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991.
Giulia Di Rocco Gianantonio Battistuzzi Carlo Augusto Bortolotti Marco Borsari Erika Ferrari Stefano Monari Marco Sola 《Journal of biological inorganic chemistry》2011,16(3):461-471
The 16-kDa diheme cytochrome c from the bacterium Shewanella baltica OS155 (Sb-DHC) was cloned and expressed in Escherichia coli and investigated through UV–vis, magnetic circular dichroism, and 1H NMR spectroscopies and protein voltammetry. The model structure was obtained by means of comparative modeling using the
X-ray structure of Rhodobacter sphaeroides diheme cytochrome c (Rs-DHC) (with a 37% pairwise sequence identity) as a template. Sb-DHC folds into two distinct domains, each containing one
heme center with a bis-His axial ligation. Both secondary and tertiary structures of the N-terminal domain resemble those
of class I cytochrome c, displaying three α-helices and a compact overall folding. The C-terminal domain is less helical than the corresponding domain
of Rs-DHC. The two heme groups are bridged by Tyr26 in correspondence with the shortest edge-to-edge distance, a feature which
would facilitate fast internal electron transfer. The electronic properties of the two prosthetic centers are equivalent and
sensitive to two acid–base equilibria with pK
a values of approximately 2.4 and 5, likely corresponding to protonation and detachment of the axial His ligands from the heme
iron and a pH-linked conformational change of the protein, respectively. Reduction potentials of −0.144 and −0.257 V (vs.
the standard hydrogen electrode), were determined for the C- and N-terminal heme groups, respectively. An approach based on
the extended Debye–Hückel equation was applied for the first time to a two-centered metalloprotein and was found to reproduce
successfully the ionic strength dependence of E°′. 相似文献
992.
Cassano M Dellavalle A Tedesco FS Quattrocelli M Crippa S Ronzoni F Salvade A Berardi E Torrente Y Cossu G Sampaolesi M 《Development (Cambridge, England)》2011,138(20):4523-4533
Mice deficient in α-sarcoglycan (Sgca-null mice) develop progressive muscular dystrophy and serve as a model for human limb girdle muscular dystrophy type 2D. Sgca-null mice suffer a more severe myopathy than that of mdx mice, the model for Duchenne muscular dystrophy. This is the opposite of what is observed in humans and the reason for this is unknown. In an attempt to understand the cellular basis of this severe muscular dystrophy, we isolated clonal populations of myogenic progenitor cells (MPCs), the resident postnatal muscle progenitors of dystrophic and wild-type mice. MPCs from Sgca-null mice generated much smaller clones than MPCs from wild-type or mdx dystrophic mice. Impaired proliferation of Sgca-null myogenic precursors was confirmed by single fiber analysis and this difference correlated with Sgca expression during MPC proliferation. In the absence of dystrophin and associated proteins, which are only expressed after differentiation, SGCA complexes with and stabilizes FGFR1. Deficiency of Sgca leads to an absence of FGFR1 expression at the membrane and impaired MPC proliferation in response to bFGF. The low proliferation rate of Sgca-null MPCs was rescued by transduction with Sgca-expressing lentiviral vectors. When transplanted into dystrophic muscle, Sgca-null MPCs exhibited reduced engraftment. The reduced proliferative ability of Sgca-null MPCs explains, at least in part, the severity of this muscular dystrophy and also why wild-type donor progenitor cells engraft efficiently and consequently ameliorate disease. 相似文献
993.
Matias Pasquali Marco Beyer Torsten Bohn Lucien Hoffmann 《Journal of Phytopathology》2011,159(10):700-704
Genetic chemotyping is an essential tool for characterizing Fusarium populations causing head blight on wheat and other cereals. Three PCR methods, based on tri cluster polymorphism, were optimized and compared on 94 single‐spore isolates obtained from three continents belonging to F. gramineaurm, F. culmorum, F. poae, F. avenaceum and Microdochium nivale. While the methods based on the tri3, tri7 and tri12 polymorphism correctly identified all the tested strains, the method based on tri13 polymorphism was unable to discriminate between the 3‐ and 15‐acetylated DON forms in F. graminearum. It is advised to avoid the use of tri13 polymorphism for genetic chemotyping of the two acetylated chemotypes. 相似文献
994.
