Small molecules that enhance the catalytic efficiency of HLA-DM

HLA-DM (DM) plays a critical role in Ag presentation to CD4 T cells by catalyzing the exchange of peptides bound to MHC class II molecules. Large lateral surfaces involved in the DM:HLA-DR (DR) interaction have been defined, but the mechanism of catalysis is not understood. In this study, we describe four small molecules that accelerate DM-catalyzed peptide exchange. Mechanistic studies demonstrate that these small molecules substantially enhance the catalytic efficiency of DM, indicating that they make the transition state of the DM:DR/peptide complex energetically more favorable. These compounds fall into two functional classes: two compounds are active only in the presence of DM, and binding data for one show a direct interaction with DM. The remaining two compounds have partial activity in the absence of DM, suggesting that they may act at the interface between DM and DR/peptide. A hydrophobic ridge in the DMbeta1 domain was implicated in the catalysis of peptide exchange because the activity of three of these enhancers was substantially reduced by point mutations in this area.     

Highly Sensitive Surface Plasmon Resonance Biosensors Utilizing Prism-Waveguide Configuration for Detection of Alzheimer Disease Biomarker

Plasmonics - Alzheimer’s disease (AD) is a disease in cognitive regions in the human brain. Also the detection of AD in the early stage is too important. In this regard, a surface plasmon...     

Inhibitory effects of deferasirox on the structure and function of bovine liver catalase: a spectroscopic and theoretical study

Deferasirox (DFX), as an oral chelator, is used for treatment of transfusional iron overload. In this study, we have investigated the effects of DFX as an iron chelator, on the function and structure of bovine liver catalase (BLC) by different spectroscopic methods of UV–visible, fluorescence, and circular dichroism (CD) at two temperatures of 25 and 37 °C. In vitro kinetic studies showed that DFX can inhibit the enzymatic activity in a competitive manner. K _I value was calculated 39 nM according to the Lineweaver–Burk plot indicating a high rate of inhibition of the enzyme. Intrinsic fluorescence data showed that increasing the drug concentrations leads to a significant decrease in the intrinsic emission of the enzyme indicating a significant change in the three-dimensional environment around the chromophores of the enzyme structure. By analyzing the fluorescence quenching data, it was found that the BLC has two binding sites for DFX and the values of binding constant at 25 and 37 °C were calculated 1.7 × 10⁷ and 3 × 10⁷ M^?1, respectively. The static type of quenching mechanism is involved in the quenching of intrinsic emission of enzyme. The thermodynamic data suggest that hydrophobic interactions play a major role in the binding reaction. UV–vis spectroscopy results represented the changes in tryptophan (Trp) absorption and Soret band spectra, which indicated changes in Trp and heme group position caused by the drug binding. Also, CD data represented that high concentrations of DFX lead to a significant decreasing in the content of β-sheet and random coil accompanied an increasing in α-helical content of the protein. The molecular docking results indicate that docking may be an appropriate method for prediction and confirmation of experimental results and also useful for determining the binding mechanism of proteins and drugs. According to above results, it can be concluded that the DFX can chelate the Fe(III) on the enzyme active site leading to changes in the function and structure of catalase which can be considered as a side effect of this drug and consequently has an important role in hepatic complications and fibrosis.     

Synthesis and in vitro evaluation of glycosyl derivatives of luteinizing hormone-releasing hormone (LHRH)

Luteinizing hormone-releasing hormone (LHRH) analogues are used extensively for the treatment of various hormone-dependent diseases. However, none of the currently marketed derivatives can be administered orally. Modification of peptide sequences by attachment of carbohydrate moieties is a promising strategy that may increase the metabolic stability of the target peptide and enhance its transport across cell membranes, subsequently improving peptide bioavailability. In this study, either the N- or C-terminus of the LHRH peptide was altered by attachment of carbohydrate moieties. Caco-2 cells were chosen as an in vitro model to investigate both the permeability and stability of the new LHRH analogues. Our findings show that conjugating sugar moieties to the N-terminus of the LHRH peptide significantly increased both permeability and metabolic stability of most of the modified LHRH derivatives.     

Modular construction of multifunctional bioresponsive cell-targeted nanoparticles for gene delivery

Multifunctional and modular block copolymers prepared from biocompatible monomers and linked by a bioreducible disulfide linkage have been prepared using a combination of ring-opening and atom-transfer radical polymerizations (ATRP). The presence of terminal functionality via ATRP allowed cell-targeting folic acid groups to be attached in a controllable manner, while the block copolymer architecture enabled well-defined nanoparticles to be prepared by a water-oil-water double emulsion procedure to encapsulate DNA with high efficiency. Gene delivery assays in a Calu-3 cell line indicated specific folate-receptor-mediated uptake of the nanoparticles, and triggered release of the DNA payload via cleavage of the disulfide link resulted in enhanced transgene expression compared to nonbioreducible analogues. These materials offer a promising and generic means to deliver a wide variety of therapeutic payloads to cells in a selective and tunable way.     

