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TitleIn vitro functional analysis of gRNA sites regulating assembly of hepatitis B virus.
Journal, issue, pagesCommun Biol, Vol. 4, Issue 1, Page 1407, Year 2021
Publish dateDec 16, 2021
AuthorsNikesh Patel / Sam Clark / Eva U Weiß / Carlos P Mata / Jen Bohon / Erik R Farquhar / Daniel P Maskell / Neil A Ranson / Reidun Twarock / Peter G Stockley /
PubMed AbstractThe roles of RNA sequence/structure motifs, Packaging Signals (PSs), for regulating assembly of an HBV genome transcript have been investigated in an efficient in vitro assay containing only core ...The roles of RNA sequence/structure motifs, Packaging Signals (PSs), for regulating assembly of an HBV genome transcript have been investigated in an efficient in vitro assay containing only core protein (Cp) and RNA. Variants of three conserved PSs, within the genome of a strain not used previously, preventing correct presentation of a Cp-recognition loop motif are differentially deleterious for assembly of nucleocapsid-like particles (NCPs). Cryo-electron microscopy reconstruction of the T = 4 NCPs formed with the wild-type gRNA transcript, reveal that the interior of the Cp shell is in contact with lower resolution density, potentially encompassing the arginine-rich protein domains and gRNA. Symmetry relaxation followed by asymmetric reconstruction reveal that such contacts are made at every symmetry axis. We infer from their regulation of assembly that some of these contacts would involve gRNA PSs, and confirmed this by X-ray RNA footprinting. Mutation of the ε stem-loop in the gRNA, where polymerase binds in vivo, produces a poor RNA assembly substrate with Cp alone, largely due to alterations in its conformation. The results show that RNA PSs regulate assembly of HBV genomic transcripts in vitro, and therefore may play similar roles in vivo, in concert with other molecular factors.
External linksCommun Biol / PubMed:34916604 / PubMed Central
MethodsEM (single particle)
Resolution3.2 - 4.2 Å
Structure data

11700

7abl

EMDB-11700, PDB-7abl:
HBV pgRNA T=4 NCP icosahedral symmetry
Method: EM (single particle) / Resolution: 3.2 Å

EMDB-11701:
HBV pgRNA T=4 NCP non-icosahedral symmetry
Method: EM (single particle) / Resolution: 4.2 Å

EMDB-11702:
HBV pgRNA T=4 NCP symmetry expansion + genome focused classification
Method: EM (single particle) / Resolution: 3.5 Å

Source
  • hepatitis b virus
KeywordsVIRUS LIKE PARTICLE / HBV / VLP / pgRNA

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