Complete nucleotide sequence of the influenza A/PR/8/34 virus NS gene and comparison with the NS genes of the A/Udorn/72 and A/FPV/Rostock/34 strains

Baez, Melvyn; Taussig, Ronald; Zazra, James J.; Young, James F.; Palese, Peter; Reisfeld, Avi; Skalka, Anna Marie

doi:10.1093/nar/8.23.5845

Cited by 89 publications

(46 citation statements)

References 42 publications

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“…If the NS2 preparation was subjected to denaturing IEF, only one band was observed as well. Matrix-assisted laser desorption ionization-time of flight mass spectrometry revealed a single species of 14,382.8, which is consistent with the molecular weight of NS2 calculated from the amino acid sequence of the native protein (14,380.3) (40). N-terminal sequencing of the purified protein yielded the sequence MDPNT as expected for InfA/PR/ 8/34 NS2.…”

Section: Resultssupporting

confidence: 78%

Structural Plasticity in Influenza Virus Protein NS2 (NEP)

Lommer

Luo

2002

Journal of Biological Chemistry

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The cellular nuclear transport machinery relies on the assembly of specialized transport complexes between soluble transport receptors, transport substrates, and additional accessory proteins. This study focuses on the structural characteristics of influenza virus protein NS2 (NEP), which interacts with the nuclear export machinery during viral replication, and has been proposed to act as an adapter molecule between the nuclear export machinery and the viral ribonucleoprotein complex. For this purpose, we have purified recombinant NS2 under nondenaturing conditions, and have investigated its structure and aggregation state using optical spectroscopy, differential scanning calorimetry, as well as hydrodynamic techniques. Our results indicate that isolated NS2 exists as a monomer in solution, and adopts a compact, but very flexible conformation, which shows characteristics of the molten globule state under near physiological conditions. Proteolytic sensitivity suggests that, despite its overall plasticity, the structure of NS2 is heterogeneous. While the C terminus of the protein adopts a relatively rigid conformation, its N terminus, which is recognized by the nuclear export machinery, exists in a highly mobile and exposed state. It is proposed that the flexibility observed in the nuclear export domain of NS2 is an important element in the recognition of substrate proteins by the nuclear export machinery.Active transport of macromolecules across the nuclear membrane is an important mechanism in the regulation of gene transcription and translation. Signal-mediated transport is typically initiated by soluble transport receptors that recognize cis-acting nuclear transport signals in their substrates and target their cargo to the nuclear pore complex. Translocation through the nuclear pore, release of substrate after transport, and directionality of the transport process are thought to be achieved mainly through action of the GTPase Ran and its regulatory factors (1-3).Influenza virus, like many viruses that replicate in the nucleus, takes advantage of these cellular pathways to translocate its genome, mRNAs, and proteins across the nuclear membrane (4, 5). Three influenza virus proteins in particular have been found to interact with the nuclear transport machinery: nucleoprotein (NP), 1 matrix protein (M1), and the nonstructural protein 2 (NS2, also called NEP). NP encapsidates the viral genomic RNA (6, 7), is able to shuttle between the nucleus and the cytoplasm (8), and is thought to catalyze the import of the viral ribonucleoprotein complexes (vRNPs) into the nucleus at early times after infection (9 -11). In addition, NP interacts with the nuclear export receptor Crm1 in vitro and has been proposed to achieve the nuclear export of progeny vRNPs during virion assembly (12).M1 is also transported into the nucleus (13) where it associates with the vRNPs (14). While it has been shown to be essential for vRNP export (14, 15), its role in this process is not clearly defined. M1 may escort the progeny vRNPs out of the ...

