DNA–DNA kissing complexes as a new tool for the assembly of DNA nanostructures

Kissing-loop annealing of nucleic acids occurs in nature in several viruses and in prokaryotic replication, among other circumstances. Nucleobases of two nucleic acid strands (loops) interact with each other, although the two strands cannot wrap around each other completely because of the adjacent double-stranded regions (stems). In this study, we exploited DNA kissing-loop interaction for nanotechnological application. We functionalized the vertices of DNA tetrahedrons with DNA stem-loop sequences. The complementary loop sequence design allowed the hybridization of different tetrahedrons via kissing-loop interaction, which might be further exploited for nanotechnology applications like cargo transport and logical elements. Importantly, we were able to manipulate the stability of those kissing-loop complexes based on the choice and concentration of cations, the temperature and the number of complementary loops per tetrahedron either at the same or at different vertices. Moreover, variations in loop sequences allowed the characterization of necessary sequences within the loop as well as additional stability control of the kissing complexes. Therefore, the properties of the presented nanostructures make them an important tool for DNA nanotechnology.     

Extracellular nucleases of Bacillus subtilis II. The nucleases as 5′-end-group reagents

The cleavage of the Bacillus subtilis extracellular nucleases was determined with DNA substrates differing in the number of phosphates on the 5′-terminus. The scission consistently is at the terminal phosphodiester bond thereby releasing mononucleoside 3′-monophosphates (Np), mononucleoside 3′,5′-diphosphates (pNp), and mononucleoside tetraphosphates (pppNp) from 5′-hydroxyl, 5′-phosphoryl and 5′-triphosphoryl terminated substrates, respectively. The recovery of the end-group is stoichiometric, and only mononucleoside monophosphates are released from internal portions of the DNA chain. The degradation mode is the same for the three nucleases studied: nucleases Bs-IA, Bs-IB and Bs-II. Therefore, these nucleases are excellent 5′-end-group reagents.     

Artificial feedback for invasive brain–computer interfaces

During the last two decades, considerable progress has been made in the studies of brain–computer interfaces (BCIs)—devices in which motor signals from the brain are registered by multi-electrode arrays and transformed into commands for artificial actuators such as cursors and robotic devices. This review is focused on one problem. Voluntary motor control is based on neurophysiological processes, which strongly depend on the afferent innervation of skin, muscles, and joints. Thus, invasive BCI has to be based on a bidirectional system in which motor control signals are registered by multichannel microelectrodes implanted in motor areas, whereas tactile, proprioceptive, and other useful signals are transported back to the brain through spatiotemporal patterns of intracortical microstimulation (ICMS) delivered to sensory areas. In general, the studies of invasive BCIs have advanced in several directions. The progress of BCIs with artificial sensory feedback will not only help patients, but will also expand base knowledge in the field of human cortical functions.     

Structure-based method for analyzing protein–protein interfaces

Hydrogen bond, hydrophobic and vdW interactions are the three major non-covalent interactions at protein–protein interfaces. We have developed a method that uses only these properties to describe interactions between proteins, which can qualitatively estimate the individual contribution of each interfacial residue to the binding and gives the results in a graphic display way. This method has been applied to analyze alanine mutation data at protein–protein interfaces. A dataset containing 13 protein–protein complexes with 250 alanine mutations of interfacial residues has been tested. For the 75 hot-spot residues (G1.5 kcal mol^-1), 66 can be predicted correctly with a success rate of 88%. In order to test the tolerance of this method to conformational changes upon binding, we utilize a set of 26 complexes with one or both of their components available in the unbound form. The difference of key residues exported by the program is 11% between the results using complexed proteins and those from unbound ones. As this method gives the characteristics of the binding partner for a particular protein, in-depth studies on protein–protein recognition can be carried out. Furthermore, this method can be used to compare the difference between protein–protein interactions and look for correlated mutation. Figure Key interaction grids at the interface between barnase and barstar. Key interaction grid for barnase and barstar are presented in one figure according to their coordinates. In order to distinguish the two proteins, different icons were assigned. Crosses represent key grids for barstar and dots represent key grids for barnase. The four residues in ball and stick are Asp40 in barstar and Arg83, Arg87, His102 in barnase.     