Crisma M Moretto A Peggion C Panella L Kaptein B Broxterman QB Formaggio F Toniolo C 《Amino acids》2011,41(3):629-641
The synthesis of the N-protected (blocked) homo-peptide esters from the chiral C(α)-ethyl, C(α)-n-pentylglycine was performed in solution to the hexapeptide level. The conformational propensity exhibited by these oligomers in chloroform solution and in the crystal state was assessed by use of FTIR absorption, NMR, and X-ray diffraction. The results indicated that fully extended helical structures (2.0(5)-helices) are overwhelmingly adopted irrespective of the peptide main-chain length. This oligomeric series is of great interest as it is characterized by the longest C ( i )(α) ,…, C ( i+1 )(α) (per residue) separation achievable in the class of chiral, rigid, helical peptide spacers based on α-amino acids. 相似文献
995.
Root-feeding insects are important drivers in ecosystems, and links between aboveground oviposition preference and belowground
larval performance have been suggested. The root-colonizing arbuscular mycorrhizal fungi (AMF) play a central role in plant
nutrition and are known to change host quality for root-feeding insects. However, it is not known if and how AMF affect the
aboveground oviposition of insects whose offspring feed on roots. According to the preference–performance hypothesis, insect
herbivores oviposit on plants that will maximize offspring performance. In a greenhouse experiment with rice (Oryza sativa), we investigated the effects of AMF (Glomus intraradices) on aboveground oviposition of rice water weevil (Lissorhoptrus oryzophilus), the larvae of which feed belowground on the roots. Oviposition (i.e., the numbers of eggs laid by weevil females in leaf
sheaths) was enhanced when the plants were colonized by AMF. However, the leaf area consumed by adult weevils was not affected.
Although AMF reduced plant biomass, it increased nitrogen (N) and phosphorus concentrations in leaves and N in roots. The
results suggest that rice water weevil females are able to discriminate plants for oviposition depending on their mycorrhizal
status. The discrimination is probably related to AMF-mediated changes in plant quality, i.e., the females choose to oviposit
more on plants with higher nutrient concentrations to potentially optimize offspring performance. AMF-mediated change in plant
host choice for chewing insect oviposition is a novel aspect of below- and aboveground interactions. 相似文献
996.
Serini S Donato V Piccioni E Trombino S Monego G Toesca A Innocenti I Missori M De Spirito M Celleno L Fasano E Ranelletti FO Calviello G 《The Journal of nutritional biochemistry》2011,22(9):874-885
The dramatic increase in the incidence of nonmelanoma skin cancer over the last decades has been related to the augmented exposure to ultraviolet (UV) radiation (UVR). It is known that apoptosis is induced as a protective mechanism after the acute irradiation of keratinocytes, whereas apoptotic resistance and carcinogenesis may follow the chronic exposure to UVR. We found that not all the human keratinocytes lines studied underwent apoptosis following acute exposure to UVR (10-60 mJ/cm2). Whereas UVR induced apoptosis in the HaCaT cells, NCTC 2544 and nr-HaCaT cells showed apoptosis resistance. The cytokeratin pattern of the apoptosis-resistant cells indicated that they possessed a degree of differentiation lower than that of HaCaT cells. They also showed an enhanced expression of cyclooxygenase-2 (COX-2), an early marker of carcinogenesis in various tissues, including skin. n-3 polyunsaturated fatty acids have drawn increasing interest as nutritional factors with the potential to reduce UVR carcinogenesis, and since they are apoptosis inducers and COX-2 inhibitors in cancer cells, we investigated the ability of n-3 polyunsaturated fatty acids to influence the resistance to UVR-induced apoptosis in keratinocytes. We observed that docosahexaenoic acid (DHA) reverted the resistance of nr-HaCaT cells to UVR-induced apoptosis, increasing the Bax/Bcl-2 ratio and caspase-3 activity, and reduced COX-2 levels by inhibiting the expression of the human antigen R (HuR), a known COX-2 mRNA stabilizer in keratinocytes. The transfection of nr-HaCaT cells with HuR siRNA mimicked the proapoptotic effect of DHA. Overall, our findings further support the role of DHA as a suitable anticarcinogenic factor against nonmelanoma skin cancers. 相似文献
997.
