Tetrahydropyrimidine derivatives inhibit binding of a Tat-like, arginine-containing peptide, to HIV TAR RNA in vitro

AbstractThe ability of a small molecule, 2-methyl,4-carboxy,5-hydroxy-3,4,5,6-tetrahydropyrimidine (THP(A)), which accumulates intracellularly in various streptomyces, to inhibit the interaction of Tat peptide (R52) with TAR RNA is presented. Using gel-shift assay, we found that the inhibition constant Ki of THP(A) is 50–100 nM, which is in the range of the binding constants of Tat peptide and protein. THP(A) is ∼ 106 times more tightly bound than the free l-arginine. The high binding affinity may be attributed to the special delocalized positive charge on the NCN group and the hydroxyl group at the 5 position of this molecule. A model for THP(A)-TAR interaction, analogous to the arginine guanidinum group-TAR interaction, is presented. The relatively high uptake of THP(A) by mammalian cells warrants in vivo Tat/TAR inhibition studies

Hypertext versus Footnotes: High School English Learners’ Online Reading Recall

This study considers forty adolescent English Language Learners who read a passage online containing additional information available through either hypertext links or footnotes. Participants were attending a special high school for English learners at the time of the study. Two versions of the text were offered, one with hypertext and the other with footnotes, and participants were randomly assigned to the footnote or hypertext condition. Answers to multiple choice questions showed no significant difference between groups in recall of the reading under the two conditions, in contrast with an earlier study of learners in higher education settings whose recall of reading with hypertext was significantly lower than with footnotes. Learners’ ratings of perceived comprehensibility of the 2 texts was also not significantly different. Additional interpretive data came from focus group interviews involving all of the participants

The Calbindin-D28k binding site on inositol monophosphatase may allow inhibition independent of the lithium site of action

Among numerous reported biochemical effects the lithium-inhibitable enzyme inositol-monophosphatase (IMPase) remains a viable target for lithium's therapeutic mechanism of action. Calbindin-D28k (calbindin) interacts with IMPase enhancing its activity. In the present study in silico modeling of IMPase-calbindin binding using the program MolFit indicated that the 55-66 amino acid segment of IMPase anchors calbindin via Lys59 and Lys61 with a glutamate in between (Lys-Glu-Lys motif). The model further suggested that the Lys-Glu-Lys motif interacts with residues Asp24 and Asp26 of calbindin. Indeed, we found that differently from wildtype calbindin, IMPase was not activated by mutated calbindin in which Asp24 and Asp26 were replaced by alanine. Calbindin's effect was significantly reduced by a peptide with the sequence of amino acids 58-63 of IMPase (peptide 1) and by six amino-acid peptides including at least part of the Lys-Glu-Lys motif. The three amino-acid peptide Lys-Glu-Lys or five amino-acid peptides containing this motif were ineffective. Intracerebroventricular administration of peptide 1 resulted in a significant antidepressant-like reduced immobility in the Porsolt forced swim test (FST) compared with mice treated with a scrambled peptide or artificial cerebrospinal fluid. Based on the sequence of peptide 1, and to potentially increase the peptide's stability, cyclic and linear pre-cyclic analog peptides were synthesized. One cyclic and one linear pre-cyclic analog peptides exhibited an inhibitory effect on calbindin-activated brain IMPase activity in vitro. These findings may lead to the development of molecules capable of inhibiting IMPase activity at an alternative site than that of lithium

Identification of the Amino Acid Subsets Accounting for the Ligand Binding Specificity of a Glutamate Receptor

AbstractIn a situation so far unique among neurotransmitter receptors, glutamate receptors share amino acid sequence similarities with the bacterial periplasmic binding proteins (PBPs). On the basis of the primary structure similarity of two bacterial periplasmic proteins (lysine/arginine/ornithine- and phosphate-binding proteins) with the chick cerebellar kainate-binding protein (KBP), a member of the ionotropic glutamate receptor family, we have generated a three-dimensional model structure of the KBP extracellular domain. By an interplay between homology modeling and site-directed mutagenesis, we have investigated the kainate binding properties of 55 different mutants (corresponding to 43 positions) and studied the interactions of some of these mutants with various glutamatergic ligands. As a result, we present here the subsets of amino acids accounting for the binding free energies and specificities of KBP for kainate, glutamate, and CNQX and propose a three-dimensional model, at the microarchitectural level, of the glutamatergic binding domain

The Evolutionary Origin of the Runx/CBFbeta Transcription Factors – Studies of the Most Basal Metazoans

BACKGROUND. Members of the Runx family of transcriptional regulators, which bind DNA as heterodimers with CBFβ, are known to play critical roles in embryonic development in many triploblastic animals such as mammals and insects. They are known to regulate basic developmental processes such as cell fate determination and cellular potency in multiple stem-cell types, including the sensory nerve cell progenitors of ganglia in mammals. RESULTS. In this study, we detect and characterize the hitherto unexplored Runx/CBFβ genes of cnidarians and sponges, two basal animal lineages that are well known for their extensive regenerative capacity. Comparative structural modeling indicates that the Runx-CBFβ-DNA complex from most cnidarians and sponges is highly similar to that found in humans, with changes in the residues involved in Runx-CBFβ dimerization in either of the proteins mirrored by compensatory changes in the binding partner. In situ hybridization studies reveal that Nematostella Runx and CBFβ are expressed predominantly in small isolated foci at the base of the ectoderm of the tentacles in adult animals, possibly representing neurons or their progenitors. CONCLUSION. These results reveal that Runx and CBFβ likely functioned together to regulate transcription in the common ancestor of all metazoans, and the structure of the Runx-CBFβ-DNA complex has remained extremely conserved since the human-sponge divergence. The expression data suggest a hypothesis that these genes may have played a role in nerve cell differentiation or maintenance in the common ancestor of cnidarians and bilaterians.National Science Foundation (IBN-0212773, FP-91656101-0); Boston University SPRInG (20-202-8103-9); Israel Science Foundation (825/07

