Tick-borne encephalitis virus (TBEV), a member of flaviviruses, represents a serious health threat by causing human encephalitis mainly in central and eastern Europe, Russia, and northeastern Asia. As no specific therapy is available, there is an urgent need to understand all steps of the TBEV replication cycle at the molecular level. One of the critical events is the packaging of flaviviral genomic RNA by TBEV C protein to form a nucleocapsid. We purified recombinant TBEV C protein and used a combination of physical-chemical approaches, such as size-exclusion chromatography, circular dichroism, NMR spectroscopies, and transmission electron microscopy, to analyze its structural stability and its ability to dimerize/oligomerize. We compared the ability of TBEV C protein to assemble in vitro into a nucleocapsid-like structure with that of dengue C protein.
- MeSH
- cirkulární dichroismus MeSH
- gelová chromatografie MeSH
- koncentrace vodíkových iontů MeSH
- magnetická rezonanční spektroskopie MeSH
- nukleokapsida chemie metabolismus MeSH
- rekombinantní proteiny chemie genetika izolace a purifikace MeSH
- virové proteiny chemie genetika izolace a purifikace metabolismus MeSH
- virus dengue chemie MeSH
- viry klíšťové encefalitidy chemie MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Klíčová slova
- pandemie prasečí chřipky, vakcína Pandemrix, vakcína stažena z trhu, protein hypokretinového receptoru 2,
- MeSH
- antagonisté orexinového receptoru MeSH
- hodnotící studie jako téma MeSH
- lidé MeSH
- narkolepsie * diagnóza genetika chemicky indukované komplikace prevence a kontrola MeSH
- orexinové receptory fyziologie imunologie účinky léků MeSH
- protilátky virové biosyntéza diagnostické užití farmakologie krev škodlivé účinky MeSH
- vakcíny proti chřipce * analýza aplikace a dávkování farmakologie škodlivé účinky terapeutické užití MeSH
- virové proteiny * aplikace a dávkování diagnostické užití farmakologie chemie izolace a purifikace škodlivé účinky MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- novinové články MeSH
Heterologous proteins are frequently purified by immobilized metal ion affinity chromatography (IMAC) based on their modification with a hexa-histidine affinity tag (His-tag). The terminal His-tag can, however, alter functional properties of the tagged protein. Numerous strategies for the tag removal have been developed including chemical treatment and insertion of protease target sequences in the protein sequence. Instead of using these approaches, we took an advantage of natural interaction of zinc finger domains with metal ions to purify functionally similar retroviral proteins from two different retroviruses. We found that these proteins exhibited significantly different affinities to the immobilized metal ions, despite that both contain the same type of zinc finger motif (i.e., CCHC). While zinc finger proteins may differ in biochemical properties, the multitude of IMAC platforms should allow relatively simple yet specific method for their isolation in native state.
- MeSH
- chromatografie afinitní metody MeSH
- Escherichia coli genetika MeSH
- exprese genu MeSH
- HIV-1 chemie genetika MeSH
- kovy chemie MeSH
- Masonův-Pfizerův opičí virus chemie genetika MeSH
- virové proteiny chemie genetika izolace a purifikace MeSH
- zinek analýza MeSH
- zinkové prsty MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Research Support, N.I.H., Extramural MeSH
Particles of DeltaProCANC, a fusion of capsid (CA) and nucleocapsid (NC) protein of Mason-Pfizer monkey virus (M-PMV), which lacks the amino terminal proline, were reassembled in vitro and visualized by atomic force microscopy (AFM). The particles, of 83-84 nm diameter, exhibited ordered domains based on trigonal arrays of prominent rings with center to center distances of 8.7 nm. Imperfect closure of the lattice on the spherical surface was affected by formation of discontinuities. The lattice is consistent only with plane group p3 where one molecule is shared between contiguous rings. There are no pentameric clusters nor evidence that the particles are icosahedral. Tubular structures were also reassembled, in vitro, from two HIV fusion proteins, DeltaProCANC and CANC. The tubes were uniform in diameter, 40 nm, but varied in length to a maximum of 600 nm. They exhibited left handed helical symmetry based on a p6 hexagonal net. The organization of HIV fusion proteins in the tubes is significantly different than for the protein units in the particles of M-PMV DeltaProCANC.
- MeSH
- HIV * ultrastruktura MeSH
- lidé MeSH
- makromolekulární látky MeSH
- Masonův-Pfizerův opičí virus * ultrastruktura MeSH
- mikroskopie atomárních sil MeSH
- sestavení viru MeSH
- virion * ultrastruktura MeSH
- virové proteiny izolace a purifikace metabolismus MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- práce podpořená grantem MeSH
- Research Support, N.I.H., Extramural MeSH
- MeSH
- financování vládou MeSH
- genetické struktury genetika imunologie MeSH
- genom virový imunologie účinky léků MeSH
- hepatitida C - protilátky genetika imunologie izolace a purifikace MeSH
- hepatitida C genetika klasifikace virologie MeSH
- interpretace statistických dat MeSH
- lidé MeSH
- techniky in vitro MeSH
- virové proteiny genetika imunologie izolace a purifikace MeSH
- Check Tag
- lidé MeSH
- MeSH
- sekvence nukleotidů MeSH
- virové proteiny izolace a purifikace MeSH
- viry klíšťové encefalitidy MeSH
- Geografické názvy
- Slovenská republika MeSH
