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PDBsum entry 1pjm
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Protein transport
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PDB id
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1pjm
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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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Structural basis for the specificity of bipartite nuclear localization sequence binding by importin-Alpha.
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Authors
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M.R.Fontes,
T.Teh,
D.Jans,
R.I.Brinkworth,
B.Kobe.
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Ref.
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J Biol Chem, 2003,
278,
27981-27987.
[DOI no: ]
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PubMed id
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Abstract
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Importin-alpha is the nuclear import receptor that recognizes cargo proteins
carrying conventional basic monopartite and bipartite nuclear localization
sequences (NLSs) and facilitates their transport into the nucleus. Bipartite
NLSs contain two clusters of basic residues, connected by linkers of variable
lengths. To determine the structural basis of the recognition of diverse
bipartite NLSs by mammalian importin-alpha, we co-crystallized a
non-autoinhibited mouse receptor protein with peptides corresponding to the NLSs
from human retinoblastoma protein and Xenopus laevis phosphoprotein N1N2,
containing diverse sequences and lengths of the linker. We show that the basic
clusters interact analogously in both NLSs, but the linker sequences adopt
different conformations, whereas both make specific contacts with the receptor.
The available data allow us to draw general conclusions about the specificity of
NLS binding by importin-alpha and facilitate an improved definition of the
consensus sequence of a conventional basic/bipartite NLS (KRX10-12KRRK) that can
be used to identify novel nuclear proteins.
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Figure 2.
FIG. 2. Structures of complexes. A, structure of RB
peptide-m-Imp  complex. m-Imp is shown
as a ribbon diagram (yellow; drawn with the program RIBBONS
(40)). The superhelical axis of the repetitive part of the
molecule is approximately horizontal. The NLS peptide is shown
in a ball-and-stick representation, colored blue. B, structure
of N1N2 peptide-m-Imp complex, shown as in A.
The bound peptide is colored red.
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Figure 3.
FIG. 3. Peptide-importin- interaction. A,
schematic diagram of the interactions between the RB peptide and
m-Imp . Polar contacts are
shown with dashed lines, and hydrophobic contacts are indicated
by arcs with radiating spokes. The NLS peptide residues are
labeled with R. The water molecules are labeled with S. Carbon,
nitrogen, and oxygen atoms are shown in black, white, and gray,
respectively. This figure was prepared with the program LIGPLOT
(41). B, schematic diagram of the interactions between the N1N2
peptide and m-Imp , shown as in A. The
NLS peptide residues are labeled with N.
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The above figures are
reprinted
by permission from the ASBMB:
J Biol Chem
(2003,
278,
27981-27987)
copyright 2003.
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Secondary reference #1
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Title
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Structural basis of recognition of monopartite and bipartite nuclear localization sequences by mammalian importin-Alpha.
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Authors
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M.R.Fontes,
T.Teh,
B.Kobe.
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Ref.
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J Mol Biol, 2000,
297,
1183-1194.
[DOI no: ]
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PubMed id
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Figure 2.
Figure 2. (a) Structure of the Impa(70-529)-SV40 NLS
complex. Importin-a is shown as a ribbon diagram (lavender;
drawn with program RIBBONS [Carson 1997]). The superhelical axis
of the repetitive part of the molecule is approximately
horizontal. The two SV40 NLS peptides are shown in a
ball-and-stick representation; the peptide bound to the major
site is colored yellow, and the peptide bound to the minor site
is colored orange. (b) Structure of Impa(70-529)-nucleoplasmin
NLS complex, shown as in (a). The nucleoplasmin NLS peptide is
colored cyan.
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Figure 4.
Figure 4. Schematic diagram of the interactions of NLS
peptides with importin-a. The NLS backbone is shown as a black
line, with the side-chains shown as perpendicular lines
radiating from it. Individual Arm repeats of importin-a are
separated by tilted lines. Some importin-a side-chains
interacting with the NLS peptides are indicated: the invariant
asparagine residues in magenta, the invariant tryptophan
residues in green, and some nearby negatively charged residues
are shown in red. Y277 and R315 that interrupt the regular
asparagine and tryptophan array are also shown.
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The above figures are
reproduced from the cited reference
with permission from Elsevier
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Secondary reference #2
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Title
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Biophysical characterization of interactions involving importin-Alpha during nuclear import.
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Authors
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B.Catimel,
T.Teh,
M.R.Fontes,
I.G.Jennings,
D.A.Jans,
G.J.Howlett,
E.C.Nice,
B.Kobe.