Ideais e realidade: uma aula reservada para crianças autistas bilíngües = Ideals and reality: a self-contained class for bilingual autistic children

Este estudo qualitativo abordou as experiências de crianças bilíngües, selecionadas com desordem do espectro autista em uma sala de aula de Educação Infantil Especial Bilíngüe. Os dados incluíram observação ao vivo, gravações em vídeo e entrevistas com os funcionários e pais. Apesar da dedicação dos professores, dos funcionários e da família, os desafios incluíram limitações lingüísticas de clínicos gerais, uso inconsistente de espanhol e/ou inglês, falta de compreensão dos pais sobre a natureza do autismo e demanda irreal de funcionários. O ambiente da barulhenta sala de aula, às vezes refletia aprendizagem, que freqüentemente significava transtorno. A preparação profissional de professores também era inadequad

Dual array EEG-fMRI : An approach for motion artifact suppression in EEG recorded simultaneously with fMRI

Objective: Although simultaneous recording of EEG and MRI has gained increasing popularity in recent years, the extent of its clinical use remains limited by various technical challenges. Motion interference is one of the major challenges in EEG-fMRI. Here we present an approach which reduces its impact with the aid of an MR compatible dual-array EEG (daEEG) in which the EEG itself is used both as a brain signal recorder and a motion sensor. Methods: We implemented two arrays of EEG electrodes organized into two sets of nearly orthogonally intersecting wire bundles. The EEG was recorded using referential amplifiers inside a 3 T MR-scanner. Virtual bipolar measurements were taken both along bundles (creating a small wire loop and therefore minimizing artifact) and across bundles (creating a large wire loop and therefore maximizing artifact). Independent component analysis (ICA) was applied. The resulting ICA components were classified into brain signal and noise using three criteria: 1) degree of two-dimensional spatial correlation between ICA coefficients along bundles and across bundles; 2) amplitude along bundles vs. across bundles; 3) correlation with ECG. The components which passed the criteria set were transformed back to the channel space. Motion artifact suppression and the ability to detect interictal epileptic spikes following daEEG and Optimal Basis Set (OBS) procedures were compared in 10 patients with epilepsy. Results: The SNR achieved by daEEG was 11.05 +/- 3.10 and by OBS was 8.25 +/- 1.01 (p <0.00001). In 9 of 10 patients, more spikes were detected after daEEG than after OBS (p <0.05). Significance: daEEG improves signal quality in EEG-fMRI recordings, expanding its clinical and research potential. (C) 2016 Elsevier Inc. All rights reserved.Peer reviewe

Conformation-specific inhibitory anti-MMP-7 monoclonal antibody sensitizes pancreatic ductal adenocarcinoma cells to chemotherapeutic cell kill

Matrix metalloproteases (MMPs) undergo post-translational modifications including pro-domain shedding. The activated forms of these enzymes are effective drug targets, but generating potent biological inhibitors against them remains challenging. We report the generation of anti-MMP-7 inhibitory monoclonal antibody (GSM-192), using an alternating immunization strategy with an active site mimicry antigen and the activated enzyme. Our protocol yielded highly selective anti-MMP-7 monoclonal antibody, which specifically inhibits MMP-7′ s enzyme activity with high affinity (IC50 = 132 ± 10 nM). The atomic model of the MMP-7-GSM-192 Fab complex exhibited antibody binding to unique epitopes at the rim of the enzyme active site, sterically preventing entry of substrates into the catalytic cleft. In human PDAC biopsies, tissue staining with GSM-192 showed characteristic spatial distribution of activated MMP-7. Treatment with GSM-192 in vitro induced apoptosis via stabilization of cell surface Fas ligand and retarded cell migration. Co-treatment with GSM-192 and chemotherapeutics, gemcitabine and oxaliplatin elicited a synergistic effect. Our data illustrate the advantage of precisely targeting catalytic MMP-7 mediated disease specific activity

FitEM2EM—Tools for Low Resolution Study of Macromolecular Assembly and Dynamics

Studies of the structure and dynamics of macromolecular assemblies often involve comparison of low resolution models obtained using different techniques such as electron microscopy or atomic force microscopy. We present new computational tools for comparing (matching) and docking of low resolution structures, based on shape complementarity. The matched or docked objects are represented by three dimensional grids where the value of each grid point depends on its position with regard to the interior, surface or exterior of the object. The grids are correlated using fast Fourier transformations producing either matches of related objects or docking models depending on the details of the grid representations. The procedures incorporate thickening and smoothing of the surfaces of the objects which effectively compensates for differences in the resolution of the matched/docked objects, circumventing the need for resolution modification. The presented matching tool FitEM2EMin successfully fitted electron microscopy structures obtained at different resolutions, different conformers of the same structure and partial structures, ranking correct matches at the top in every case. The differences between the grid representations of the matched objects can be used to study conformation differences or to characterize the size and shape of substructures. The presented low-to-low docking tool FitEM2EMout ranked the expected models at the top