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Journal articlePaudel BP, Rueda D, 2014,
Molecular Crowding Accelerates Ribozyme Docking and Catalysis
, Journal of the American Chemical Society, Vol: 136, Pages: 16700-16703, ISSN: 1520-5126All biological processes take place in highly crowded cellular environments. However, the effect that molecular crowding agents have on the folding and catalytic properties of RNA molecules remains largely unknown. Here, we have combined single-molecule fluorescence resonance energy transfer (smFRET) and bulk cleavage assays to determine the effect of a molecular crowding agents on the folding and catalysis of a model RNA enzyme, the hairpin ribozyme. Our single-molecule data reveal that PEG favors the formation of the docked (active) structure by increasing the docking rate constant with increasing PEG concentrations. Furthermore, Mg2+ ion-induced folding in the presence of PEG occurs at concentrations ∼7-fold lower than in the absence of PEG, near the physiological range (∼1 mM). Lastly, bulk cleavage assays in the presence of the crowding agent show that the ribozyme’s activity increases while the heterogeneity decreases. Our data is consistent with the idea that molecular crowding plays an important role in the stabilization of ribozyme active conformations in vivo.
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Journal articleElderfield RA, Watson SJ, Godlee A, et al., 2014,
Accumulation of human-adapting mutations during circulation of A(H1N1)pdm09 influenza virus in humans in the United Kingdom
, Journal of Virology, Vol: 88, Pages: 13269-13283, ISSN: 0022-538XThe influenza pandemic that emerged in 2009 provided an unprecedented opportunity to study adaptation of a virus recently acquired from an animal source during human transmission. In the United Kingdom, the novel virus spread in three temporally distinct waves between 2009 and 2011. Phylogenetic analysis of complete viral genomes showed that mutations accumulated over time. Second- and third-wave viruses replicated more rapidly in human airway epithelial (HAE) cells than did the first-wave virus. In infected mice, weight loss varied between viral isolates from the same wave but showed no distinct pattern with wave and did not correlate with viral load in the mouse lungs or severity of disease in the human donor. However, second- and third-wave viruses induced less alpha interferon in the infected mouse lungs. NS1 protein, an interferon antagonist, had accumulated several mutations in second- and third-wave viruses. Recombinant viruses with the third-wave NS gene induced less interferon in human cells, but this alone did not account for increased virus fitness in HAE cells. Mutations in HA and NA genes in third-wave viruses caused increased binding to α-2,6-sialic acid and enhanced infectivity in human mucus. A recombinant virus with these two segments replicated more efficiently in HAE cells. A mutation in PA (N321K) enhanced polymerase activity of third-wave viruses and also provided a replicative advantage in HAE cells. Therefore, multiple mutations allowed incremental changes in viral fitness, which together may have contributed to the apparent increase in severity of A(H1N1)pdm09 influenza virus during successive waves.
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Journal articleJia N, Barclay WS, Roberts K, et al., 2014,
Glycomic characterization of respiratory tract tissues of ferrets IMPLICATIONS FOR ITS USE IN INFLUENZA VIRUS INFECTION STUDIES
, Journal of Biological Chemistry, Vol: 289, Pages: 28489-28504, ISSN: 0021-9258The initial recognition between influenza virus and the host cell is mediated by interactions between the viral surface protein hemagglutinin and sialic acid-terminated glycoconjugates on the host cell surface. The sialic acid residues can be linked to the adjacent monosaccharide by α2–3- or α2–6-type glycosidic bonds. It is this linkage difference that primarily defines the species barrier of the influenza virus infection with α2–3 binding being associated with avian influenza viruses and α2–6 binding being associated with human strains. The ferret has been extensively used as an animal model to study the transmission of influenza. To better understand the validity of this model system, we undertook glycomic characterization of respiratory tissues of ferret, which allows a comparison of potential viral receptors to be made between humans and ferrets. To complement the structural analysis, lectin staining experiments were performed to characterize the regional distributions of glycans along the respiratory tract of ferrets. Finally, the binding between the glycans identified and the hemagglutinins of different strains of influenza viruses was assessed by glycan array experiments. Our data indicated that the respiratory tissues of ferret heterogeneously express both α2–3- and α2–6-linked sialic acids. However, the respiratory tissues of ferret also expressed the Sda epitope (NeuAcα2-3(GalNAcβ1–4)Galβ1–4GlcNAc) and sialylated N,N′-diacetyllactosamine (NeuAcα2–6GalNAcβ1–4GlcNAc), which have not been observed in the human respiratory tract surface epithelium. The presence of the Sda epitope reduces potential binding sites for avian viruses and thus may have implications for the usefulness of the ferret in the study of influenza virus infection.
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Journal articleWarnasooriya C, Rueda D, 2014,
Single-molecule fluorescence-based studies on the dynamics, assembly and catalytic mechanism of the spliceosome
, BIOCHEMICAL SOCIETY TRANSACTIONS, Vol: 42, Pages: 1211-1218, ISSN: 0300-5127- Cite
- Citations: 9
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Journal articleGoujon C, Moncorge O, Bauby H, et al., 2014,
Transfer of the Amino-Terminal Nuclear Envelope Targeting Domain of Human MX2 Converts MX1 into an HIV-1 Resistance Factor
, JOURNAL OF VIROLOGY, Vol: 88, Pages: 9017-9026, ISSN: 0022-538X- Cite
- Citations: 80
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Journal articleTzellos S, Correia PB, Karstegl CE, et al., 2014,
A Single Amino Acid in EBNA-2 Determines Superior B Lymphoblastoid Cell Line Growth Maintenance by Epstein-Barr Virus Type 1 EBNA-2
, JOURNAL OF VIROLOGY, Vol: 88, Pages: 8743-8753, ISSN: 0022-538X- Author Web Link
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- Citations: 42
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Journal articleOjala PM, Schulz TF, 2014,
Manipulation of endothelial cells by KSHV: Implications for angiogenesis and aberrant vascular differentiation
, SEMINARS IN CANCER BIOLOGY, Vol: 26, Pages: 69-77, ISSN: 1044-579X- Author Web Link
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- Citations: 24
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Journal articleCauldwell AV, Long JS, Moncorge O, et al., 2014,
Viral determinants of influenza A virus host range
, JOURNAL OF GENERAL VIROLOGY, Vol: 95, Pages: 1193-1210, ISSN: 0022-1317- Cite
- Citations: 120
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Journal articleBajorek M, Caly L, Tran KC, et al., 2014,
The Thr205 Phosphorylation Site within Respiratory Syncytial Virus Matrix (M) Protein Modulates M Oligomerization and Virus Production
, JOURNAL OF VIROLOGY, Vol: 88, Pages: 6380-6393, ISSN: 0022-538X- Cite
- Citations: 30
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Journal articleMarriott AC, Dove BK, Whittaker CJ, et al., 2014,
Low Dose Influenza Virus Challenge in the Ferret Leads to Increased Virus Shedding and Greater Sensitivity to Oseltamivir
, PLOS ONE, Vol: 9, ISSN: 1932-6203- Author Web Link
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- Citations: 38
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