Numerous attempts to elucidate the strength of cadherin dimerization that mediates intercellular adhesion have produced controversial and inconclusive results. To clarify this issue, we compared E-cadherin dimerization on the surface of living cells with how the same process unfolds on agarose beads. In both cases, dimerization was monitored by the same site-specific cross-linking assay, greatly simplifying data interpretation. We showed that on the agarose surface under physiological conditions, E-cadherin produced a weak dimer that immediately dissociated after the depletion of calcium ions. However, either at pH 5 or in the presence of cadmium ions, E-cadherin produced a strong dimer that was unable to dissociate upon calcium depletion. Both types of dimers were W156-dependent. Remarkably, only the strong dimer was found on the surface of living cells. We also showed that the intracellular cadherin region, the clustering of which through catenins had been proposed as stabilizer of weak intercadherin interactions, was not needed, in fact, for cadherin junction assembly. Taken together, our data present convincing evidence that cadherin adhesion is based on high-affinity cadherin-cadherin interactions. 相似文献
The AAA(+) chaperone ClpB mediates the reactivation of aggregated proteins in cooperation with the DnaK chaperone system. ClpB consists of two AAA domains that drive the ATP-dependent threading of substrates through a central translocation channel. Its unique middle (M) domain forms a coiled-coil structure that laterally protrudes from the ClpB ring and is essential for aggregate solubilization. Here, we demonstrate that the conserved helix 3 of the M domain is specifically required for the DnaK-dependent shuffling of aggregated proteins, but not of soluble denatured substrates, to the pore entrance of the ClpB translocation channel. Helix 3 exhibits nucleotide-driven conformational changes possibly involving a transition between folded and unfolded states. This molecular switch controls the ClpB ATPase cycle by contacting the first ATPase domain and establishes the M domain as a regulatory device that acts in the disaggregation process by coupling the threading motor of ClpB with the DnaK chaperone activity. 相似文献
Brazil has 22 genera and 75 species of Cetoniidae, with the Cerrado hosting the greatest diversity among Brazilian biomes. However, the diversity of groups among the different phytophysiognomies of the biome is not known. The objectives of this study are to assess Cetoniidae diversity and to verify the seasonality of these beetles in three Cerrado phytophysiognomies (gallery forest, cerrado sensu stricto, and campo sujo) located in three conservation units in Brasilia, Distrito Federal, Brazil. We collected adults monthly (October/2016 to September/2018) using 180 traps baited with bananas fermented with sugarcane juice, totaling 1,574 specimens, 8 genera, and 17 species. Cetoniidae diversity was higher in the phytophysiognomies with lower tree density (campo sujo and cerrado sensu stricto) than in gallery forests (forest formation), confirming our hypothesis that more open areas favor the dispersal of these insects due to their diurnal long-flying behavior. The seasonality of Cetoniidae was directly related to the precipitation, with higher numbers of individuals and species in the rainy season. However, the distribution varies among the phytophysiognomies, with aggregated distribution in campo sujo and gallery forest and dispersed in cerrado sensu stricto. Our results suggest that the Cetoniidae take advantage of the open Cerrado physiognomies to locate resources faster and with less energy expenditure, presenting higher diversity in these environments, despite the more ephemeral and dispersed food resources.
