The publication data currently available has been vetted by Vanderbilt faculty, staff, administrators and trainees. The data itself is retrieved directly from NCBI's PubMed and is automatically updated on a weekly basis to ensure accuracy and completeness.
If you have any questions or comments, please contact us.
Increasing evidence supports the hypothesis that soluble misfolded protein assemblies contribute to the degeneration of postmitotic tissue in amyloid diseases. However, there is a dearth of reliable nonantibody-based probes for selectively detecting oligomeric aggregate structures circulating in plasma or deposited in tissues, making it difficult to scrutinize this hypothesis in patients. Hence, understanding the structure-proteotoxicity relationships driving amyloid diseases remains challenging, hampering the development of early diagnostic and novel treatment strategies. We report peptide-based probes that selectively label misfolded transthyretin (TTR) oligomers circulating in the plasma of TTR hereditary amyloidosis patients exhibiting a predominant neuropathic phenotype. These probes revealed that there are much fewer misfolded TTR oligomers in healthy controls, in asymptomatic carriers of mutations linked to amyloid polyneuropathy, and in patients with TTR-associated cardiomyopathies. The absence of misfolded TTR oligomers in the plasma of cardiomyopathy patients suggests that the tissue tropism observed in the TTR amyloidoses is structure-based. Misfolded oligomers decrease in TTR amyloid polyneuropathy patients treated with disease-modifying therapies (tafamidis or liver transplant-mediated gene therapy). In a subset of TTR amyloid polyneuropathy patients, the probes also detected a circulating TTR fragment that disappeared after tafamidis treatment. Proteomic analysis of the isolated TTR oligomers revealed a specific patient-associated signature composed of proteins that likely associate with the circulating TTR oligomers. Quantification of plasma oligomer concentrations using peptide probes could become an early diagnostic strategy, a response-to-therapy biomarker, and a useful tool for understanding structure-proteotoxicity relationships in the TTR amyloidoses.
Copyright © 2017 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works.
Caveolins mediate the formation of caveolae, which are small omega-shaped membrane invaginations involved in a variety of cellular processes. There are three caveolin isoforms, the third of which (Cav3) is expressed in smooth and skeletal muscles. Mutations in Cav3 cause a variety of human muscular diseases. In this work, we characterize the secondary structure, dynamics, and topology of the monomeric form of the full-length lipidated protein. Cav3 consists of a series of membrane-embedded or surface-associated helical elements connected by extramembrane connecting loops or disordered domains. Our results also reveal that the N-terminal domain undergoes a large scale pH-mediated topological rearrangement between soluble and membrane-anchored forms. Considering that roughly one-third of pathogenic mutations in Cav3 influence charged residues located in this domain, we hypothesize that this transition is likely to be relevant to the molecular basis of Cav3-linked diseases. These results provide insight into the structure of Cav3 and set the stage for mechanistic investigations of the effects of pathogenic mutations.
Copyright © 2016 Biophysical Society. Published by Elsevier Inc. All rights reserved.
UNLABELLED - Extraordinary antibodies capable of near pan-neutralization of HIV-1 have been identified. One of the broadest is antibody 10E8, which recognizes the membrane-proximal external region (MPER) of the HIV-1 envelope and neutralizes >95% of circulating HIV-1 strains. If delivered passively, 10E8 might serve to prevent or treat HIV-1 infection. Antibody 10E8, however, is markedly less soluble than other antibodies. Here, we describe the use of both structural biology and somatic variation to develop optimized versions of 10E8 with increased solubility. From the structure of 10E8, we identified a prominent hydrophobic patch; reversion of four hydrophobic residues in this patch to their hydrophilic germ line counterparts resulted in an ∼10-fold decrease in turbidity. We also used somatic variants of 10E8, identified previously by next-generation sequencing, to optimize heavy and light chains; this process yielded several improved variants. Of these, variant 10E8v4 with 26 changes versus the parent 10E8 was the most soluble, with a paratope we showed crystallographically to be virtually identical to that of 10E8, a potency on a panel of 200 HIV-1 isolates also similar to that of 10E8, and a half-life in rhesus macaques of ∼10 days. An anomaly in 10E8v4 size exclusion chromatography that appeared to be related to conformational isomerization was resolved by engineering an interchain disulfide. Thus, by combining a structure-based approach with natural variation in potency and solubility from the 10E8 lineage, we successfully created variants of 10E8 which retained the potency and extraordinary neutralization breadth of the parent 10E8 but with substantially increased solubility.
IMPORTANCE - Antibody 10E8 could be used to prevent HIV-1 infection, if manufactured and delivered economically. It suffers, however, from issues of solubility, which impede manufacturing. We hypothesized that the physical characteristic of 10E8 could be improved through rational design, without compromising breadth and potency. We used structural biology to identify hydrophobic patches on 10E8, which did not appear to be involved in 10E8 function. Reversion of hydrophobic residues in these patches to their hydrophilic germ line counterparts increased solubility. Next, clues from somatic variants of 10E8, identified by next-generation sequencing, were incorporated. A combination of structure-based design and somatic variant optimization led to 10E8v4, with substantially improved solubility and similar potency compared to the parent 10E8. The cocrystal structure of antibody 10E8v4 with its HIV-1 epitope was highly similar to that with the parent 10E8, despite 26 alterations in sequence and substantially improved solubility. Antibody 10E8v4 may be suitable for manufacturing.
Copyright © 2016, American Society for Microbiology. All Rights Reserved.
