Bandwidth Enhancement of Planar Terahertz Metasurfaces via Overlapping of Dipolar Modes

Plasmonics - We present enhancement of operational bandwidths of planar terahertz metasurfaces by incorporating a complex unit cell that consists of a pair of concentric ring resonators. The inner...     

Computational and Functional Analyses of a Small-Molecule Binding Site in ROMK

The renal outer medullary potassium channel (ROMK, or Kir1.1, encoded by KCNJ1) critically regulates renal tubule electrolyte and water transport and hence blood volume and pressure. The discovery of loss-of-function mutations in KCNJ1 underlying renal salt and water wasting and lower blood pressure has sparked interest in developing new classes of antihypertensive diuretics targeting ROMK. The recent development of nanomolar-affinity small-molecule inhibitors of ROMK creates opportunities for exploring the chemical and physical basis of ligand-channel interactions required for selective ROMK inhibition. We previously reported that the bis-nitro-phenyl ROMK inhibitor VU591 exhibits voltage-dependent knock-off at hyperpolarizing potentials, suggesting that the binding site is located within the ion-conduction pore. In this study, comparative molecular modeling and in silico ligand docking were used to interrogate the full-length ROMK pore for energetically favorable VU591 binding sites. Cluster analysis of 2498 low-energy poses resulting from 9900 Monte Carlo docking trajectories on each of 10 conformationally distinct ROMK comparative homology models identified two putative binding sites in the transmembrane pore that were subsequently tested for a role in VU591-dependent inhibition using site-directed mutagenesis and patch-clamp electrophysiology. Introduction of mutations into the lower site had no effect on the sensitivity of the channel to VU591. In contrast, mutations of Val¹⁶⁸ or Asn¹⁷¹ in the upper site, which are unique to ROMK within the Kir channel family, led to a dramatic reduction in VU591 sensitivity. This study highlights the utility of computational modeling for defining ligand-ROMK interactions and proposes a mechanism for inhibition of ROMK.     

Controlling the reactivity of radical intermediates by coenzyme B(12)-dependent methylmalonyl-CoA mutase

Adenosylcobalamin or coenzyme B(12)-dependent enzymes are members of the still relatively small group of radical enzymes and catalyse 1,2-rearrangement reactions. A member of this family is methylmalonyl-CoA mutase, which catalyses the isomerization of methylmalonyl-CoA to succinyl-CoA and, unlike the others, is present in both bacteria and animals. Enzymes that catalyse some of the most chemically challenging reactions are the ones that tend to deploy radical chemistry. The use of radical intermediates in an active site lined with amino acid side chains that threaten to extinguish the reaction by presenting alternative groups for abstraction poses the conundrum of how the enzymes control their reactivity. In this review, insights into this issue that have emerged from kinetic, mutagenesis and structural studies are described for methylmalonyl-CoA mutase.     

Bone marrow transplantation in a patient with drug-induced aplastic anemia

A 23-year-old woman gravely ill with Pseudomonas septicemia secondary to presumed drug-induced bone marrow aplasia received marrow transplantation from two male HL-A identical sibling donors. She had a successful engraftment with excellent but temporary clinical improvement. Subsequently she succumbed to graft-versus-host disease manifested by Pseudomonas and Candida albicans septicemia, cytomegalovirus pneumonitis, three phases of dermatitis, nausea, vomiting, dysphagia, diarrhea, fever, edema and bone pain, with gradual but complete graft suppression by the 74th day after the transplantation. A second marrow transplant on the 70th day was unsuccessful.     

An insecticidal GroEL protein with chitin binding activity from Xenorhabdus nematophila

Xenorhabdus nematophila secretes insecticidal proteins to kill its larval prey. We have isolated an approximately 58-kDa GroEL homolog, secreted in the culture medium through outer membrane vesicles. The protein was orally insecticidal to the major crop pest Helicoverpa armigera with an LC50 of approximately 3.6 microg/g diet. For optimal insecticidal activity all three domains of the protein, apical, intermediate, and equatorial, were necessary. The apical domain alone was able to bind to the larval gut membranes and manifest low level insecticidal activity. At equimolar concentrations, the apical domain contained approximately one-third and the apical-intermediate domain approximately one-half bioactivity of that of the full-length protein. Interaction of the protein with the larval gut membrane was specifically inhibited by N-acetylglucosamine and chito-oligosaccharides. Treatment of the larval gut membranes with chitinase abolished protein binding. Based on the three-dimensional structural model, mutational analysis demonstrated that surface-exposed residues Thr-347 and Ser-356 in the apical domain were crucial for both binding to the gut epithelium and insecticidal activity. Double mutant T347A,S356A was 80% less toxic (p < 0.001) than the wild type protein. The GroEL homolog showed alpha-chitin binding activity with Kd approximately 0.64 microm and Bmax approximately 4.68 micromol/g chitin. The variation in chitin binding activity of the mutant proteins was in good agreement with membrane binding characteristics and insecticidal activity. The less toxic double mutant XnGroEL showed an approximately 8-fold increase of Kd in chitin binding assay. Our results demonstrate that X. nematophila secretes an insecticidal GroEL protein with chitin binding activity.     

