Molecular and dimensional profiling of highly purified extracellular vesicles by fluorescence fluctuation spectroscopy.

Cells secrete extracellular vesicles (EVs) into their microenvironment that act as mediators of intercellular communication under physiological conditions and in this context also actively participate in spreading various diseases. Large efforts are currently made to produce reliable EV samples and to develop, improve, and standardize techniques allowing their biophysical characterization. Here, we used ultrafiltration and size-exclusion chromatography for the isolation and a model-free fluorescence fluctuation analysis for the investigation of the physical and biological properties of EVs secreted by mammalian cells.

Structure-based engineering of a minimal porin reveals loop-independent channel closure.

Porins, like outer membrane protein G (OmpG) of Escherichia coli, are ideal templates among ion channels for protein and chemical engineering because of their robustness and simple architecture. OmpG shows fast transitions between open and closed states, which were attributed to loop 6 (L6). As flickering limits single-channel-based applications, we pruned L6 by either 8 or 12 amino acids.

Flexibility of the N-terminal mVDAC1 segment controls the channel's gating behavior.

Since the solution of the molecular structures of members of the voltage dependent anion channels (VDACs), the N-terminal α-helix has been the main focus of attention, since its strategic location, in combination with its putative conformational flexibility, could define or control the channel's gating characteristics. Through engineering of two double-cysteine mVDAC1 variants we achieved fixing of the N-terminal segment at the bottom and midpoint of the pore. Whilst cross-linking at the midpoint resulted in the channel remaining constitutively open, cross-linking at the base resulted in an "asymmetric" gating behavior, with closure only at one electric field's orientation depending on the channel's orientation in the lipid bilayer.

Chemical engineering of Mycobacterium tuberculosis dodecin hybrids.

The suitability for chemical engineering of the highly symmetrical Mycobacterium tuberculosis dodecin was investigated, its inner cavity providing a large compartment shields introduced compounds from bulk solvent. Hybrids were obtained by S-alkylation of cysteine mutants and characterized by spectroscopic methods, including the crystal structures of wild type and biohybrid dodecins.

Strategies and perspectives in ion-channel engineering.

Membranes form natural barriers that need to be permeable to diverse matter like ions and substrates. This permeability is controlled by ion-channel proteins, which have attracted great interest for pharmaceutical applications. Ion-channel engineering (ICE) modifies biological ion channels by chemical/biological synthetis means.

Structural characterization of a beta-turn mimic within a protein-protein interface.

β-Turns are secondary structure elements not only exposed on protein surfaces, but also frequently found to be buried in protein-protein interfaces. Protein engineering so far considered mainly the backbone-constraining properties of synthetic β-turn mimics as parts of surface-exposed loops. A β-turn mimic, Hot═Tap, that is available in gram amounts, provides two hydroxyl groups that enhance its turn-inducing properties besides being able to form side-chain-like interactions.

Structural and functional characterization of a synthetically modified OmpG.

Chemical modification of ion channels has recently attracted attention due to their potential use in stochastic sensing and neurobiology. Among the available channel templates stable β-barrel proteins have shown their potential for large scale chemical modifications due to their wide pore lumen. Ion-channel hybrids using the outer membrane protein OmpG were generated by S-alkylation with a synthetic modulator and functionally as well as structurally characterized.

Xenobiotics removal from polluted water by a multifunctional constructed wetland.

Removal efficiencies on xenobiotics from polluted water in a twin-shaped constructed wetland consisting of a vertical flow chamber with the crop plant Colocasia esculenta L. Schott and a reverse vertical flow one with Ischaemum aristatum var. glaucum Honda, were assessed by chemical analysis and bioassays.

Sulphonated aromatic pollutants. Limits of microbial degradability and potential of phytoremediation.

Many synthetic sulphonated aromatic compounds are used as starting material to produce dyes and pigments, or are released as by-products in the effluents of the textile and dye industry. A large number of these chemicals are poorly biodegradable and cannot be eliminated by classical wastewater treatment plants. To limit the impact of these pollutants on the environment, new processes, based on the use of higher plants (constructed wetlands or hydroponic systems), are under development.