Thermodynamic aspects of DsbD-mediated electron transport

DsbD from Escherichia coli transports electrons from cytoplasmic thioredoxin across the inner membrane to the periplasmic substrate proteins DsbC, DsbG and CcmG. DsbD consists of three domains: a periplasmic N-terminal domain, a central transmembrane domain (tmDsbD) and a periplasmic C-terminal domain. Each domain contains two essential cysteine residues that are required for electron transport. In contrast to the quinone reductase DsbB, HPLC analysis of the methanol/hexane extracts of purified DsbD revealed no presence of quinones, suggesting that the tmDsbD interacts with thioredoxin and the periplasmic C-terminal domain exclusively via disulfide exchange. We also demonstrate that a DsbD variant containing only the redox-active cysteine pair C163 and C285 in tmDsbD, reconstituted into liposomes, has a redox potential of − 0.246 V. The results show that all steps in the DsbD-mediated electron flow are thermodynamically favorable.     

Two types of information processing in cercal systems of insects: directional sensitivity of giant interneurons

Summary Cercal systems of seven insect species (cricketMelanogryllus desertus, mole cricketGryllotalpa gryllotalpa, katydidsPholidoptera pustulipes andTettigonia viridissima, cockroachesPeriplaneta americana andBlatta orientalis, and locustLocusta migratoria) were examined for direction-sensitive giant interneurons (GIs) that are excited by cercal receptors but have directional preferences independent of cercus position. Such GIs are known for the cricketsAcheta domesticus andGryllus bimaculatus. Directional sensitivity diagrams (DSDs) of GIs were obtained by recording and analysing the electrical responses of abdominal connectives to sound stimuli from various directions. For each animal DSDs were plotted in the form of polar graphs for two or three positions of the stimulated cercus so that the effect of cercus position on the orientation of the DSD could be evaluated.All insects studied had GIs whose DSDs for fixed cercus positions were similar in appearance to the DSDs described for GIs of the cricketsAcheta domesticus andGryllus bimaculatus. Most of these DSDs were shaped like a figure 8 (when airflow is used as the stimulus instead of sound, each DSD has only one lobe). However, not all GIs demonstrated a constant directional preference. GIs with constant directionality were found only inMelanogryllus desertus, Pholidoptera pustulipes, Tettigonia viridissima andLocusta migratoria. In these insects DSDs from one GI plotted for different cercus positions had approximately the same orientation (Figs. 4–7). In contrast, GIs inGryllotalpa gryllotalpa, Periplaneta americana andBlatta orientalis had DSDs whose orientation changed in accordance with a change in position of the stimulated cercus (Figs. 8–10).Thus, direction-sensitive GIs investigated here can be divided into two types: (1) GIs with constant directionality (whose DSDs are fixed to the body, and (2) GIs with variable directionality (whose DSDs are fixed to the cerci). To date, in each species only GIs of the same type have been encountered. This may be an indication that cercal systems can be divided into two categories according to how they process information. However, since we have not tested all GIs in each species, we cannot rule out the possibility that a species might have both types of GIs.Abbreviations DSD directional sensitivity diagram - GI giant interneuron - TAG terminal abdominal ganglion     

Polymorphism and inheritance of seed storage protein in sunflower

The data on polymorphism and inheritance of the seed storage protein helianthinin are presented. The results of hybrid analysis indicate that in the annual sunflower Helianthus annuus, helianthinin synthesis is controlled by at least three loci: HelA, HelB, HelB, and HelC. Codominant alleles controlling different electrophoretic variants of polypeptides were identified at each of the loci. The HelA locus was inherited independently of HelB and HelC in a series of dihybrid crosses. The frequencies of recombination between loci HelB and HelC estimated in F2 and BC of two crossing combinations were respectively 21.8 and 19.0%. Segregation of the Hel-C-controlled variants in the progenies from the crosses of cultured sunflower with annual wild species and forms corresponded to that theoretically expected for Mendelian inheritance. The maternal type of helianthinin inheritance was observed in the progenies from the crosses of inbred H. annuus lines with perennial diploid and polyploid Helianthus species. Altered expression of the HelC locus was detected in some hybrid combinations. These alterations appeared in early (F1, F2) hybrid generations and were similar in different hybrid combinations. They did not depend on the perennial paternal species being more influenced by the maternal genotype and by the mode of obtaining hybrids (in an embryo culture or in the field). These results are explained by "genomic shock" generated by hybridization of genetically incompatible species.     

