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PDBsum entry 1v1i
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Contents |
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* Residue conservation analysis
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References listed in PDB file
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Key reference
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Title
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Adenovirus fibre shaft sequences fold into the native triple beta-Spiral fold when n-Terminally fused to the bacteriophage t4 fibritin foldon trimerisation motif.
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Authors
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K.Papanikolopoulou,
S.Teixeira,
H.Belrhali,
V.T.Forsyth,
A.Mitraki,
M.J.Van raaij.
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Ref.
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J Mol Biol, 2004,
342,
219-227.
[DOI no: ]
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PubMed id
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Abstract
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Adenovirus fibres are trimeric proteins that consist of a globular C-terminal
domain, a central fibrous shaft and an N-terminal part that attaches to the
viral capsid. In the presence of the globular C-terminal domain, which is
necessary for correct trimerisation, the shaft segment adopts a triple
beta-spiral conformation. We have replaced the head of the fibre by the
trimerisation domain of the bacteriophage T4 fibritin, the foldon. Two different
fusion constructs were made and crystallised, one with an eight amino acid
residue linker and one with a linker of only two residues. X-ray
crystallographic studies of both fusion proteins shows that residues 319-391 of
the adenovirus type 2 fibre shaft fold into a triple beta-spiral fold
indistinguishable from the native structure, although this is now resolved at a
higher resolution of 1.9 A. The foldon residues 458-483 also adopt their natural
structure. The intervening linkers are not well ordered in the crystal
structures. This work shows that the shaft sequences retain their capacity to
fold into their native beta-spiral fibrous fold when fused to a foreign
C-terminal trimerisation motif. It provides a structural basis to artificially
trimerise longer adenovirus shaft segments and segments from other trimeric
beta-structured fibre proteins. Such artificial fibrous constructs, amenable to
crystallisation and solution studies, can offer tractable model systems for the
study of beta-fibrous structure. They can also prove useful for gene therapy and
fibre engineering applications.
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Figure 2.
Figure 2. Space-filling models of the long-linker (a) and
short-linker (b) adenovirus fibre shaft-fibritin foldon chimeras
in the same orientation as Figure 1.
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Figure 3.
Figure 3. Ribbon diagrams of the long-linker (a) and
short-linker (b) adenovirus fibre shaft-fibritin foldon chimeras
in the same orientation as Figure 1. The N and the C termini in
the A subunits are labelled.
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The above figures are
reprinted
by permission from Elsevier:
J Mol Biol
(2004,
342,
219-227)
copyright 2004.
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Secondary reference #1
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Title
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Formation of highly stable chimeric trimers by fusion of an adenovirus fiber shaft fragment with the foldon domain of bacteriophage t4 fibritin.
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Authors
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K.Papanikolopoulou,
V.Forge,
P.Goeltz,
A.Mitraki.
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Ref.
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J Biol Chem, 2004,
279,
8991-8998.
[DOI no: ]
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PubMed id
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Figure 1.
FIG. 1. A, schematic representation of the domain structure
of the proteins described in this study. 1, the stable
adenovirus fiber fragment (fiber residues 319-582). Residues
belonging to the shaft domain (Val319-Gly392) are symbolized
with a rectangle, and residues belonging to the globular head
(Leu399-Glu582) are symbolized with a circle. Residues 393-398
(Asn-Lys-Asn-Asp-Asp-Lys) form the linker that connects the two
parts and are drawn in italics. 2, the chimeric protein that
comprises the fibritin foldon domain (fibritin residues
Gly457-Leu483, oval shape) fused to the C terminus of the shaft
domain with use of the natural linker between the two domains.
To avoid confusion, the numbers corresponding to the fibritin
residues are underlined. The residues Gly-Ser, highlighted in
bold, are not part of the coding sequence and are introduced as
a result of the cloning strategy. 3, the chimeric protein
carrying the foldon domain at the C-terminal end of the shaft
domain without the use of the natural linker sequence. 4, the
chimeric protein carrying the foldon domain at the N-terminal
end of the shaft domain. The residues Gly-Ser-Gly, highlighted
in bold, do not belong to the coding sequence and are introduced
as a result of the cloning strategy. B, amino acid sequences of
the fiber shaft residues 319-392 and of the fibritin foldon
residues 457-483. The fiber shaft sequence repeat numbers
(repeats 18-22 according to Ref. 9) are indicated on the left.