Using microalgae for remediation of crude petroleum oil–water emulsions

Crude petroleum oil spills are among the most important organic contaminations. While the separated oils accumulated on the surface water are relatively easily removed, the emulsified portions are more difficult to remove and pose significant threats to the environment. Bioremediation using bacteria has proven to be an effective method, but the biomass produced in this case does not have any significant remunerative value. In this work, microalgae were proposed to combine emulsified oil remediation process with the potential of microalgae as a biofuel feedstock, thus enhancing the economic and environmental benefits of the process. A freshwater strain of Chlorella vulgaris was grown in water containing different concentrations of emulsified crude oil at different temperatures. The specific growth rate (μ_max) of the microalgae for each initial oil concentration was determined and was found to increase with the increase in initial oil concentration. For example, at 30°C, the specific growth rate, μ increased from 0.477 to 0.784 per day as the oil concentration increased from 57 to 222 mg/L. At 30°C, the effect of substrate concentration agreed with that of the microalgae growth, whereas at 40°C, the drop in oil concentration decreased with the increase in concentration. The results were fitted to a modified Monod kinetics model that used specific interfacial area as the influential substrate instead of the actual concentration. The results of this study clearly show the potential of using microalgae for emulsified oil remediation at relatively high concentrations.     

An information-theoretic classification of amino acids for the assessment of interfaces in protein–protein docking

Docking represents a versatile and powerful method to predict the geometry of protein–protein complexes. However, despite significant methodical advances, the identification of good docking solutions among a large number of false solutions still remains a difficult task. We have previously demonstrated that the formalism of mutual information (MI) from information theory can be adapted to protein docking, and we have now extended this approach to enhance its robustness and applicability. A large dataset consisting of 22,934 docking decoys derived from 203 different protein–protein complexes was used for an MI-based optimization of reduced amino acid alphabets representing the protein–protein interfaces. This optimization relied on a clustering analysis that allows one to estimate the mutual information of whole amino acid alphabets by considering all structural features simultaneously, rather than by treating them individually. This clustering approach is fast and can be applied in a similar fashion to the generation of reduced alphabets for other biological problems like fold recognition, sequence data mining, or secondary structure prediction. The reduced alphabets derived from the present work were converted into a scoring function for the evaluation of docking solutions, which is available for public use via the web service score-MI: http://score-MI.biochem.uni-erlangen.de     

Melt-in-Mouth Pellets of Fexofenadine Hydrochloride Using Crospovidone as an Extrusion–Spheronisation Aid

Microcrystalline cellulose (MCC) is well established as an extrusion spheronisation aid for the preparation of pellets. Crospovidone (Polyplasdone® XL-10) is compared with microcrystalline cellulose for the preparation of melt-in-mouth pellets. Taste-masked fexofenadine hydrochloride was incorporated in the melt-in-mouth formulation. Crospovidone was found to be well suited as extrusion–spheronisation aid for the preparation of melt-in-mouth pellets. The great advantage of crospovidone is, however, the disintegrating properties of the pellets after only a short time of exposure to liquid. Crospovidone was successfully employed as an extrusion–spheronisation aid to produce melt-in-mouth pellets obviating the need of a traditional extrusion–spheronisation aid, MCC. Dual properties of Crospovidone were explored viz. as an extrusion–spheronisation aid and a disintegrant.     