Five known secondary metabolites, chrysophanol ( 1 ), 7,7′‐biphyscion ( 2 ), secalonic acid D ( 3 ), mannitol ( 4 ) and trehalose ( 5 ) were isolated for the first time from the extracts of the fungus Phialomyces macrosporus. Their structures were elucidated by NMR methods (1D and 2D NMR analysis), optical activity and ESI‐MS. Complete 1H and 13C assignments were performed for compound 2 . The antimicrobial activity was evaluated by serial microdilution assay for compounds 2 and 3 and results showed that compound 3 exhibited a significant growth inhibition at concentrations of 15.6 mg/ml (S. aureus and S. choleraesius) and 0.97 mg/mL (B. subtilis), comparable to the positive control. 相似文献
Compact single-domain antibodies (sdAbs) are nearly 13 times smaller than full-size monoclonal antibodies (mAbs) and have a number of advantages for biotechnological applications, such as small size, high specificity, solubility, stability, and great refolding capacity. Carcinoembryonic antigen (CEA) is a tumor-associated glycoprotein expressed in a variety of cancers. Detection of CEA on the tumor cell surface may be carried out using anti-CEA antibodies and conventional fluorescent dyes. Semiconductor quantum dots (QDs) are brighter and more photostable than organic dyes; they provide the possibility for labeling of different recognition molecules with QDs of different colors but excitable with the same wavelength of excitation. In this study, the abilities for specific detection of CEA expressed by tumor cells with anti-CEA sdAbs biotinylated in vitro and in vivo, as well as with anti-CEA mAbs biotinylated in vitro, were compared using flow cytometry and the conjugates of streptavidin with QDs (SA-QDs). The results demonstrated that either in vitro or in vivo biotinylated anti-CEA sdAbs are more sensitive for cell staining compared to biotinylated anti-CEA mAbs. The data also show that simultaneous use of biotinylated sdAbs with highly fluorescent SA-QDs can considerably improve the sensitivity of detection of CEA on tumor cell surfaces. 相似文献
A rapid decrease in parasitaemia remains the major goal for new antimalarial drugs
and thus, in vivo models must provide precise results concerning parasitaemia
modulation. Hydroxyethylamine comprise an important group of alkanolamine compounds
that exhibit pharmacological properties as proteases inhibitors that has already been
proposed as a new class of antimalarial drugs. Herein, it was tested the antimalarial
property of new nine different hydroxyethylamine derivatives using the green
fluorescent protein (GFP)-expressing Plasmodium berghei strain. By
comparing flow cytometry and microscopic analysis to evaluate parasitaemia
recrudescence, it was observed that flow cytometry was a more sensitive methodology.
The nine hydroxyethylamine derivatives were obtained by inserting one of the
following radical in the para position: H, 4Cl, 4-Br, 4-F, 4-CH3,
4-OCH3, 4-NO2, 4-NH2 and 3-Br. The antimalarial test showed that the compound that
received the methyl group (4-CH3) inhibited 70% of parasite growth. Our results
suggest that GFP-transfected P. berghei is a useful tool to study the recrudescence
of novel antimalarial drugs through parasitaemia examination by flow cytometry.
Furthermore, it was demonstrated that the insertion of a methyl group at the
para position of the sulfonamide ring appears to be critical for
the antimalarial activity of this class of compounds. 相似文献
The aim of this study was to evaluate an enzyme-linked immunoassay with recombinant
rhoptry protein 2 (ELISA-rROP2) for its ability to detectToxoplasma
gondii ROP2-specific IgG in samples from pregnant women. The study
included 236 samples that were divided into groups according to serological screening
profiles for toxoplasmosis: unexposed (n = 65), probable acute infection (n = 48),
possible acute infection (n = 58) and exposed to the parasite (n = 65). When an
indirect immunofluorescence assay forT. gondii-specific IgG was
considered as a reference test, the ELISA-rROP2 had a sensitivity of 61.8%,
specificity of 62.8%, predictive positive value of 76.6% and predictive negative
value of 45.4% (p = 0.0002). The ELISA-rROP2 reacted with 62.5% of the samples from
pregnant women with probable acute infection and 40% of the samples from pregnant
women with previous exposure (p = 0.0180). Seropositivity was observed in 50/57
(87.7%) pregnant women with possible infection. The results underscored that
T. gondii rROP2 is recognised by specific IgG antibodies in both
the acute and chronic phases of toxoplasmosis acquired during pregnancy. However, the
sensitivity of the ELISA-rROP2 was higher in the pregnant women with probable and
possible acute infections and IgM reactivity. 相似文献