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PDBsum entry 1y2a
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Protein transport
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PDB id
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1y2a
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References listed in PDB file
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Key reference
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Title
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Phospholipid scramblase 1 contains a nonclassical nuclear localization signal with unique binding site in importin alpha.
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Authors
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M.H.Chen,
I.Ben-Efraim,
G.Mitrousis,
N.Walker-Kopp,
P.J.Sims,
G.Cingolani.
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Ref.
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J Biol Chem, 2005,
280,
10599-10606.
[DOI no: ]
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PubMed id
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Abstract
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Nuclear import of proteins containing a classical nuclear localization signal
(NLS) is an energy-dependent process that requires the heterodimer importin
alpha/beta. Three to six basic contiguous arginine/lysine residues characterize
a classical NLS and are thought to form a basic patch on the surface of the
import cargo. In this study, we have characterized the NLS of phospholipid
scramblase 1 (PLSCR1), a lipid-binding protein that enters the nucleus via the
nonclassical NLS (257)GKISKHWTGI(266). This import sequence lacks a contiguous
stretch of positively charged residues, and it is enriched in hydrophobic
residues. We have determined the 2.2 A crystal structure of a complex between
the PLSCR1 NLS and the armadillo repeat core of vertebrate importin alpha. Our
crystallographic analysis reveals that PLSCR1 NLS binds to armadillo repeats 1-4
of importin alpha, but its interaction partially overlaps the classical NLS
binding site. Two PLSCR1 lysines occupy the canonical positions indicated as P2
and P5. Moreover, we present in vivo evidence that the critical lysine at
position P2, which is essential in other known NLS sequences, is dispensable in
PLSCR1 NLS. Taken together, these data provide insight into a novel nuclear
localization signal that presents a distinct motif for binding to importin alpha.
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Figure 3.
FIG. 3. Analysis of the  IBB-importin  -PLSCR1
NLS binding interface. A, electrostatic surface potential of
importin . Acidic and basic
surfaces of the protein are in red and blue, respectively. The
PLSCR1 NLS, in blue sticks, is docked to the importin surface.
B, recognition of PLSCR1 Lys261. The conserved lysine at
position P5 is sandwiched by two importin Trp184/ Trp142
(shown in yellow) and by the PLSCR1-Trp263 (in orange).
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Figure 5.
FIG. 5. Nuclear import of PLSCR1 is blocked by triple
mutation of the residues K258A, K261A/H263A, but not by single
point mutations. Confocal fluorescence images of murine SVT2
fibroblasts transiently transfected with (top to bottom): pcDNA3
containing cDNA for wild-type (WT) human PLSCR1 or the mutants
PLSCR1 (184AAAPAA^189) or PLSCR1 (184AAAPAA^189, K258A), or
(184-AAAPAA^189, K261A) or PLSCR1(184AAAPAA^189, W263A) or
(184AAAPAA^189, 258-GAISAAWTGI266). First column PL-SCR1 label,
second column nuclear label, third column merge. Expressed human
PLSCR1 antigen was detected with monoclonal antibody 4D2 and
fluorescein isothiocyanate-conjugated goat anti-mouse IgG.
Following immunostaining and DNA labeling with propidium iodide
(PI), cells were visualized by confocal fluorescence microscopy.
Data are from a single experiment, representative of four so
performed.
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The above figures are
reprinted
by permission from the ASBMB:
J Biol Chem
(2005,
280,
10599-10606)
copyright 2005.
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