The protein–protein interactions required for assembly of the Tn3 resolution synapse

The site-specific recombinase Tn3 resolvase initiates DNA strand exchange when two res recombination sites and six resolvase dimers interact to form a synapse. The detailed architecture of this intricate recombination machine remains unclear. We have clarified which of the potential dimer–dimer interactions are required for synapsis and recombination, using a novel complementation strategy that exploits a previously uncharacterized resolvase from Bartonella bacilliformis (“Bart”). Tn3 and Bart resolvases recognize different DNA motifs, via diverged C-terminal domains (CTDs). They also differ substantially at N-terminal domain (NTD) surfaces involved in dimerization and synapse assembly. We designed NTD-CTD hybrid proteins, and hybrid res sites containing both Tn3 and Bart dimer binding sites. Using these components in in vivo assays, we demonstrate that productive synapsis requires a specific “R” interface involving resolvase NTDs at all three dimer-binding sites in res. Synapses containing mixtures of wild-type Tn3 and Bart resolvase NTD dimers are recombination-defective, but activity can be restored by replacing patches of Tn3 resolvase R interface residues with Bart residues, or vice versa. We conclude that the Tn3/Bart family synapse is assembled exclusively by R interactions between resolvase dimers, except for the one special dimer–dimer interaction required for catalysis.     

Structural basis for catalyzed assembly of the Sonic hedgehog–Patched1 signaling complex

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Molecular dynamics simulation approach for the prediction of transmembrane helix–helix heterodimers assembly

Computational methods are useful to identify favorable structures of transmembrane (TM) helix oligomers when experimental data are not available or when they cannot help to interpret helix-helix association. We report here a global search method using molecular dynamics (MD) simulations to predict the structures of transmembrane homo and heterodimers. The present approach is based only on sequence information without any experimental data and is first applied to glycophorin A to validate the protocol and to the HER2-HER3 heterodimer receptor. The method successfully reproduces the experimental structures of the TM domain of glycophorin A (GpA(TM)) with a root mean square deviation of 1.5 A. The search protocol identifies three energetically stable models of the TM domain of HER2-HER3 receptor with favorable helix-helix arrangement, including right-handed and left-handed coiled-coils. The predicted TM structures exhibit the GxxxG-like motif at the dimer interface which is presumed to drive receptor oligomerization. We demonstrate that native structures of TM domain can be predicted without quantitative experimental data. This search protocol could help to predict structures of the TM domain of HER heterodimer family.     

Functional–structural models optimize the placement of foliage units for multiple whole-canopy functions

I present examples of plant functional–structural models (FSMs) that are used to evaluate how foliage units affect whole-canopy functions, and I show that multi-criteria optimization is an effective tool for these models. FSMs produce plant structures through the repeated application of a set of rules for the placement of foliage units. The models are blind (rules are the same regardless of dynamic simulation conditions), sighted (rules change with interference from other foliage units) or self-regulatory (rules change depending on the conditions of the simulation, i.e., internal conditions). In the examples presented, the models are used to optimize plant morphology for one or more measures of plant performance; these measures include movement of materials and associated hydraulic functions, foliage display, light interception and net carbon, mechanical support and stability, and reproductive success. It is consistently found that no morphology is optimal for any single measure of plant performance, and the rules for plant development are not stationary in space and time. In multi-criteria optimization, alternative morphologies are compared against multiple measures of plant performance; these are optimized simultaneously using Pareto optimality, which yields the set of mutually co-dominant solutions not dominated by any other solution. Two solutions are considered to be mutually co-dominant if improvement with respect to one criterion is at the expense of another criterion. I conclude that multi-criteria optimization is an essential tool for the use of FSMs to relate processes at the foliage level to whole-canopy function and to explain the structural diversity of old-growth forests.     

Implications of disease-related mutations at protein–protein interfaces

Protein–protein interfaces have been attracting great attention owing to their critical roles in protein–protein interactions and the fact that human disease-related mutations are generally enriched in them. Recently, substantial research progress has been made in this field, which has significantly promoted the understanding and treatment of various human diseases. For example, many studies have discovered the properties of disease-related mutations. Besides, as more large-scale experimental data become available, various computational approaches have been proposed to advance our understanding of disease mutations from the data. Here, we overview recent advances in characteristics of disease-related mutations at protein–protein interfaces, mutation effects on protein interactions, and investigation of mutations on specific diseases.     