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Ref.
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J Biol Chem, 2001,
276,
34189-34198.
[DOI no: ]
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PubMed id
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Figure 5.
Fig. 5. Crystal structure of the complex between Imp  (44-54) and
Imp (70-529).
A, stereoview of the electron density (drawn with the program
BOBSCRIPT (52)) in the region of the peptide bound to the major
binding site of Imp (70-529).
All peptide residues were omitted from the model and simulated
annealing run with the starting temperature of 1000 K. The
electron density map was calculated with coefficients 3|F[obs]|
 2|F[calc]|
and data between 30 and 2.8 Å resolution and contoured at
1.3 standard deviations. Superimposed is the refined model of
the peptide. B, schematic diagram of the complex. Importin- is shown as
a ribbon diagram (yellow; drawn with program RIBBONS (53)). The
superhelical axis of the repetitive part of the molecule is
approximately horizontal. The two peptides are shown in a
ball-and-stick representation; the peptide bound to the major
site is colored cyan, and the peptide bound to the minor site is
colored red. C, superposition of the Imp (44-54)
peptide (cyan) and the corresponding region of full-length
importin (magenta)
bound to the major NLS-binding site of importin- . The C
atoms of
residues 70-496 were used in the superposition (drawn with the
program RIBBONS (53)).
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Figure 7.
Fig. 7. Schematic diagram of the NLS-dependent nuclear
import pathway. Yellow, importin- ; green,
importin-  ; cyan,
NLS-containing cargo protein; magenta, Ran-GTP. For simplicity,
other factors involved in the pathway such as nuclear transport
factor-2, the nuclear export receptor for importin- , and
Ran-binding proteins have been omitted from the diagram.
Dissociation constants for the different binding events are
shown.
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The above figures are
reproduced from the cited reference
with permission from the ASBMB
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Secondary reference #3
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Title
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Autoinhibition by an internal nuclear localization signal revealed by the crystal structure of mammalian importin alpha.
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Author
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B.Kobe.
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Ref.
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Nat Struct Biol, 1999,
6,
388-397.
[DOI no: ]
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PubMed id
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Figure 4.
Figure 4. Mechanism of autoinhibition. a, Molecular surface
(probe radius 1.4 Å) of the arm repeat domain of importin
 color-coded
according to surface complementarity^49 with the N-terminal
segment (residues 44−54). Red, Sc (shape correlation
statistic) > 0.76; yellow, 0.76 > Sc > 0.3; green, 0.3 > Sc >
-0.3; light blue, Sc < -0.3. The overall Sc equals 0.75. The
N-terminal segment is shown in magenta. Drawn with the program
GRASP^46. b, Molecular surface of the arm repeat domain of
importin color-coded
according to electrostatic potential mapped to it. Lys and Arg
were assigned a single positive charge, and Glu and Asp were
assigned a single negative charge. A uniform dielectric constant
of 80 was assumed for the solvent and 2 for the protein
interior; the ionic strength was set to zero. Coloring is
continuous going from blue (potential +10 kt/e; 1 kt = 0.6 kcal,
e is the charge of an electron) through white to red (potential
-10 kt/e). The N-terminal segment is shown in magenta.
Calculated and drawn with the program GRASP^46. c, Schematic
diagram of the interactions between the autoinhibitory segment
(residues 44−54) and the arm repeat domain of mouse importin
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Polar contacts are shown with dashed lines, and hydrophobic
contacts are indicated by arcs with radiating spokes. Atom
colors: black, C; medium gray, N; light gray, O. Prepared with
the program LIGPLOT^51.
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Figure 6.
Figure 6. NLS-binding determinants. a, Schematic diagram of
the interactions between the SV40 NLS (S) and yeast Kap 50
at the larger NLS-binding site^21, prepared as Fig. 4c. b, Model
of the bipartite NLS from nucleoplasmin (residues 154−173;
green) bound to mouse importin (gray
ribbon diagram). Although the side chains of all residues of the
NLS were used in the modeling, only the side chains occupying
P1'−P2' and P1−P4 positions are shown in this figure.
Superimposed are the backbones of the autoinhibitory segment of
mouse importin (residues
44−45; magenta) and the SV40 NLS bound to the smaller
NLS-binding site in yeast Kap 50
(21) after superposition of that structure onto the structure of
mouse importin .
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The above figures are
reproduced from the cited reference
with permission from Macmillan Publishers Ltd
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