Search results for "570"
showing 10 items of 1074 documents
Simulazione della propagazione di difetti a fatica mediante il modello di zona coesiva
2009
Le giunzioni incollate guadagnano sempre più mercato, nel campo delle costruzioni in genere, dove è necessario un alleggerimento delle strutture. Nel caso di geometrie di giunto semplici il dimensionamento avviene attraverso relazioni analitiche che restituiscono il valore massimo delle tensioni, il quale deve essere inferiore al limite di utilizzo dell’adesivo stesso. Quando le geometrie sono complesse l’approccio analitico diventa impraticabile, di conseguenza si provvede a verificare la correttezza della soluzione mediante analisi agli elementi finiti (EF). L’introduzione del modello di zona coesiva nell'analisi EF permette di simulare il danneggiamento ed il cedimento del giunto in cond…
Changes in the hydrogen-bonding strength of internal water molecules and cysteine residues in the conductive state of channelrhodopsin-1
2014
Water plays an essential role in the structure and function of proteins, particularly in the less understood class of membrane proteins. As the first of its kind, channelrhodopsin is a light-gated cation channel and paved the way for the new and vibrant field of optogenetics, where nerve cells are activated by light. Still, the molecular mechanism of channelrhodopsin is not understood. Here, we applied time-resolved FT-IR difference spectroscopy to channelrhodopsin-1 from Chlamydomonas augustae. It is shown that the (conductive) P2(380) intermediate decays with τ ≈ 40 ms and 200 ms after pulsed excitation. The vibrational changes between the closed and the conductive states were analyzed in…
Structure of Rhodococcus erythropolis limonene-1,2-epoxide hydrolase reveals a novel active site
2003
Epoxide hydrolases are essential for the processing of epoxide-containing compounds in detoxification or metabolism. The classic epoxide hydrolases have an alpha/beta hydrolase fold and act via a two-step reaction mechanism including an enzyme-substrate intermediate. We report here the structure of the limonene-1,2-epoxide hydrolase from Rhodococcus erythropolis, solved using single-wavelength anomalous dispersion from a selenomethionine-substituted protein and refined at 1.2 A resolution. This enzyme represents a completely different structure and a novel one-step mechanism. The fold features a highly curved six-stranded mixed beta-sheet, with four alpha-helices packed onto it to create a …
Insights into the catalytic mechanism of human sEH phosphatase by site-directed mutagenesis and LC-MS/MS analysis
2008
We have recently reported that human soluble epoxide hydrolase (sEH) is a bifunctional enzyme with a novel phosphatase enzymatic activity. Based on a structural relationship with other members of the haloacid dehalogenase superfamily, the sEH N-terminal phosphatase domain revealed four conserved sequence motifs, including the proposed catalytic nucleophile D9, and several other residues potentially implicated in substrate turnover and/or Mg(2+) binding. To enlighten the catalytic mechanism of dephosphorylation, we constructed sEH phosphatase active-site mutants by site-directed mutagenesis. A total of 18 mutants were constructed and recombinantly expressed in Escherichia coli as soluble pro…
The Protein Structure Context of PolyQ Regions.
2016
Proteins containing glutamine repeats (polyQ) are known to be structurally unstable. Abnormal expansion of polyQ in some proteins exceeding a certain threshold leads to neurodegenerative disease, a symptom of which are protein aggregates. This has led to extensive research of the structure of polyQ stretches. However, the accumulation of contradictory results suggests that protein context might be of importance. Here we aimed to evaluate the structural context of polyQ regions in proteins by analysing the secondary structure of polyQ proteins and their homologs. The results revealed that the secondary structure in polyQ vicinity is predominantly random coil or helix. Importantly, the region…
Structure of Aspergillus niger epoxide hydrolase at 1.8 A resolution: implications for the structure and function of the mammalian microsomal class o…
2000
AbstractBackground: Epoxide hydrolases have important roles in the defense of cells against potentially harmful epoxides. Conversion of epoxides into less toxic and more easily excreted diols is a universally successful strategy. A number of microorganisms employ the same chemistry to process epoxides for use as carbon sources.Results: The X-ray structure of the epoxide hydrolase from Aspergillus niger was determined at 3.5 Å resolution using the multiwavelength anomalous dispersion (MAD) method, and then refined at 1.8 Å resolution. There is a dimer consisting of two 44 kDa subunits in the asymmetric unit. Each subunit consists of an α/β hydrolase fold, and a primarily helical lid over the…
Structures of yeast peroxisomal Δ(3),Δ(2)-enoyl-CoA isomerase complexed with acyl-CoA substrate analogues: the importance of hydrogen-bond networks f…
2015
Δ3,Δ2-Enoyl-CoA isomerases (ECIs) catalyze the shift of a double bond from 3Z- or 3E-enoyl-CoA to 2E-enoyl-CoA. ECIs are members of the crotonase superfamily. The crotonase framework is used by many enzymes to catalyze a wide range of reactions on acyl-CoA thioesters. The thioester O atom is bound in a conserved oxyanion hole. Here, the mode of binding of acyl-CoA substrate analogues to peroxisomalSaccharomyces cerevisiaeECI (ScECI2) is described. The best defined part of the bound acyl-CoA molecules is the 3′,5′-diphosphate-adenosine moiety, which interacts with residues of loop 1 and loop 2, whereas the pantetheine part is the least well defined. The catalytic base, Glu158, is hydrogen-bo…
Crystallization and Preliminary Analysis of Crystals of the 24-Meric Hemocyanin of the Emperor Scorpion (Pandinus imperator)
2011
Hemocyanins are giant oxygen transport proteins found in the hemolymph of several invertebrate phyla. They constitute giant multimeric molecules whose size range up to that of cell organelles such as ribosomes or even small viruses. Oxygen is reversibly bound by hemocyanins at binuclear copper centers. Subunit interactions within the multisubunit hemocyanin complex lead to diverse allosteric effects such as the highest cooperativity for oxygen binding found in nature. Crystal structures of a native hemocyanin oligomer larger than a hexameric substructure have not been published until now. We report for the first time growth and preliminary analysis of crystals of the 24-meric hemocyanin (M(…
Structure of the altitude adapted hemoglobin of Guinea pig in the R2-state
2010
Background: Guinea pigs are considered to be genetically adapted to a high altitude environment based on the consistent finding of a high oxygen affinity of their blood. Methodology/Principal Findings: The crystal structure of guinea pig hemoglobin at 1.8 A u resolution suggests that the increased oxygen affinity of guinea pig hemoglobin can be explained by two factors, namely a decreased stability of the Tstate and an increased stability of the R2-state. The destabilization of the T-state can be related to the substitution of a highly conserved proline (P44) to histidine (H44) in the a-subunit, which causes a steric hindrance with H97 of the b-subunit in the switch region. The stabilizatio…
Evaluation of consistency in spheroid invasion assays
2016
Multicellular tumor spheroids embedded in a matrix represent invaluable tools to analyze cell invasion. Spheroid sizes and invasiveness are the main observables easily measurable to evaluate effects of biological or pharmaceutical manipulations on invasion. They largely account for these 3-D platforms variability, leading to flaws in data interpretation. No method has been established yet that characterizes this variability and guarantees a reliable use of 3-D platforms. Spheroid initial/end sizes and invasiveness were systematically analyzed and compared in spheroids of U87MG cells generated by three different methods and embedded at different times in a collagen matrix. A normality test w…