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Section: Resultssupporting

confidence: 78%

Structural Plasticity in Influenza Virus Protein NS2 (NEP)

Lommer

Luo

2002

Journal of Biological Chemistry

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“…The fusion peptide contains a Trp residue at fusion sequence position l l (Baez et al, 1980) and there are no Trps in tropomyosin. The stability of the fusion peptide region of fusion-tropomyosin was therefore studied by monitoring its Trp fluorescence.…”

Section: Resultsmentioning

confidence: 99%

Unfolding domains of recombinant fusion αα‐tropomyosin

et al. 1992

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The thermal unfolding of the coiled-coil a-helix of recombinant acu-tropomyosin from rat striated muscle containing an additional 80-residue peptide of influenza virus NSI protein at the N-terminus (fusion-tropomyosin) was studied with circular dichroism and fluorescence techniques. Fusion-tropomyosin unfolded in four cooperative transitions: (1) a pretransition starting at 35 "C involving the middle of the molecule; (2) a major transition at 46 "C involving no more than 36% of the helix from the C-terminus; (3) a major transition at 56 "C involving about 46% of the helix from the N-terminus; and (4) a transition from the nonhelical fusion domain at about 70 "C. Rabbit skeletal muscle tropomyosin, which lacks the fusion peptide but has the same tropomyosin sequence, does not exhibit the 56 "C or 70 "C transition. The very stable fusion unfolding domain of fusion-tropomyosin, which appears in electron micrographs as a globular structural domain at one end of the tropomyosin rod, acts as a crosslink to stabilize the adjacent N-terminal domain. The least stable middle of the molecule, when unfolded, acts as a boundary to allow the independent unfolding of the C-terminal domain at 46 "C from the stabilized N-terminal unfolding domain at 56 "C. Thus, strong localized interchain interactions in coiled-coil molecules can increase the stability of neighboring domains.

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“…Purification of virus and extraction of RNA from the virus were as described (11). Plasmid pBR322 containing cloned virus-specific double-stranded DNA was propagated in Escherichia coli C600 cells as described (12).…”

Section: Methodsmentioning

confidence: 99%

“…The synthesis ofdouble-stranded cDNA molecules from influenza virus RNA by using synthetic dodecamer nucleotide primers has been described (12). The sequence of the HA gene of influenza B/Lee/40 virus was obtained from a pair of overlapping cDNA clones.…”

Section: Methodsmentioning

confidence: 99%

Evolution of influenza A and B viruses: conservation of structural features in the hemagglutinin genes.

Krystal¹,

Elliott²,

Benz³

et al. 1982

Proc. Natl. Acad. Sci. U.S.A.

156

125

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The complete nucleotide sequence of the hemagglutinin (HA) gene of a type B influenza virus (B/Lee/40) was obtained by using cloned cDNA derived from the RNA segment. The gene is 1,882 nucleotides long and can code for a protein precursor of 584 amino acids. Structural features common to type A virus HAs are also conserved in the B virus HA. These include a hydrophobic signal peptide, hydrophobic NH2 and COOH termini of the HA2 subunit, and a HA1/HA2 cleavage site involving an arginine residue. The sequence of the B HA gene and its deduced amino acid sequence were compared to those ofa type A influenza virus (A/PR/8/34). When these two genes were aligned, it was found that 24% of the amino acids in the HAI subunits and 39% of the amino acids in the HA2 subunits are conserved. This degree of relatedness between type B virus and type A virus HAs (intertypic comparison) is similar to the homologies observed among certain type A virus HAs (intratypic comparison). A close evolutionary relationship is therefore suggested between the HAs of type A and type B influenza viruses.

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Complete nucleotide sequence of the influenza A/PR/8/34 virus NS gene and comparison with the NS genes of the A/Udorn/72 and A/FPV/Rostock/34 strains

Cited by 89 publications

References 42 publications

Structural Plasticity in Influenza Virus Protein NS2 (NEP)

Structural Plasticity in Influenza Virus Protein NS2 (NEP)

Unfolding domains of recombinant fusion αα‐tropomyosin

Evolution of influenza A and B viruses: conservation of structural features in the hemagglutinin genes.

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