‘Click’ assembly of selective inhibitors for MAO-A

In this Letter, an efficient strategy for the fast construction of 108 compounds library was developed using click chemistry. The fingerprint of inhibitory activity toward MAO-A/B against this library was obtained, and four hit compounds were identified as selective inhibitors toward MAO-A. Docking study was carried out to demonstrate the binding mode between a9 and MAO-A/B, and the result reveals that a9 localized in the ‘aromatic cage’ and oriented to establish π–π stacking interactions with Tyr407, Tyr444 and FAD in MAO-A rather than in MAO-B.     

Structural features of peptoid–peptide hybrids in lipid–water interfaces

The inclusion of peptoid monomers into antimicrobial peptides (AMPs) increases their proteolytic resistance, but introduces conformational flexibility (reduced hydrogen bonding ability and cis/trans isomerism). We here use NMR spectroscopy to answer how the insertion of a peptoid monomer influences the structure of a regular α-helical AMP upon interaction with a dodecyl phosphocholine (DPC) micelle. Insertion of [(2-methylpropyl)amino]acetic acid in maculatin-G15 shows that the structural change and conformational flexibility depends on the site of insertion. This is governed by the micelle interaction of the amphipathic helices flanking the peptoid monomer and the side chain properties of the peptoid and its preceding residue.     

Brain–computer interfaces for dissecting cognitive processes underlying sensorimotor control

Sensorimotor control engages cognitive processes such as prediction, learning, and multisensory integration. Understanding the neural mechanisms underlying these cognitive processes with arm reaching is challenging because we currently record only a fraction of the relevant neurons, the arm has nonlinear dynamics, and multiple modalities of sensory feedback contribute to control. A brain–computer interface (BCI) is a well-defined sensorimotor loop with key simplifying advantages that address each of these challenges, while engaging similar cognitive processes. As a result, BCI is becoming recognized as a powerful tool for basic scientific studies of sensorimotor control. Here, we describe the benefits of BCI for basic scientific inquiries and review recent BCI studies that have uncovered new insights into the neural mechanisms underlying sensorimotor control.     

A Highlights from MBoC Selection: Cell type–dependent mechanisms for formin-mediated assembly of filopodia

Filopodia are finger-like protrusions from the plasma membrane and are of fundamental importance to cellular physiology, but the mechanisms governing their assembly are still in question. One model, called convergent elongation, proposes that filopodia arise from Arp2/3 complex–nucleated dendritic actin networks, with factors such as formins elongating these filaments into filopodia. We test this model using constitutively active constructs of two formins, FMNL3 and mDia2. Surprisingly, filopodial assembly requirements differ between suspension and adherent cells. In suspension cells, Arp2/3 complex is required for filopodial assembly through either formin. In contrast, a subset of filopodia remains after Arp2/3 complex inhibition in adherent cells. In adherent cells only, mDia1 and VASP also contribute to filopodial assembly, and filopodia are disproportionately associated with focal adhesions. We propose an extension of the existing models for filopodial assembly in which any cluster of actin filament barbed ends in proximity to the plasma membrane, either Arp2/3 complex dependent or independent, can initiate filopodial assembly by specific formins.     

NMR relaxation parameters of methyl groups as a tool to map the interfaces of helix–helix interactions in membrane proteins

A number of recent advances in the field of magic-angle-spinning (MAS) solid-state NMR have enabled its application to a range of biological systems of ever increasing complexity. To retain biological relevance, these samples are increasingly studied in a hydrated state. At the same time, experimental feasibility requires the sample preparation process to attain a high sample concentration within the final MAS rotor. We discuss these considerations, and how they have led to a number of different approaches to MAS NMR sample preparation. We describe our experience of how custom-made (or commercially available) ultracentrifugal devices can facilitate a simple, fast and reliable sample preparation process. A number of groups have since adopted such tools, in some cases to prepare samples for sedimentation-style MAS NMR experiments. Here we argue for a more widespread adoption of their use for routine MAS NMR sample preparation.