Helicobacter pylori colonizes the human stomach and is associated with an increased risk of gastric cancer and peptic ulcer disease. Analysis of H. pylori protein secretion is complicated by the occurrence of bacterial autolysis. In this study, we analyzed the exoproteome of H. pylori at multiple phases of bacterial growth and identified 74 proteins that are selectively released into the extracellular space. These include proteins known to cause alterations in host cells, antigenic proteins, and additional proteins that have not yet been studied in any detail. The composition of the H. pylori exoproteome is dependent on the phase of bacterial growth. For example, the proportional abundance of the vacuolating toxin VacA in culture supernatant is higher during late growth phases than early growth phases, whereas the proportional abundance of many other proteins is higher during early growth phases. We detected marked variation in the subcellular localization of putative secreted proteins within soluble and membrane fractions derived from intact bacteria. By providing a comprehensive view of the H. pylori exoproteome, these results provide new insights into the array of secreted H. pylori proteins that may cause alterations in the gastric environment.
Published by Elsevier B.V.
We report the synthesis and encapsulation of polyester nanosponge particles (NPs) co-loaded with tamoxifen (TAM) and quercetin (QT) to investigate the loading, release and in vitro metabolism of a dual drug formulation. The NPs are made in two variations, 4% and 8% crosslinking densities, to evaluate the effects on metabolism and release kinetics. The NP-4% formulation with a particle size of 89.3 ± 14.8 nm was found to have loading percentages of 6.91 ± 0.13% TAM and 7.72 ± 0.15% QT after targeting 10% (w/w) each. The NP-8% formulation with a particle size of 91.5 ± 9.8 nm was found to have loading percentages of 7.26 ± 0.10% TAM and 7.80 ± 0.12% QT. The stability of the formulation was established in simulated gastrointestinal fluids, and the metabolism of TAM was shown to be reduced 2-fold and 3-fold for NP-4%s and NP-8%s, respectively, while QT metabolism was reduced 3 and 4-fold. The implications for improved bioavailability of the NP formulations were supported by cytotoxicity results that showed a similar efficacy to free dual drug formulations and even enhanced anti-cancer effects in the recovery condition. This work demonstrates the suitability of the nanosponges not only as a dual release drug delivery system but also enabling a regulated metabolism through the capacity of a nanonetwork. The variation in crosslinking enables a dual release with tailored release kinetics and suggests improved bioavailability aided by a reduced metabolism.
Copyright © 2015 Elsevier B.V. All rights reserved.
Carbaboranes are increasingly studied as pharmacophores, particularly as replacements for aromatic systems. However, especially ortho-carbaborane is prone to degradation of the cluster, which hampers biological application. This study demonstrates that deboronation of the cluster may not only lead to a more active analogue, but can also improve the solubility and stability of a carbaborane-containing inhibitor. Notably, introduction of a nido-dicarbaborate cluster into the cyclooxygenase (COX) inhibitor indomethacin results in remarkably increased inhibitory potency and selectivity for COX-2 relative to the respective phenyl analogue. The first crystal structure of a carbaborane-containing inhibitor bound to COX-2 further reveals a novel binding mode for the inhibitor that is strikingly different from that of indomethacin. These results indicate that nido-dicarbaborate is a promising pharmacophore that exhibits properties which are also highly beneficial for its introduction into other inhibitor classes.
© 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.
Activation of a catalyst [IrCl(COD)(IMes)] (IMes = 1,3-bis(2,4,6-trimethylphenyl)imidazol-2-ylidene; COD = cyclooctadiene)] for signal amplification by reversible exchange (SABRE) was monitored by in situ hyperpolarized proton NMR at 9.4 T. During the catalyst-activation process, the COD moiety undergoes hydrogenation that leads to its complete removal from the Ir complex. A transient hydride intermediate of the catalyst is observed via its hyperpolarized signatures, which could not be detected using conventional nonhyperpolarized solution NMR. SABRE enhancement of the pyridine substrate can be fully rendered only after removal of the COD moiety; failure to properly activate the catalyst in the presence of sufficient substrate can lead to irreversible deactivation consistent with oligomerization of the catalyst molecules. Following catalyst activation, results from selective RF-saturation studies support the hypothesis that substrate polarization at high field arises from nuclear cross-relaxation with hyperpolarized (1)H spins of the hydride/orthohydrogen spin bath. Importantly, the chemical changes that accompanied the catalyst's full activation were also found to endow the catalyst with water solubility, here used to demonstrate SABRE hyperpolarization of nicotinamide in water without the need for any organic cosolvent--paving the way to various biomedical applications of SABRE hyperpolarization methods.
Recently, significant progress has been made in developing “stimuli-sensitive” biomaterials as a new therapeutic approach to interact with dynamic physiological conditions. Reactive oxygen species (ROS) production has been implicated in important pathophysiological events, such as atherosclerosis,aging, and cancer. ROS are often overproduced locally in diseased cells and tissues, and they individually and synchronously contribute to many of the abnormalities associated with local pathogenesis. Therefore, the advantages of developing ROS-responsive materials extend beyond site-specific targeting of therapeutic delivery, and potentially include navigating,sensing, and repairing the cellular damages via programmed changes in material properties. Here we review the mechanism and development of biomaterials with ROS-induced solubility switch or degradation, as well as their performance and potential for future biomedical applications.
We report the synthesis and photophysical properties of three biperylene-based dendrimers, which show red fluorescence in water. A fluorescence microscopy study demonstrated uptake of biperylene-based dendrimers in living cells. Our results indicate that these biperylene-based dendrimers are promising candidates in fluorescence imaging applications with the potential as therapeutic carriers.