Structural basis and kinetics of inter- and intramolecular disulfide exchange in the redox catalyst DsbD

DsbD from Escherichia coli catalyzes the transport of electrons from cytoplasmic thioredoxin to the periplasmic disulfide isomerase DsbC. DsbD contains two periplasmically oriented domains at the N- and C-terminus (nDsbD and cDsbD) that are connected by a central transmembrane (TM) domain. Each domain contains a pair of cysteines that are essential for catalysis. Here, we show that Cys109 and Cys461 form a transient interdomain disulfide bond between nDsbD and cDsbD in the reaction cycle of DsbD. We solved the crystal structure of this catalytic intermediate at 2.85 A resolution, which revealed large relative domain movements in DsbD as a consequence of a strong overlap between the surface areas of nDsbD that interact with DsbC and cDsbD. In addition, we have measured the kinetics of all functional and nonfunctional disulfide exchange reactions between redox-active, periplasmic proteins and protein domains from the oxidative DsbA/B and the reductive DsbC/D pathway. We show that both pathways are separated by large kinetic barriers for nonfunctional disulfide exchange between components from different pathways.     

Influence of the cycle number in processing of cellulose from waste paper on its ability to hydrolysis by cellulases

Hydrolytic ability of laboratory enzyme preparations from fungus of the Penicillium genus was investigated using kraft pulp from nonbleached softwood and bleached hardwood cellulose as substrates. The enzyme preparations were shown to efficiently hydrolyze both softwood and hardwood cellulose. The yields of glucose and reducing sugars were 24–36 g/l and 27–37 g/l from 100 g/l of dry substrate in 48 h, respectively, and depended on the number of substrate grinding cycles.     

Chemical modification of epsilon-amino lysine groups in horseradish peroxidase. Its effect on catalytic properties and spatial structure of the enzyme]

Effect of chemical modification of horseradish peroxidase lysine epsilon-amino groups by propionic, butyric, valeric, succinic anhydrides and trinitrobenzolsulfonic acid (TNBS) on catalytic properties of the enzyme is investigated. All the preparations of modified peroxidase have 100% peroxidase activity for o-dianizidine at pH 7.0, which indicates the absence of lysine epsilon-amino group in the enzyme active site. pH-dependencies of modified peroxidase relative activity are studied; modification by anhydrides of monobasic acids is not found to result in changes of the relative activity pH-profile, while modification by succinic anhydride widens it. Absorption and circular dichoism spectra of native and modified peroxidase within 260--270 nm are obtained, some changes in the enzyme tertiary structure after its epsilon-amino groups modification are observed. Modification of four epsilon-amino groups by buturic and succinic anhydrides and of three epsilon-amino groups by TNBS is found to increase the regidity of protein surrounding of heme, and modification of six epsilon-amino groups by TNBS results in more unwrapped enzyme structure as compared with its native molecule.     

Production of Industrial Enzymes Based on the Expression System of the Fungus Penicillium verruculosum

Applied Biochemistry and Microbiology - The review presents data on the creation of highly active recombinant producer strains of a wide range of enzymes for various fields of industry and...     

The synthesis and antioxidant activity of azole derivatives of hemin

New derivatives of proto- and deuterohemin IX containing tri- and tetrazole rings were synthesized and characterized. A pronounced antioxidant activity was found for these compounds in the Fe(II)/ascorbate-dependent system of lipid peroxidation in murine liver homogenates.     

Feeding of the stone sculpin Paracottus knerii (Cottidae) in the littoral of southern Baikal

The food spectrum and seasonal dynamics of feeding of the stone sculpin Paracottus knerii in the littoral of southern Baikal is investigated. Altogether, 29 species of invertebrates are recorded. All year round, the bulk of food, similarly to other areas of the lake, consisted of amphipods; other groups of zoobenthos were represented insignificantly. The significance of particular amphipod species in the food of fish depended on their ecological and morphofunctional traits. The seasonal changes in the species composition and the ratio of food items in the food depended on life cycles of fish and on the migrations of zoobenthic organisms.