Leber congenital amaurosis associated with AIPL1: challenges in ascribing disease causation, clinical findings, and implications for gene therapy

Leber Congenital Amaurosis (LCA) and Early Childhood Onset Severe Retinal Dystrophy are clinically and genetically heterogeneous retinal disorders characterised by visual impairment and nystagmus from birth or early infancy. We investigated the prevalence of sequence variants in AIPL1 in a large cohort of such patients (n = 392) and probed the likelihood of disease-causation of the identified variants, subsequently undertaking a detailed assessment of the phenotype of patients with disease-causing mutations. Genomic DNA samples were screened for known variants in the AIPL1 gene using a microarray LCA chip, with 153 of these cases then being directly sequenced. The assessment of disease-causation of identified AIPL1 variants included segregation testing, assessing evolutionary conservation and in silico predictions of pathogenicity. The chip identified AIPL1 variants in 12 patients. Sequencing of AIPL1 in 153 patients and 96 controls found a total of 46 variants, with 29 being novel. In silico analysis suggested that only 6 of these variants are likely to be disease-causing, indicating a previously unrecognized high degree of polymorphism. Seven patients were identified with biallelic changes in AIPL1 likely to be disease-causing. In the youngest subject, electroretinography revealed reduced cone photoreceptor function, but rod responses were within normal limits, with no measurable ERG in other patients. An increasing degree and extent of peripheral retinal pigmentation and degree of maculopathy was noted with increasing age in our series. AIPL1 is significantly polymorphic in both controls and patients, thereby complicating the establishment of disease-causation of identified variants. Despite the associated phenotype being characterised by early-onset severe visual loss in our patient series, there was some evidence of a degree of retinal structural and functional preservation, which was most marked in the youngest patient in our cohort. This data suggests that there are patients who have a reasonable window of opportunity for gene therapy in childhood.     

Novel coumarin derivatives bearing N-benzyl pyridinium moiety: Potent and dual binding site acetylcholinesterase inhibitors

A novel series of coumarin derivatives linked to benzyl pyridinium group were synthesized and biologically evaluated as inhibitors of both acetylcholinesterase (AChE) and butyrylcholinesterase (BuChE). The enzyme inhibitory activity of synthesized compounds was measured using colorimetric Ellman’s method. It was revealed that compounds 3e, 3h, 3l, 3r and 3s have shown higher activity compared with donepezil hydrochloride as standard drug. Most of the compounds in these series had nanomolar range IC₅₀ in which compound 3r (IC₅₀ = 0.11 nM) was the most active compound against acetylcholinesterase enzyme.     

QTL mapping of complex binary traits in an advanced intercross line

An advanced intercross line (AIL) is an easier and more cost-effective approach compared to recombinant inbred lines for fine mapping of quantitative trait loci (QTL) identified by F(2) designs. In an AIL, a complex binary trait can be mapped through analysis of either continuously distributed proxy traits for the liability of the binary trait or the liability itself, the latter presenting the greater statistical challenge. In another work, we successfully applied both approaches in an AIL to fine map previously identified QTL underlying anatomical parameters of the cardiac inter-atrial septum including patent foramen ovale. Here, we describe the statistical methods that we used to analyse complex binary traits in our AIL design. This is achieved using a likelihood-based method, with the expectation-maximisation algorithm allowing use of standard logistic regression methods for model fitting.     

Are Long-Range Structural Correlations Behind the Aggregration Phenomena of Polyglutamine Diseases?

We have characterized the conformational ensembles of polyglutamine peptides of various lengths (ranging from to ), both with and without the presence of a C-terminal polyproline hexapeptide. For this, we used state-of-the-art molecular dynamics simulations combined with a novel statistical analysis to characterize the various properties of the backbone dihedral angles and secondary structural motifs of the glutamine residues. For (i.e., just above the pathological length for Huntington''s disease), the equilibrium conformations of the monomer consist primarily of disordered, compact structures with non-negligible -helical and turn content. We also observed a relatively small population of extended structures suitable for forming aggregates including - and -strands, and - and -hairpins. Most importantly, for we find that there exists a long-range correlation (ranging for at least residues) among the backbone dihedral angles of the Q residues. For polyglutamine peptides below the pathological length, the population of the extended strands and hairpins is considerably smaller, and the correlations are short-range (at most residues apart). Adding a C-terminal hexaproline to suppresses both the population of these rare motifs and the long-range correlation of the dihedral angles. We argue that the long-range correlation of the polyglutamine homopeptide, along with the presence of these rare motifs, could be responsible for its aggregation phenomena.