The repeats are not aligned.
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Figure 7.
FIG. 7. Proteolytic digestion of the purified protein. A,
time course of proteolytic digestion of the purified protein
with the linker in the absence of SDS. Chymotrypsin was added to
a purified protein solution (0.3 mg/ml in 50 mM phosphate
buffer, pH 7, 1 mM EDTA) at a 1:10 ratio w/w, and digestion was
allowed to proceed for the following times: 30 min (lane 2), 1 h
(lane 3), 3 h (lane 4), and 5 h (lane 5). Lane 1, protein
control in the absence of protease; lane 6, protease control in
the absence of protein. B, time course of proteolytic digestion
in the presence of SDS. The digestion protocol is the same as in
A, except that the protein was incubated in the presence of 0.1%
SDS before adding the protease. Lane 1, protein control in the
absence of protease. Lanes 2-5, same times of digestion as the
corresponding lanes in A. In lane 6, the protein was boiled for
4 min in 0.1% SDS, cooled on ice for 1 min, and then the
protease was added. Digestion was subsequently carried out at 22
°C for 30 min (lane 7), 1 h (lane 8), and 3 h (lane 9).
Aliquots were taken at the corresponding times, and sample
buffer was added to give a 2% final SDS concentration; the
samples were frozen at -20 °C until electrophoresis. The
samples have not been boiled prior to loading in the gels. All
gels (15%) were stained with Coomassie Blue. Lane M, molecular
mass markers.
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The above figures are
reproduced from the cited reference
with permission from the ASBMB
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Secondary reference #2
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Title
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A triple beta-Spiral in the adenovirus fibre shaft reveals a new structural motif for a fibrous protein.
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Authors
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M.J.Van raaij,
A.Mitraki,
G.Lavigne,
S.Cusack.
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Ref.
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Nature, 1999,
401,
935-938.
[DOI no: ]
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PubMed id
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Figure 1.
Figure 1 The adenovirus fibre fold. The two parts of this
figure are shown in stereo. a, Chain trace of the C  atoms
of one of the trimers present in the asymmetric unit. Chains A,
B and C are shown in red, blue and green respectively. Every
tenth residue of the shaft domain in chain A is numbered. b, The
shaft domain as present in the crystal. The N and C termini of
chain A are labelled.
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Figure 3.
Figure 3 Interactions in the shaft. a, Hydrophobic core. The
main chain of residues 356-389 of monomers A (red), B (blue) and
C (yellow) is shown. Residues contributing to the central
hydrophobic core (orange) and residues contributing to the
peripheral hydrophobic patches (green) are also given. b,
Diagram of the hydrogen-bonding network within one monomer and
between adjacent monomers. Residues are labelled according to
Fig. 2. Intra-chain (dotted lines) and inter-chain hydrogen
bonds (solid black lines) that are conserved in each of the four
repeats of the structure are shown.
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The above figures are
reproduced from the cited reference
with permission from Macmillan Publishers Ltd
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Secondary reference #3
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Title
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Structure of bacteriophage t4 fibritin m: a troublesome packing arrangement.
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Authors
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S.V.Strelkov,
Y.Tao,
M.M.Shneider,
V.V.Mesyanzhinov,
M.G.Rossmann.
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Ref.
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Acta Crystallogr D Biol Crystallogr, 1998,
54,
805-816.
[DOI no: ]
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PubMed id
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Figure 1.
Fig. 1. Schematic diagrams of the wild-type tibritin and the engineered
fragments. The black boxes represent the individual coiled-coil
segments and the ovals show the N- and C- terminal domains.
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Figure 7.
Fig. 7. Crystal contacts. Surfaces of trimeric molecules were calculated
with program
GRASP
(Nicholls
et al.,
1993) and colored by
approximate electrostatic potential revealing the location of acidic
(red) and basic (blue) side chains. (a) Contact between the C-
terminal domains of molecules A and B. (b) Contact between the
coiled-coil
domain of A and the C-terminal domain of C. There is a
similar contact between the coiled-coil domain of C and the C-
terminal domain of B. (c) The C-terminal domain of A and the
coiled-coil domain of C make no contacts. The same is true for the
C-terminal domain of C and the coiled-coil
domain of B.
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The above figures are
reproduced from the cited reference
with permission from the IUCr
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