Safety, efficacy and delivery of multiple nucleoside analogs via drug encapsulated carbon (DECON) based sustained drug release platform.

Acyclovir and similar nucleoside analogs form an essential frontline treatment for various herpesvirus infections of the eye. However, these drugs have low ocular retention when delivered topically and need to be administered several times every day. We have previously demonstrated that acyclovir loaded into activated carbon can be used to significantly decrease dosage frequency in a murine model of ocular infection.

Evidence for distinct mechanisms of small molecule inhibitors of filovirus entry.

Many small molecules have been identified as entry inhibitors of filoviruses. However, a lack of understanding of the mechanism of action for these molecules limits further their development as anti-filoviral agents. Here we provide evidence that toremifene and other small molecule entry inhibitors have at least three distinctive mechanisms of action and lay the groundwork for future development of anti-filoviral agents.

NMR-based metabolite studies with N amino acids.

N labeled amino acids are routinely used to label proteins or nucleic acids for study by NMR. However, NMR studies of N labeled amino acids in metabolite studies have not been pursued extensively, presumably due to line broadening present under standard experimental conditions. In this work, we show that lowering the temperature to -5 °C allows facile characterization of N-labeled amino acids.

Interplay of actin, ADP and Mg interactions with striated muscle myosin: Implications of their roles in ATPase.

The effects of Mg on the interaction between ADP, a product of the ATPase reaction, and striated muscle myosin-subfragment 1 (S1) were investigated with both functional and spectroscopic methods. Mg inhibited striated muscle myosin ATPase in the presence of F-actin. Significant effects of Mg were observed in both rate constants of NOE build-up and maximal intensities in WaterLOGSY NMR experiments as F-actin concentration increased.

Mechanism of Action of G-Quadruplex-Forming Oligonucleotide Homologous to the Telomere Overhang in Melanoma.

T-oligo, a guanine-rich oligonucleotide homologous to the 3'-telomeric overhang of telomeres, elicits potent DNA-damage responses in melanoma cells; however, its mechanism of action is largely unknown. Guanine-rich oligonucleotides can form G-quadruplexes (G4), which are stabilized by the hydrogen bonding of guanine residues. In this study, we confirmed the G4-forming capabilities of T-oligo using nondenaturing PAGE, nuclear magnetic resonance, and immunofluorescence.

Probing the metastable state of influenza hemagglutinin.

Viral entry into host cells is mediated by membrane proteins in a metastable state that transition to a more stable state upon a stimulus. For example, in the influenza envelope protein hemagglutinin (HA), the low pH in the endosome triggers a transition from the metastable prefusion conformation to the stable fusion conformation. To identify probes that interfere with HA function, here we screened a library of H7 HA peptides for inhibition of H7 HA-mediated entry.

The Polybasic Region of the Polysialyltransferase ST8Sia-IV Binds Directly to the Neural Cell Adhesion Molecule, NCAM.

Polysialic acid (polySia) is a unique post-translational modification found on a small set of mammalian glycoproteins. Composed of long chains of α2,8-linked sialic acid, this large, negatively charged polymer attenuates protein and cell adhesion and modulates signaling mediated by its carriers and proteins that interact with these carriers. PolySia is crucial for the proper development of the nervous system and is upregulated during tissue regeneration and in highly invasive cancers.

Subtle Chemical Shifts Explain the NMR Fingerprints of Oligomeric Proanthocyanidins with High Dentin Biomodification Potency.

The ability of certain oligomeric proanthocyanidins (OPACs) to enhance the biomechanical properties of dentin involves collagen cross-linking of the 1.3-4.5 nm wide space via protein-polyphenol interactions.

Structure-Function Analysis of the Non-Muscle Myosin Light Chain Kinase (nmMLCK) Isoform by NMR Spectroscopy and Molecular Modeling: Influence of MYLK Variants.

The MYLK gene encodes the multifunctional enzyme, myosin light chain kinase (MLCK), involved in isoform-specific non-muscle and smooth muscle contraction and regulation of vascular permeability during inflammation. Three MYLK SNPs (P21H, S147P, V261A) alter the N-terminal amino acid sequence of the non-muscle isoform of MLCK (nmMLCK) and are highly associated with susceptibility to acute lung injury (ALI) and asthma, especially in individuals of African descent. To understand the functional effects of SNP associations, we examined the N-terminal segments of nmMLCK by 1H-15N heteronuclear single quantum correlation (HSQC) spectroscopy, a 2-D NMR technique, and by in silico molecular modeling.

The lifestyle switch protein Bd0108 of Bdellovibrio bacteriovorus is an intrinsically disordered protein.

Bdellovibrio bacteriovorus is a δ-proteobacterium that preys upon Salmonella spp., E. coli, and other Gram-negative bacteria.

Comparison of the sensitivities of WaterLOGSY and saturation transfer difference NMR experiments.

