Thermostabilization of viruses via complex coacervation

Widespread vaccine coverage for viral diseases could save the lives of millions of people each year. For viral vaccines to be effective, they must be transported and stored in a narrow temperature range of 2–8 °C. If temperatures are not maintained, the vaccine may lose its potency and would no longer be effective in fighting disease; this is called the cold storage problem. Finding a way to thermally stabilize a virus and end the need to transport and store vaccines at refrigeration temperatures will increase access to life-saving vaccines. We explore the use of polymer-rich complex coacervates to stabilize viruses. We have developed a method of encapsulating virus particles in liquid complex coacervates that relies on the electrostatic interaction of viruses with polypeptides. In particular, we tested the incorporation of two model viruses; a non-enveloped porcine parvovirus (PPV) and an enveloped bovine viral diarrhea virus (BVDV) into coacervates formed from poly(lysine) and poly(glutamate). We identified optimal conditions (i.e., the relative amount of the two polypeptides) for virus encapsulation, and trends in this composition matched differences in the isoelectric point of the two viruses. Furthermore, we were able to achieve a ∼103–104-fold concentration of virus into the coacervate phase, such that the level of virus remaining in the bulk solution approached our limit of detection. Lastly, we demonstrated a significant enhancement of the stability of non-enveloped PPV during an accelerated aging study at 60 °C over the course of a week. Our results suggest the potential for using coacervation to aid in the purification and formulation of both enveloped and non-enveloped viruses, and that coacervate-based formulations could help limit the need for cold storage throughout the transportation and storage of vaccines based on non-enveloped viruses

Thermostabilization of Viruses via Complex Coacervation

Widespread vaccine coverage for viral diseases could save the lives of millions of people each year. For viral vaccines to be effective, they must be transported and stored in a narrow temperature range of 2-8°C. If temperatures are not maintained, the vaccine may lose its potency and would no longer be effective in fighting disease; this is called the cold storage problem. Finding a way to thermally stabilize a virus and end the need to transport and store vaccines at refrigeration temperatures will increase access to life-saving vaccines. We explore the use of polymer-rich complex coacervates to stabilize viruses. We have developed a method of encapsulating virus particles in liquid complex coacervates that relies on the electrostatic interaction of viruses with polypeptides. In particular, we tested the incorporation of two model viruses; a non-enveloped porcine parvovirus (PPV) and an enveloped bovine viral diarrhea virus (BVDV) into coacervates formed from poly(lysine) and poly(glutamate). We identified optimal conditions (i.e., the relative amount of the two polypeptides) for virus encapsulation, and trends in this composition matched differences in the isoelectric point of the two viruses. Furthermore, we were able to achieve a ~103 – 104-fold concentration of virus into the coacervate phase, such that the level of virus remaining in the bulk solution approached our limit of detection. Lastly, we demonstrated a significant enhancement of the stability of non-enveloped PPV during an accelerated aging study at 60°C over the course of a week. Our results suggest the potential for using coacervation to aid in the purification and formulation of both enveloped and non-enveloped viruses, and that coacervate-based formulations could help limit the need for cold storage throughout the transportation and storage of vaccines based on non-enveloped viruses

Requirement of a Membrane Potential for the Posttranslational Transfer of Proteins into Mitochondsria

Posttranslational transfer of most precursor proteins into mitochondria is dependent on energization of the mitochondria. Experiments were carried out to determine whether the membrane potential or the intramitochondrial ATP is the immediate energy source. Transfer in vitro of precursors to the ADP/ATP carrier and to ATPase subunit 9 into isolated Neurospora mitochondria was investigated. Under conditions where the level of intramitochondrial ATP was high and the membrane potential was dissipated, import and processing of these precursor proteins did not take place. On the other hand, precursors were taken up and processed when the intramitochondrial ATP level was low, but the membrane potential was not dissipated. We conclude that a membrane potential is involved in the import of those mitochondrial precursor proteins which require energy for intracellular translocatio

Effects of supplemental degradable intake protein on intake and digestibility of low-quality brome hay

The effects of increasing levels of degradable intake and digestion of low-quality brome hay were evaluated using 16 ruminally fistulated beef steers. Trends were evident for small, positive changes in total intake and digestion with increasing level of DIP supplementation. As a result, total digestible to feeding forage to provide .041, .082, and OM intake (TDOMI) increased with DIP supplementation but tended to plateau below the highest supplementation level

Influence of Prototropic Reactions on the Absorption and Fluorescence Spectra of Methyl p-dimethylaminobenzoate and Its Two Ortho Derivatives

