Characterization of a putative phosphorylation switch: adaptation of SPOT synthesis to analyze PDZ domain regulation mechanisms

Transient macromolecular complexes are often formed by protein-protein interaction domains (e.g., PDZ, SH2, SH3, WW), which are often regulated (positively or negatively) by phosphorylation. To address the in vitro analysis of PDZ domain regulation by such phosphorylation, we improved the inverted peptide method. This method is based on standard SPOT synthesis, followed by inversion of the peptide under acidic conditions to generate the free C termini necessary for PDZ domain ligand recognition. The benefit of the newly introduced acidic conditions is the preservation of the incorporated phosphate group during peptide synthesis. Furthermore, the improved method is more robust and shows an increased signal-to-noise ratio. As representative examples, we used the AF6, ERBIN, and SNA1 (alpha-1-syntrophin) PDZ domains to analyze the influence of ligand-position-dependent phosphorylation. We could clearly demonstrate severe down-regulation by phosphorylation of the PDZ ligand position -2 (<50 %) and slightly less at position -1 ( approximately 50 %). These results are specific and reproducible for all three PDZ domains. Finally, we confirmed the influence of negative regulation by using the protein kinase BCR as the AF6 PDZ domain ligand. For the first time, this approach allows the SPOT synthesis technique to be used to screen large libraries of phosphorylated peptides in vitro. This should ultimately help in the identification of phosphorylation-dependent regulation mechanisms in vivo.     

Laser optical feedback imaging insensitive to parasitic optical feedback

We present an optical architecture for the laser optical feedback imaging (LOFI) technique that makes it possible to avoid the effect of the optical parasitic reflections introduced by the optical components located between the laser source and the studied object. These reflections damage phase and amplitude information contained in the images. This phenomenon is a leading problem that strongly limits the LOFI performance for weak feedback detection. Consequently, it is essential to be able to limit or avoid the effect of these parasitic reflections to reach the optimal LOFI performance.     

From bacterial genome to functionality; case bifidobacteria

The availability of complete bacterial genome sequences has significantly furthered our understanding of the genetics, physiology and biochemistry of the microorganisms in question, particularly those that have commercially important applications. Bifidobacteria are among such microorganisms, as they constitute mammalian commensals of biotechnological significance due to their perceived role in maintaining a balanced gastrointestinal (GIT) microflora. Bifidobacteria are therefore frequently used as health-promoting or probiotic components in functional food products. A fundamental understanding of the metabolic activities employed by these commensal bacteria, in particular their capability to utilize a wide range of complex oligosaccharides, can reveal ways to provide in vivo growth advantages relative to other competing gut bacteria or pathogens. Furthermore, an in depth analysis of adaptive responses to nutritional or environmental stresses may provide methodologies to retain viability and improve functionality during commercial preparation, storage and delivery of the probiotic organism.     

On computing dominant frequency from bipolar intracardiac electrograms

Dominant frequency (DF) computed from action potentials is a key parameter for investigating atrial fibrillation in animal studies and computer models. A recent clinical trial reported consistent results computing DF from 30 Hz to 400 Hz bandpass filtered bipolar electrograms in humans. The DF (< 15 Hz and, thus, filtered out) was recovered by rectifying the signal, while the theoretical background of this approach was left uncommented. It is the focus of this paper to provide this background by a Fourier analysis. We demonstrate that it is mainly the timing of the narrow deflections (local activation at the catheter tip) which contribute to the DF peak in the frequency spectrum. Due to the typical signal morphology pronounced harmonic peaks occur in the spectrum. This is a disadvantage when computing the regularity index (RI) as a parameter for local organization and signal quality. It is demonstrated for synthetical and patient data that at low DF the RI is far below the optimal value one even for high underlying organization and good signal quality. The insight obtained promotes the development of better measures for organization. The finding that mainly timing of activation contributes to DF might promote the development of powerful realtime signal processing tools for computing DF.     

Model-based development of neural prostheses for movement

Neural prostheses for restoration of limb movement in paralyzed and amputee patients tend to be complex systems. Subjective intuition and trial-and-error approaches have been applied to the design and clinical fitting of simple systems with limited functionality. These approaches are time consuming, difficult to apply in larger scale, and not applicable to limbs under development with more anthropomorphic motion and actuation. The field of neural prosthetics is in need of more systematic methods, including tools that will allow users to develop accurate models of neural prostheses and simulate their behavior under various conditions before actual manufacturing or clinical application. Such virtual prototyping would provide an efficient and safe test-bed for narrowing the design choices and tuning the control parameters before actual clinical application. We describe a software environment that we have developed to facilitate the construction and modification of accurate mathematical models of paralyzed and prosthetic limbs and simulate their movement under various neural control strategies. These simulations can be run in real time with a stereoscopic display to enable design engineers and prospective users to evaluate a candidate neural prosthetic system and learn to operate it before actually receiving it.     