Beres C de Nazaré FV de Souza NC Miguel MA Werneck MM 《Biosensors & bioelectronics》2011,30(1):328-332
Cells detection is crucial in microbiological analysis of clinical, food, water or environmental samples. However, currently employed methods are time consuming. Plastic optical fiber (POF) biosensors consist in a viable alternative for rapid and inexpensive scheme for detection. In order to study the sensitivity of tapers for microbiological detection, geometric parameters are studied, such as the taper waist diameter since the formation of taper regions are the key sensing element in this particular type of sensors. In this study, a series of POF taper sensors were prepared using a specially developed tapering machine, and the dispersion of geometric dimensions is evaluated, aiming to achieve the best tapering characteristics which will provide a better sensitivity on the sensor response. The fiber tapers that presented the finest results were those constructed in U-shaped (bended) configurations, with taper waist diameters ranging from 0.40 mm up to 0.50 mm. These fiber tapers were used as the main section of the monitoring device, and when chemically treated as immunosensors for the detection of bacteria, yeast and erythrocytes. 相似文献
998.
The development of a voltammetric immunosensor for determination of alpha-fetoprotein (AFP) in serum is presented. ELISA assays with voltammetric reading were carried out exploiting the peculiar properties of nanobiocomposite materials based on gold nanoparticles for the immobilization of Antibody (Ab)/Antigen/Antibody-HRP (Horseradish Peroxidase) sandwich on the glassy carbon (GC) electrode surface. The electrochemical transduction was mediated by thionin, which was used in its monomeric form dissolved in the reading solution, so avoiding critical immobilization procedures. The study was aimed at the development and validation of an immunosensor able to provide results in short time, simple to use, rugged and cost-effective for AFP monitoring purposes. A crucial aspect of the study was the development of an experimental protocol leading to highly standardized and consequently reproducible sensors. Two-way analysis of variance (ANOVA) was applied to study the effect of the concentration of the solutions used for the incubation of the antibodies. The sensor was validated in serum assessing stability of the immunocomplex, linearity of response, limit of detection (3.7 ng/ml) and limit of quantitation (11 ng/ml), precision (intra- and inter-sensor repeatability) and recovery rate (103%). The stability of the GC/Ab functionalized substrate was demonstrated over one month, showing variation coefficients below 5%. Experiments carried out with real samples of clinical interest evidenced that the developed immunosensor can be considered as powerful tool in cancer screening programmes. 相似文献
999.
The possibility of introducing a computationally assisted method to study aptamer-protein interaction was evaluated with the aim of streamlining the screening and selection of new aptamers. Starting from information on the 15-mer (5'-GGTTGGTGTGGTTGG-3') thrombin binding aptamer (TBA), a library of mutated DNA sequences (994 elements) was generated and screened using shapegauss a shape-based scoring function from openeye software to generate computationally derived binding scores. The TBA and three other mutated oligonucleotides, selected on the basis of their binding score (best, medium, worst), were incorporated into surface plasmon resonance (SPR) biosensors. By reducing the ionic strength (binding buffer, 50 mM TrisHCl pH 7.4, 140 mM NaCl, 1mM MgCl?, diluted 1:50) in order to match the simulated condition, the analytical performances of the four oligonucleotide sequences were compared using signal amplitude, sensitivity (slope), linearity (R2) and reproducibility (CVav %). The experimental results were in agreement with the simulation findings. 相似文献
1000.
A wide variety of human diseases are associated with the formation of highly organized protein aggregates termed amyloid fibrils, whose growth (elongation) is due to the assembly of the basic molecular units (monomers) in a sequential polymerization process. Surface plasmon resonance (SPR) technology has been proposed as a powerful approach to study in detail the fibril elongation of some amyloidogenic peptides. In particular, the injection of monomers over immobilized fibrils allows to follow in real time, and on a very short time-scale, the kinetics of fibril growth. In the present study we confirmed and extended this application of SPR to Aβ(1-42), hampered till now by the very pronounced aggregation propensity of this peptide, involved in Alzheimer disease. We took advantage of a new synthetic strategy ("depsi-peptide" technique) which allows to obtain reliable seed-free solutions (monomers) as well as fibrils of Aβ(1-42). SPR data were consistent with a "dock-and-lock" mechanism underlying Aβ(1-42) elongation process. The setup of an assay monitoring the elongation kinetics is very useful for investigating potential anti-amyloidogenic compounds. Moreover, the possibility to reliably immobilize both Aβ(1-42) monomers and fibrils allows to measure the binding affinities of putative ligands for these different species. The approach applied here to Aβ(1-42) might well be also applied to the study of other fibrillogenic peptides/proteins or to the study of polymerization reactions in general. 相似文献