Post on 06-Jul-2015
Adviser:Dr.Hosin Keyvani
Sherko Naseri
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Today we discuss about that how adenovirus, a DNA
virus, recruits cellular and viral factors and makes use of
its own cysteine protease to regulate capsid assembly…
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With the discovery of the genetic code it was recognised
that any given viral genome was too small to encode a
single polypeptide making up the entire capsid. It was
found instead, that a capsid was constructed of one or
several smaller polypeptides arranged as symmetric
building elements
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Capsid formation may be assisted by scaffolding
structures or cellular chaperones, both of which are not
parts of the final capsid structure
Packaging of the nucleic acid can occur by at least three
different mechanisms.
(I) A proteinaceous capsid is built around a condensed
nucleic acid,
(II) a capsid shell is built and the nucleic acid
packed and condensed within the shell
(III) the nucleic acid is condensed sequentially within the
growing capsid
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Directionality of assembly can also be conferred by
limited proteolysis of capsid proteins. These reactions
are catalysed by either host or virally encoded enzymes
and may stabilise or destabilise the capsid depending on
the virus. In the case of enveloped viruses, proteolytic
processing of surface spike proteins also confers
additional functions, such as the ability to fuse
membranes
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Protein IIIa links adjacent facets by spanning the capsid
Protein IX stabilises groups of nine trimeric hexons
protein VI anchors the ring of peripentonal hexons by
bridging to the DNA core inside the capsid
Other minor proteins, such as protein VIII, X, XI or XII
have not been localised unequivocally but are likely to
occur at the inside of the capsid
The DNA is condensed with proteins V, VII and the minor
component µ and covalently linked to two terminal
proteins sitting at each a 5' end.
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Also inside the adenovirus capsid are 10 to 50
copies of the cysteine protease p23.
the cysteine protease L3/p23, located in the
internal cavity at ~10 copies per virion
This protease, encoded in the late cassette L3
as a 23 kDa polypeptide, is essential during the
assembly of the virus in the nucleus of infected
cells (Weber, 1995).
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L3/p23 has eight cysteine residues, of which at least one is needed for proteolyticactivity (Weber, 1995). A thiol-disulfide exchange mechanism is thought to provide activation of the protease during virus assembly via a disulfide-linked peptide dimer corresponding to the C terminal peptide of the protein VI precursor (Mangelet al.,1993; Webster et al., 1993).
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The first step to assembly is the formation of hexon,penton base and fibre capsomers. While fibre and penton base oligomerisations occur independently of chaperones, hexon trimers are formed in the cytoplasm by a transient,directassociation of nascent polypeptide with a chaperone of Mr 100 000 in a 1:1 complex. This assisted folding pathway is thought to help to avoid assembly errors. A completely folded hexon trimeris then imported into the nucleus perhaps in association with the precursor of protein VI
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Empty capsids have a characteristic buoyant density of
1·31 g/mL and are made up of hexon, penton base, fibre,
and precursors of proteins IIIa, VI and VIII, but lack DNA.
Empty capsids also contain scaffolding proteins L1
52/55K and perhaps minor proteins of unknown function
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the late region 1 (L1), such as the scaffolding proteins of
Mr 52 000 and 55 000, respectively, and the precursor of
protein IIIa, are involved in DNA encapsidation
It is also clear that packaging somehow depends on a
cis-acting DNA sequence containing AT-rich repeats at
the left end and trans-acting in the right of the viral
genome.
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Possibly, specific trans-acting packaging factors
recognise this and adjacent DNA regions and thus
facilitate polar encapsidation of viral DNA into empty
capsids. Whether specific proteins are added to the
capsid once it is filled with DNA in order to lock-in the
DNA is unknown
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mutant viruses that lack the functional protease (tsl)
failed at releasing fibers and penetrating into the cytosol.
The mutation in ts1 responsible for the lack of processing
has been mapped to a C-T transition resulting in a
proline to leucine exchange at position 137 of the p23
proteinase
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They are most prominent in the ts1 mutant at the
restrictive temperature and arise due to the lack of
functional protease p23. ‘Young virions’ have the same
specific density as wild particles (1·34 g/mL), but lack
processing of precursor polypeptides pIIIa, pVI, pVII,
pVIII, pµ and preterminal protein (pTP) and are devoid of
polypeptides X, XI and XII and mr55000 scaffolding
protein
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The first cofactor identified has been the 11 amino acid
C-terminal peptide of the precursor to protein VI, pVI.
This activating peptide contains a critical cysteine residue
at the C-terminal position -2, which engages in a
disulphide exchange reaction with cysteine 104 of p23 as
indicated by biochemical studies and the crystal structure
of p23 bound to the activator peptide.
Possibly, a second activator of p23, polyanions, including
DNA, either stimulate p23 activity or stabilise the
enzyme.
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It is possible that p23 enters the capsid, perhaps in
association with the viral DNA as a complex with proteins V
and VII
This interaction would directly or indirectly lead to N- and C-
terminal cleavage of pVI and the production of the activating
peptide.
This could be part of a conformational change enabling the
fully processed pVI to stably lock-in with hexon
In agreement with such a scenario, blot overlay experiments
have detected an interaction between hexon and processed
pVI, but not with the precursor of VI.
How the scaffolding proteins exit the capsid is unknown. It is
possible that they leave through openings in an incomplete
capsid, or via transient holes created by conformational
changes in the shell, analogous to bacteriophage maturation
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Lytic release of adenovirus is facilitated by a virally
encoded Mr 11 600 polypeptide also called ADP
(adenovirus death protein) of the delayed early
transcription unit E3.
Whether wild type p23 protease, which cleaves parts of
the cytoplasmic intermediate filament network, directly or
indirectly facilitates virus exit rather than cell lysis is not
known
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The role of the adenovirus protease in virus entry into cells Urs F.Greberl'2, Paul Webster, Joseph Weber3 and Ari Helenius
Virus Assembly and Disassembly: the Adenovirus Cysteine Protease as a Trigger FactorUrs F. Greber
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