Improved nearest-neighbor parameters for the stability of RNA/DNA hybrids under a physiological condition

The stability of Watson–Crick paired RNA/DNA hybrids is important for designing optimal oligonucleotides for ASO (Antisense Oligonucleotide) and CRISPR (Clustered Regularly Interspaced Short Palindromic Repeats)–Cas9 techniques. Previous nearest-neighbour (NN) parameters for predicting hybrid stability in a 1 M NaCl solution, however, may not be applicable for predicting stability at salt concentrations closer to physiological condition (e.g. ∼100 mM Na⁺ or K⁺ in the presence or absence of Mg²⁺). Herein, we report measured thermodynamic parameters of 38 RNA/DNA hybrids at 100 mM NaCl and derive new NN parameters to predict duplex stability. Predicted ΔG°₃₇ and T_m values based on the established NN parameters agreed well with the measured values with 2.9% and 1.1°C deviations, respectively. The new results can also be used to make precise predictions for duplexes formed in 100 mM KCl or 100 mM NaCl in the presence of 1 mM Mg²⁺, which can mimic an intracellular and extracellular salt condition, respectively. Comparisons of the predicted thermodynamic parameters with published data using ASO and CRISPR–Cas9 may allow designing shorter oligonucleotides for these techniques that will diminish the probability of non-specific binding and also improve the efficiency of target gene regulation.     

Interactions of amyloid Aβ(1–42) peptide with self‐assembled peptide nanospheres

In this work we have probed the interactions of the amyloid Aβ(1–42) peptide with self‐assembled nanospheres. The nanospheres were formed by self‐assembly of a newly developed bolaamphiphile bis(N‐alpha‐amido‐methionine)‐1,8 octane dicarboxylate under aqueous conditions. It was found that the interactions of the Aβ(1–42) peptide with the nanospheres were concentration as well as pH dependent and the peptide largely adopts a random coil structure upon interacting with the nanospheres. Further, upon incorporation with the nanospheres, we observed a relative diminution in the aggregation of Aβ(1–42) at low concentrations of Aβ(1–42). The interactions between the nanospheres and the Aβ(1–42) peptide were investigated by atomic force microscopy, transmission electron microscopy, circular dichroism, FTIR and fluorescence spectroscopy, and the degree of fibrillation in the presence and absence of nanospheres was monitored by the Thioflavine T assay. We believe that the outcome from this work will help further elucidate the binding properties of Aβ peptide as well as designing nanostructures as templates for further investigating the nucleation and fibrillation process of Aβ‐like peptides. Copyright © 2010 European Peptide Society and John Wiley & Sons, Ltd.     

Characterization of the Cardiac Overexpression of HSPB2 Reveals Mitochondrial and Myogenic Roles Supported by a Cardiac HspB2 Interactome

Small Heat Shock Proteins (sHSPs) are molecular chaperones that transiently interact with other proteins, thereby assisting with quality control of proper protein folding and/or degradation. They are also recruited to protect cells from a variety of stresses in response to extreme heat, heavy metals, and oxidative-reductive stress. Although ten human sHSPs have been identified, their likely diverse biological functions remain an enigma in health and disease, and much less is known about non-redundant roles in selective cells and tissues. Herein, we set out to comprehensively characterize the cardiac-restricted Heat Shock Protein B-2 (HspB2), which exhibited ischemic cardioprotection in transgenic overexpressing mice including reduced infarct size and maintenance of ATP levels. Global yeast two-hybrid analysis using HspB2 (bait) and a human cardiac library (prey) coupled with co-immunoprecipitation studies for mitochondrial target validation revealed the first HspB2 “cardiac interactome” to contain many myofibril and mitochondrial-binding partners consistent with the overexpression phenotype. This interactome has been submitted to the Biological General Repository for Interaction Datasets (BioGRID). A related sHSP chaperone HspB5 had only partially overlapping binding partners, supporting specificity of the interactome as well as non-redundant roles reported for these sHSPs. Evidence that the cardiac yeast two-hybrid HspB2 interactome targets resident mitochondrial client proteins is consistent with the role of HspB2 in maintaining ATP levels and suggests new chaperone-dependent functions for metabolic homeostasis. One of the HspB2 targets, glyceraldehyde 3-phosphate dehydrogenase (GAPDH), has reported roles in HspB2 associated phenotypes including cardiac ATP production, mitochondrial function, and apoptosis, and was validated as a potential client protein of HspB2 through chaperone assays. From the clientele and phenotypes identified herein, it is tempting to speculate that small molecule activators of HspB2 might be deployed to mitigate mitochondrial related diseases such as cardiomyopathy and neurodegenerative disease.