Conformational changes during protein adsorption. FT-IR spectroscopic imaging of adsorbed fibrinogen layers

The influence of hydrophobicity of the substrate surface on structural changes during protein adsorption was investigated. Plasma fibrinogen was chosen to model this effect as it is the most important protein in the body that adsorbs to foreign surfaces. Only conformations of adsorbed fibrinogen similar to that of the protein in solution do not activate the process of blood coagulation. Small spots on the substrate surface with conformational changes within the adsorbed protein are already sufficient to deteriorate biocompatibility. Mid-infrared hyperspectral imaging permits the identification of coagulated spots down to a few micrometers in size. The spectra of the FT-IR images that were assessed to be of suitable quality were clustered by a fuzzy c-means algorithm. The determination of the appropriate number of clusters was based on cluster variance. Subsequent evaluation of the centroid spectra of each cluster showed that their amide I band was separated into contributions from different structural units, with the alpha-helix content always being dominant. Significant differences between hydrophobic and hydrophilic surfaces were observed for turn and sheet contributions. Lower sheet/turn ratios appear to indicate inferior biocompatibility. Spots on hydrophilic surfaces could be identified, which exhibit structural changes similar to those on hydrophobic surfaces.     

Virtual polymorphism: finding divergent peptide matches in mass spectrometry data

The prevailing method of analyzing tandem-MS data for protein identification involves the comparison of peptide molecular weight and fragmentation data to theoretically predicted values, based on known protein sequences in databases. This is generally effective since proteins from most species under study are in the database or have sufficient homology to allow significant matching. We have encountered difficulties identifying proteins from fungal species Alternaria alternata due to significant interspecies protein sequence differences (divergence) and its absence from the database. This common household mold causes asthma and allergy problems, but the genome has not been sequenced. De novo sequencing and error-tolerant methods can facilitate protein identifications in divergent, unsequenced species. But these standard methods can be laborious and only allow single amino acid substitution, respectively. We have developed an alternative approach focusing on database engineering, predicting biologically rational polymorphism using statistically weighted amino acid substitution information held in BLOSUM62. Like other second pass methods, it is based on the initially identified protein. However, this approach allows more control over sequences to be considered, including multiple changes per peptide. The results show considerable improvement for routine protein identification and the potential for rescuing otherwise unconvincing identifications in unusually divergent species.     

石墨烯在防腐防污涂料中的应用进展

石墨烯具有极好的阻隔性能、屏蔽性能及化学稳定性,其在防腐防污涂料中的应用已经被深入研究。介绍了石墨烯对防腐防污涂层性能的影响:降低水、氧气等腐蚀介质的渗透率,加强抗生物附着性,抑制微生物腐蚀。分析了石墨烯在涂料中的应用缺陷及产生原因:极强的范德华力导致石墨烯在涂料中分散性差、易团聚,高化学稳定性及疏水性导致石墨烯与成膜物质结合性差,超高的导电性导致石墨烯膜在失效时加速金属腐蚀。综述了为应对石墨烯在防腐防污涂料中的应用缺陷,国内外学者采用的主要方法:采用改性处理方法制备改性石墨烯(GO、RGO、FG)以及合成石墨烯复合颗粒(石墨烯修饰纳米粒子,即GO-Al2O3颗粒、GO-TiO2颗粒、GO-SiO2颗粒等;树脂负载石墨烯复合填料,即石墨烯/聚苯胺复合填料等)。最后展望了石墨烯及其衍生物在防腐防污涂料中的发展。     

Stepwise measurement procedure for the characterization of large‐area photovoltaic modules

In this paper, we present an indoor measurement procedure for characterizing the electrical performance of large aperture photovoltaic modules. Because of the fact that sun simulators, especially for concentrator photovoltaic applications, are strongly limited in the size of the uniformly illuminated area, we developed a measurement procedure that allows characterizing modules with a larger aperture area than the aperture provided by the sun simulator. The procedure is based on the concept of stepwise illumination of the module area and measurement of the corresponding I–V curves—without the need to contact the subunits directly. Using the additionally measured dark I–V curve of the module, the characteristic I–V curve of the full module can be calculated. Copyright © 2015 John Wiley & Sons, Ltd.     

FLATCON® CPV module with 36.7% efficiency equipped with four‐junction solar cells

In this paper we present the application of high efficiency four‐junction solar cells using SOITEC bonding technology under a Fresnel lens optic and in a FLATCON®‐type CPV module. We demonstrate very high performance. The measurement of a sub‐module, consisting of a four‐junction solar cell adjusted under a single Fresnel lens, showed an efficiency of 38.9%. An 829.6 cm² sized FLATCON®‐type CPV module yielded in an efficiency of 35.0% and 36.7% at CSOC and CSTC, respectively. Thus, both, the sub‐module and the CPV module showed record values, which prove the usefulness of high efficiency four‐junction solar cells in CPV applications. Copyright © 2014 John Wiley & Sons, Ltd.