The WaterLOGSY (WL) and saturation transfer difference (STD) NMR experiments have proven to be extremely useful techniques to characterize interactions between small molecules and large biomolecules. In this work we compare the relative sensitivities of WL and STD NMR using 3 experimental systems: ketoprofen (KET)-bovine serum albumin (BSA), tert-butyl hydroquinone (TBHQ)-hemagglutinin (HA), and chloramphenicol (CAM)-ribosome (70S). In all cases we find that WL is more sensitive than STD for a given experimental time with the ratios ranging from 3.

Distinguishing Vaccinium species by chemical fingerprinting based on NMR spectra, validated with spectra collected in different laboratories.

A method was developed to distinguish Vaccinium species based on leaf extracts using nuclear magnetic resonance spectroscopy. Reference spectra were measured on leaf extracts from several species, including lowbush blueberry (Vaccinium angustifolium), oval leaf huckleberry (Vaccinium ovalifolium), and cranberry (Vaccinium macrocarpon). Using principal component analysis, these leaf extracts were resolved in the scores plot.

Ligand screening using NMR.

NMR has proven to be an invaluable technique for identifying and characterizing ligand interactions with biomolecules. NMR-based detection of ligand binding to protein targets is described. Specifically, the use of the WaterLOGSY NMR experiment to screen mixtures of compounds from a fragment library for binding to influenza H5 hemagglutinin is detailed.

Amelogenin supramolecular assembly in nanospheres defined by a complex helix-coil-PPII helix 3D-structure.

Tooth enamel, the hardest material in the human body, is formed within a self-assembled matrix consisting mostly of amelogenin proteins. Here we have determined the complete mouse amelogenin structure under physiological conditions and defined interactions between individual domains. NMR spectroscopy revealed four major amelogenin structural motifs, including an N-terminal assembly of four α-helical segments (S9-V19, T21-P33, Y39-W45, V53-Q56), an elongated random coil region interrupted by two 3(10) helices (∼P60-Q117), an extended proline-rich PPII-helical region (P118-L165), and a charged hydrophilic C-terminus (L165-D180).

SARS-CoV heptad repeat 2 is a trimer of parallel helices.

In severe acute respiratory syndrome coronavirus, the envelope heptad repeat 2 (HR2) plays a critical role in viral entry. Moreover, HR2 is both the target for novel antiviral therapies and, as an isolated peptide, presents a potential antiviral therapeutic. The structure of HR2, as determined by NMR spectroscopy in the presence of the co-solvent trifluoroethanol (TFE), is a trimer of parallel helices, whereas the structure of HR2, as determined by X-ray crystallography, is a tetramer of anti-parallel helices.

Probing the HIV gp120 envelope glycoprotein conformation by NMR.

The HIV envelope glycoprotein gp120 plays a critical role in virus entry, and thus, its structure is of extreme interest for the development of novel therapeutics and vaccines. To date, high resolution structural information about gp120 in complex with gp41 has proven intractable. In this study, we characterize the structural properties of gp120 in the presence and absence of gp41 domains by NMR.

Diversity of mtDNA in Southern River Otter (Lontra provocax) from Argentinean Patagonia.

Lontra provocax is an endemic species from Patagonia that has been categorized as "endangered" by the International Union for Conservation of Nature. In this study, noninvasive molecular methods were used to investigate the genetic diversity and haplotype distribution of L. provocax in Argentinean Patagonia.

Chlorination diversifies Cimicifuga racemosa triterpene glycosides.

Extracts from the roots and rhizomes of black cohosh (Cimicifuga racemosa) are widely used as dietary supplements to alleviate menopausal symptoms. State-of-the-art quality control measures involve phytochemical fingerprinting of the triterpene glycosides for species identification and chemical standardization by HPLC. In the course of developing materials and methods for standardization procedures, the major C.

Yeast cox17 solution structure and Copper(I) binding.

Cox17 is a 69-residue cysteine-rich, copper-binding protein that has been implicated in the delivery of copper to the Cu(A) and Cu(B) centers of cytochrome c oxidase via the copper-binding proteins Sco1 and Cox11, respectively. According to isothermal titration calorimetry experiments, fully reduced Cox17 binds one Cu(I) ion with a K(a) of (6.15 +/- 5.

Evaluation of backbone proton positions and dynamics in a small protein by liquid crystal NMR spectroscopy.

NMR measurements of a large set of protein backbone one-bond dipolar couplings have been carried out to refine the structure of the third IgG-binding domain of Protein G (GB3), previously solved by X-ray crystallography at a resolution of 1.1 A. Besides the commonly used bicelle, poly(ethylene glycol), and filamentous phage liquid crystalline media, dipolar couplings were also measured when the protein was aligned inside either positively or negatively charged stretched acrylamide gels.