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PDBsum entry 2qk2
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Protein binding
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PDB id
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2qk2
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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 of microtubule plus end tracking by xmap215, Clip-170, And eb1.
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Authors
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K.C.Slep,
R.D.Vale.
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Ref.
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Mol Cell, 2007,
27,
976-991.
[DOI no: ]
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PubMed id
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Abstract
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Microtubule plus end binding proteins (+TIPs) localize to the dynamic plus ends
of microtubules, where they stimulate microtubule growth and recruit signaling
molecules. Three main +TIP classes have been identified (XMAP215, EB1, and
CLIP-170), but whether they act upon microtubule plus ends through a similar
mechanism has not been resolved. Here, we report crystal structures of the
tubulin binding domains of XMAP215 (yeast Stu2p and Drosophila Msps), EB1 (yeast
Bim1p and human EB1), and CLIP-170 (human), which reveal diverse tubulin binding
interfaces. Functional studies, however, reveal a common property that native or
artificial dimerization of tubulin binding domains (including chemically induced
heterodimers of EB1 and CLIP-170) induces tubulin nucleation/assembly in vitro
and, in most cases, plus end tracking in living cells. We propose that +TIPs,
although diverse in structure, share a common property of multimerizing tubulin,
thus acting as polymerization chaperones that aid in subunit addition to the
microtubule plus end.
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Figure 3.
Figure 3. Structure of the Second TOG Domains from Mini
Spindles and Stu2p
Ribbon diagram of the TOG2 domain from
Msps (A) and Stu2p (B) with the six HEAT-like repeats
represented in shades of similar color and labeled A–F. The
conserved and nonconserved regions (faces A and B, respectively)
are indicated. (C) Least-squares fit of Msps (color) and Stu2p
(gray) with TOG2 domains shown in cylindrical helix
representation. (D) Individual TOG2 HEAT-like repeats are shown
for Msps and Stu2p in ribbons format in similar orientations
after global least-squares fit of each TOG2 domain. The
definitive α helix kink that defines HEAT repeats is evident in
the α2 helices of Msps HEAT-like repeats C and D and Stu2p
HEAT-like repeats C and F. (E) 90° rotations of the Msps
TOG2 domain about its long axis shown at left in ribbons for
orientation and at right in CPK representation for conservation
mapping. TOG2 residues with 80% identity across species are
represented in green, 80% conservation in yellow (see Figure
S2). (E) 2F[o] − F[c] electron density map at 1.7 Å
resolution of the Stu2p TOG2 structure contoured at 1.0 σ
showing the surface exposed and highly conserved KEKK loop of
HEAT-like repeat C. (G) 2F[o] − F[c] electron density map at
2.1 Å resolution of the Msps TOG2 structure contoured at
1.0 σ showing the surface exposed W292 residue and the buried
R295-D331 salt bridge. Inset (upper left) indicates the relative
orientation of the TOG domain (F and G). (H) Gel filtration
tubulin binding assays for wild-type (WT) and mutant Msps
TOG1-2. Single or double mutations of the conserved TOG domain
tryptophan (TOG1: W21E, TOG2: W292E) are indicated above the
chromatogram. Tubulin alone, black; Msps TOG1-2 WT alone, red.
The plot indicates absorption at 280 nm on the y axis (mAU) and
elution volume in ml along the x axis.
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Figure 4.
Figure 4. Structure of the Calponin Homology Domains of EB1
and Bim1p
Ribbon diagram of the N-terminal CH domain from
EB1 (A) and Bim1p (B) centered on the highly conserved α6
helix. (C) Least-squares fit of EB1 (color) and Bim1p (gray) CH
domains with cylindrical helix representation showing the
overall structural conservation between these two members. (D)
2F[o] − F[c] electron density map at 1.25 Å resolution
of the EB1 CH domain structure contoured at 1.0 σ showing the
highly conserved aromatic core. (E) Ninety degrees rotation of
the EB1 CH domain about the y axis shown above in ribbon format
for orientation and below in CPK representation for conservation
mapping and to summarize mutagenesis results. Center: CH domain
residues with 80% identity across species are represented in
green, 80% conservation in yellow (D and E) (see Figure S3).
Bottom row: results of CH domain mutagenesis on the ability of
EB1[1–187]-LZ to plus end track are mapped. Ablation of
microtubule association, brick red (three cluster mutants
indicated: [S16A, R17E, H18E, D19R], [K59E, K60E], and [K66E,
L67D]). No effect on microtubule plus end tracking, slate.
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The above figures are
reprinted
from an Open Access publication published by Cell Press:
Mol Cell
(2007,
27,
976-991)
copyright 2007.
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