The influence of prototropic reactions on the spectral characteristics of methyl p-dimethylaminobenzoate (I) and its o-methoxy (II) and o-hydroxy (III) derivatives has been studied using steady-state spectroscopic technique and quantum-chemical calculations. This study concerns the solvent-induced shift of the absorption, locally excited (LE) and intramolecular charge transfer (ICT) fluorescence bands in the neat tetrahydrofuran (THF) and its hydrochloric acid solutions at different HCl concentrations. On the basis of the experimental results and quantum-chemical calculations, it was shown that in a hydrochloric acid solution the studied molecules exist as a mixture of neutral, mono-, and dicationic forms. Additionally, the results of spectroscopic measurements were used to calculate, according to the Benesi-Hildebrand method, the equilibrium constants of protopropic reactions in the ground, S0, and excited, S1, states. Our findings predestine molecules I and II to be used as acid fluorescence probes in a region of 0–2.5 M of [H+] concentrations

The determinants of election to the United Nations Security Council

This is the author's accepted manuscript. The final publication is available at Springer via http://dx.doi.org/10.1007/s11127-013-0096-4.The United Nations Security Council (UNSC) is the foremost international body responsible for the maintenance of international peace and security. Members vote on issues of global importance and consequently receive perks—election to the UNSC predicts, for instance, World Bank and IMF loans. But who gets elected to the UNSC? Addressing this question empirically is not straightforward as it requires a model that allows for discrete choices at the regional and international levels; the former nominates candidates while the latter ratifies them. Using an original multiple discrete choice model to analyze a dataset of 180 elections from 1970 to 2005, we find that UNSC election appears to derive from a compromise between the demands of populous countries to win election more frequently and a norm of giving each country its turn. We also find evidence that richer countries from the developing world win election more often, while involvement in warfare lowers election probability. By contrast, development aid does not predict election

Adenosine A2A receptors modulate BDNF both in normal conditions and in experimental models of Huntington’s disease

Brain-derived neurotrophic factor (BDNF), a member of the neurotrophin family, enhances synaptic transmission and regulates neuronal proliferation and survival. Functional interactions between adenosine A2A receptors (A2ARs) and BDNF have been recently reported. In this article, we report some recent findings from our group showing that A2ARs regulate both BDNF functions and levels in the brain. Whereas BDNF (10 ng/ml) increased the slope of excitatory postsynaptic field potentials (fEPSPs) in hippocampal slices from wild-type (WT) mice, it was completely ineffective in slices taken from A2AR knock-out (KO) mice. Furthermore, enzyme immunoassay studies showed a significant reduction in hippocampal BDNF levels in A2AR KO vs. WT mice. Having found an even marked reduction in the striatum of A2AR KO mice, and as both BDNF and A2ARs have been implicated in the pathogenesis of Huntington’s disease (HD), an inherited striatal neurodegenerative disease, we then evaluated whether the pharmacological blockade of A2ARs could influence striatal levels of BDNF in an experimental model of HD-like striatal degeneration (quinolinic acid-lesioned rats) and in a transgenic mice model of HD (R6/2 mice). In both QA-lesioned rats and early symptomatic R6/2 mice (8 weeks), the systemic administration of the A2AR antagonist SCH58261 significantly reduced striatal BDNF levels. These results indicate that the presence and the tonic activation of A2ARs are necessary to allow BDNF-induced potentiation of synaptic transmission and to sustain a normal BDNF tone. The possible functional consequences of reducing striatal BDNF levels in HD models need further investigation

Macrocyclic β-Sheet Peptides That Inhibit the Aggregation of a Tau-Protein-Derived Hexapeptide

This paper describes studies of a series of macrocyclic β-sheet peptides 1 that inhibit the aggregation of a tau-protein-derived peptide. The macrocyclic β-sheet peptides comprise a pentapeptide "upper" strand, two δ-linked ornithine turn units, and a "lower" strand comprising two additional residues and the β-sheet peptidomimetic template "Hao". The tau-derived peptide Ac-VQIVYK-NH(2) (AcPHF6) aggregates in solution through β-sheet interactions to form straight and twisted filaments similar to those formed by tau protein in Alzheimer's neurofibrillary tangles. Macrocycles 1 containing the pentapeptide VQIVY in the "upper" strand delay and suppress the onset of aggregation of the AcPHF6 peptide. Inhibition is particularly pronounced in macrocycles 1a, 1d, and 1f, in which the two residues in the "lower" strand provide a pattern of hydrophobicity and hydrophilicity that matches that of the pentapeptide "upper" strand. Inhibition varies strongly with the concentration of these macrocycles, suggesting that it is cooperative. Macrocycle 1b containing the pentapeptide QIVYK shows little inhibition, suggesting the possibility of a preferred direction of growth of AcPHF6 β-sheets. On the basis of these studies, a model is proposed in which the AcPHF6 amyloid grows as a layered pair of β-sheets and in which growth is blocked by a pair of macrocycles that cap the growing paired hydrogen-bonding edges. This model provides a provocative and appealing target for future inhibitor design