Supramolecular associations between atypical oxidative phosphorylation complexes of Euglena gracilis

In vivo associations of respiratory complexes forming higher supramolecular structures are generally accepted nowadays. Supercomplexes (SC) built by complexes I, III and IV and the so-called respirasome (I/III2/IV) have been described in mitochondria from several model organisms (yeasts, mammals and green plants), but information is scarce in other lineages. Here we studied the supramolecular associations between the complexes I, III, IV and V from the secondary photosynthetic flagellate Euglena gracilis with an approach that involves the extraction with several mild detergents followed by native electrophoresis. Despite the presence of atypical subunit composition and additional structural domains described in Euglena complexes I, IV and V, canonical associations into III2/IV, III2/IV2 SCs and I/III2/IV respirasome were observed together with two oligomeric forms of the ATP synthase (V2 and V4). Among them, III2/IV SC could be observed by electron microscopy. The respirasome was further purified by two-step liquid chromatography and showed in-vitro oxygen consumption independent of the addition of external cytochrome c

The pseudo‐brookite spin‐glass system studied by means of muon spin relaxation

Zero-field muon spin relaxation (µSR) experiments have been performed on the spin glass Fe1.75Ti1.25O5. Above the spin-glass temperature of 44 K a distinct exponential µSR rate (¿) is observed, while below Tg a square-root exponential decay occurs, indicating fast spin fluctuations. Near 8 K, a maximum in ¿ is indicative of transverse spin ordering. The low ¿ values and the sharp ¿ peak at Tg are very promising for the study of spin freezing models like the Vogel–Fulcher law or the power law

Transmission Electron Microscopy as a Tool for the Characterization of Soft Materials:Application and Interpretation

Transmission electron microscopy (TEM) provides direct structural information on nano-structured materials and is popular as a characterization tool in soft matter and supramolecular chemistry. However, technical aspects of sample preparation are overlooked and erroneous image interpretations are regularly encountered in the literature. There are three most commonly used TEM methods as we derived from literature: drying, staining and cryo-TEM, which are explained here with respect to their application, limitations and interpretation. Since soft matter chemistry relies on a lot of indirect evidence, the role of TEM for the correct evaluation of the nature of an assembly is very large. Mistakes in application and interpretation can therefore have enormous impact on the quality of present and future studies. We provide helpful background information of these three techniques, the information that can and cannot be derived from them and provide assistance in selecting the right technique for soft matter imaging. This essay warns against the use of drying and explains why. In general cryo-TEM is by far the best suited method and many mistakes and over-interpretations can be avoided by the use of this technique

Purification and characterization of an alcohol dehydrogenase from 1,2-propanediol-grown Desulfovibriostrain HDv

The sulfate-reducing bacterium Desulfovibrio strain HDv (DSM 6830) grew faster on (S)- and on (R, S)-1,2-propanediol (µmax 0.053 h–1) than on (R)-propanediol (0.017 h–1) and ethanol (0.027 h–1). From (R, S)-1,2-propanediol-grown cells, an alcohol dehydrogenase was purified. The enzyme was oxygen-labile, NAD-dependent, and decameric; the subunit mol. mass was 48 kDa. The N-terminal amino acid sequence indicated similarity to alcohol dehydrogenases belonging to family III of NAD-dependent alcohol dehydrogenases, the first 21 N-terminal amino acids being identical to those of the Desulfovibrio gigas alcohol dehydrogenase. Best substrates were ethanol and propanol (Km of 0.48 and 0.33 mM, respectively). (R, S)-1,2-Propanediol was a relatively poor substrate for the enzyme, but activities in cell extracts were high enough to account for the growth rate. The enzyme showed a preference for (S)-1,2-propanediol over (R)-1,2-propanediol. Antibodies raised against the alcohol dehydrogenase of D. gigas showed cross-reactivity with the alcohol dehydrogenase of Desulfovibrio strain HDv and with cell extracts of six other ethanol-grown sulfate-reducing bacteria.

One-step isolation and biochemical characterization of a highlyactive plant PSII monomeric core

We describe a one-step detergent solubilization protocol for isolating a highly active form of Photosystem II (PSII) from Pisum sativum L. Detailed characterization of the preparation showed that the complex was a monomer having no light harvesting proteins attached. This core reaction centre complex had, however, a range of low molecular mass intrinsic proteins as well as the chlorophyll binding proteins CP43 and CP47 and the reaction centre proteins D1 and D2. Of particular note was the presence of a stoichiometric level of PsbW, a low molecular weight protein not present in PSII of cyanobacteria. Despite the high oxygen evolution rate, the core complex did not retain the PsbQ extrinsic protein although there was close to a full complement of PsbO and PsbR and partial level of PsbP. However, reconstitution of PsbP and PsbPQ was possible. The presence of PsbP in absence of LHCII and other chlorophyll a/b binding proteins confirms that LHCII proteins are not a strict requirement for the assembly of this extrinsic polypeptide to the PSII core in contrast with the conclusion of Caffarri et al. (2009)