an unidentified cellular component and that the deletion of yvcJ induced a decrease in competence efficiency. In this paper, we report that growth conditions influence both the YvcJ-dependent phosphorylation event and the localization of this protein. More precisely, we have observed that YvcJ can be localized in the cell either as a helical-like pattern or as foci close to the poles and the septa depending on growth phase and on growth medium. In addition, we show that the mutation of the catalytic lysine residue (K22) located in the Walker A motif of YvcJ, and necessary for its GTPase activity, induces a decrease in competence efficiency similar to that observed for the yvcJ null mutant. This mutation also inhibits the YvcJ-dependent phosphorylation event. Furthermore, a phylogenetic analysis of the YvcJ homologues shows that this protein is ancient in Bacteria (being possibly present in their last common ancestor) and has been conserved in a number of major bacterial phyla, suggesting that this protein has an important function in this domain of life. To sum up, even if the precise cellular role of this ancient protein remains unknown, our data show that the GTPase activity of B. subtilis YvcJ and its function in the phosphorylation of a cellular component are influenced by the growth conditions, and are important for the effect of YvcJ on competence efficiency.     

Actinoplanes teichomyceticus ATCC 31121 as a cell factory for producing teicoplanin

Background

Single-domain antibody fragments possess structural features, such as a small dimension, an elevated stability, and the singularity of recognizing epitopes non-accessible for conventional antibodies that make them interesting for several research and biotechnological applications.

Results

The discovery of the single-domain antibody's potentials has stimulated their use in an increasing variety of fields. The rapid accumulation of articles describing new applications and further developments of established approaches has made it, therefore, necessary to update the previous reviews with a new and more complete summary of the topic.

Conclusions

Beside the necessary task of updating, this work analyses in detail some applicative aspects of the single-domain antibodies that have been overseen in the past, such as their efficacy in affinity chromatography, as co-crystallization chaperones, protein aggregation controllers, enzyme activity tuners, and the specificities of the unconventional single-domain fragments.     

Combinatorial analysis and algorithms for quasispecies reconstruction using next-generation sequencing

Background  

Next-generation sequencing (NGS) offers a unique opportunity for high-throughput genomics and has potential to replace Sanger sequencing in many fields, including de-novo sequencing, re-sequencing, meta-genomics, and characterisation of infectious pathogens, such as viral quasispecies. Although methodologies and software for whole genome assembly and genome variation analysis have been developed and refined for NGS data, reconstructing a viral quasispecies using NGS data remains a challenge. This application would be useful for analysing intra-host evolutionary pathways in relation to immune responses and antiretroviral therapy exposures. Here we introduce a set of formulae for the combinatorial analysis of a quasispecies, given a NGS re-sequencing experiment and an algorithm for quasispecies reconstruction. We require that sequenced fragments are aligned against a reference genome, and that the reference genome is partitioned into a set of sliding windows (amplicons). The reconstruction algorithm is based on combinations of multinomial distributions and is designed to minimise the reconstruction of false variants, called in-silico recombinants.     

Prior parity positively regulates learning and memory in young and middle-aged rats

Reproductive experience in female rats modifies acquired behaviors, induces long-lasting functional neuroadaptations and can also modify spatial learning and memory. The present study supports and expands this knowledge base by employing the Morris water maze, which measures spatial memory. Age-matched young adult (YNG) nulliparous (NULL; nonmated) and primiparous (PRIM; one pregnancy and lactation) female rats were tested 15 d after the litter's weaning. In addition, corresponding middle-aged (AGD) PRIM (mated in young adulthood so that pregnancy, parturition, and lactation occurred at the same age as in YNG PRIM) and NULL female rats were tested at 18 mo of age. Behavioral evaluation included: 1) acquisition of reference memory (platform location was fixed for 14 to 19 d of testing); 2) retrieval of this information associated with extinction of the acquired response (probe test involving removal of the platform 24 h after the last training session); and 3) performance in a working memory version of the task (platform presented in a novel location every day for 13 d, and maintained in a fixed location within each day). YNG PRIM outperformed NULL rats and showed different behavioral strategies. These results may be related to changes in locomotor, mnemonic, and cognitive processes. In addition, YNG PRIM exhibited less anxiety-like behavior. Compared with YNG rats, AGD rats showed less behavioral flexibility but stronger memory consolidation. These data, which were obtained by using a well-documented spatial task, demonstrate long lasting modifications of behavioral strategies in both YNG and AGD rats associated with a single reproductive experience.     

TGR5 activation inhibits atherosclerosis by reducing macrophage inflammation and lipid loading

The G protein-coupled receptor TGR5 has been identified as an important component of the bile acid signaling network, and its activation has been linked to enhanced energy expenditure and improved glycemic control. Here, we demonstrate that activation of TGR5 in macrophages by 6α-ethyl-23(S)-methylcholic acid (6-EMCA, INT-777), a semisynthetic BA, inhibits proinflammatory cytokine production, an effect mediated by TGR5-induced cAMP signaling and subsequent NF-κB inhibition. TGR5 activation attenuated atherosclerosis in Ldlr^−/−Tgr5^+/+ mice but not in Ldlr^−/−Tgr5^−/− double-knockout mice. The inhibition of lesion formation was associated with decreased intraplaque inflammation and less plaque macrophage content. Furthermore, Ldlr^−/− animals transplanted with Tgr5^−/− bone marrow did not show an inhibition of atherosclerosis by INT-777, further establishing an important role of leukocytes in INT-777-mediated inhibition of vascular lesion formation. Taken together, these data attribute a significant immune modulating function to TGR5 activation in the prevention of atherosclerosis, an important facet of the metabolic syndrome.     

Leviserpin: A Serine Peptidase Inhibitor (Serpin) from the Sugarcane Weevil <Emphasis Type="Italic">Sphenophorus levis</Emphasis>

Serine peptidase inhibitors (serpins) form a superfamily of proteins covering abroad spectrum of different biological functions. Here we describe the inhibitory characterization of leviserpin, the first serpin from the sugar cane weevil Sphenophorus levis. Leviserpin was able to inhibit bovine trypsin by the formation of the covalent complex serpin-peptidase, demonstrated by SDS–PAGE and mass spectroscopy analysis. We also have determined the cleavage site at the reactive center loop, by the analysis of the polypeptides released from de C-terminus of leviserpin. Moreover we investigated the mRNA expression of leviserpin in different stages of S. levis development. Thus the specificity of leviserpin, in addition with its mRNA coding being transcribed through all lifecycle of the insect, can suggest a possible role in defense mechanism by regulating the action of prophenoloxidase (proPO) activating enzyme.     

Automatic network fingerprinting through single-node motifs

Complex networks have been characterised by their specific connectivity patterns (network motifs), but their building blocks can also be identified and described by node-motifs-a combination of local network features. One technique to identify single node-motifs has been presented by Costa et al. (L. D. F. Costa, F. A. Rodrigues, C. C. Hilgetag, and M. Kaiser, Europhys. Lett., 87, 1, 2009). Here, we first suggest improvements to the method including how its parameters can be determined automatically. Such automatic routines make high-throughput studies of many networks feasible. Second, the new routines are validated in different network-series. Third, we provide an example of how the method can be used to analyse network time-series. In conclusion, we provide a robust method for systematically discovering and classifying characteristic nodes of a network. In contrast to classical motif analysis, our approach can identify individual components (here: nodes) that are specific to a network. Such special nodes, as hubs before, might be found to play critical roles in real-world networks.