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References listed in PDB file
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Key reference
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Title
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Insights into the mechanism of partial agonism: crystal structures of the peroxisome proliferator-Activated receptor gamma ligand-Binding domain in the complex with two enantiomeric ligands.
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Authors
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G.Pochetti,
C.Godio,
N.Mitro,
D.Caruso,
A.Galmozzi,
S.Scurati,
F.Loiodice,
G.Fracchiolla,
P.Tortorella,
A.Laghezza,
A.Lavecchia,
E.Novellino,
F.Mazza,
M.Crestani.
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Ref.
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J Biol Chem, 2007,
282,
17314-17324.
[DOI no: ]
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PubMed id
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Abstract
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The peroxisome proliferator-activated receptors (PPARs) are transcriptional
regulators of glucose and lipid metabolism. They are activated by natural
ligands, such as fatty acids, and are also targets of synthetic antidiabetic and
hypolipidemic drugs. By using cell-based reporter assays, we studied the
transactivation activity of two enantiomeric ureidofibrate-like derivatives. In
particular, we show that the R-enantiomer, (R)-1, is a full agonist of
PPARgamma, whereas the S-enantiomer, (S)-1, is a less potent partial agonist.
Most importantly, we report the x-ray crystal structures of the PPARgamma ligand
binding domain complexed with the R- and the S-enantiomer, respectively. The
analysis of the two crystal structures shows that the different degree of
stabilization of the helix 12 induced by the ligand determines its behavior as
full or partial agonist. Another crystal structure of the PPARgamma.(S)-1
complex, only differing in the soaking time of the ligand, is also presented.
The comparison of the two structures of the complexes with the partial agonist
reveals significant differences and is suggestive of the possible coexistence in
solution of transcriptionally active and inactive forms of helix 12 in the
presence of a partial agonist. Mutation analysis confirms the importance of
Leu(465), Leu(469), and Ile(472) in the activation by (R)-1 and underscores the
key role of Gln(286) in the PPARgamma activity.
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Figure 7.
FIGURE 7. C^  superposition of the
complexes with the R- and the S-enantiomer (in yellow and cyan,
respectively). Protein side chains of the complex with the
R-enantiomer are shown in green; the correspondent side-chains
are in pink for the complex with the S-enantiomer.
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Figure 11.
FIGURE 11. Superposition of the C^ traces of the complexes
of PPAR  with the R-enantiomer
(green), the S-enantiomer (cyan), and rosiglitazone (purple).
Putative contacts are shown in parentheses.
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The above figures are
reprinted
by permission from the ASBMB:
J Biol Chem
(2007,
282,
17314-17324)
copyright 2007.
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Secondary reference #1
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Title
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Ligand binding and co-Activator assembly of the peroxisome proliferator-Activated receptor-Gamma.
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Authors
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R.T.Nolte,
G.B.Wisely,
S.Westin,
J.E.Cobb,
M.H.Lambert,
R.Kurokawa,
M.G.Rosenfeld,
T.M.Willson,
C.K.Glass,
M.V.Milburn.
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Ref.
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Nature, 1998,
395,
137-143.
[DOI no: ]
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PubMed id
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Figure 1.
Figure 1: Crystal structure of the apo-PPAR- bold gamma- .
a, Ribbons drawing of the apo-PPAR-  LBD,
amino acids 207–476. The nomenclature of the helices is based
on the RXR-  crystal
structure^16. -Helices
are light blue;  -strands
are green; loops are brown. b, A worm backbone tracing of PPAR-
,
with a surface representation of the unoccupied van der Waals
space in the ligand-binding site. We determined the 'unoccupied'
volume by fitting in virtual atoms that did not occupy the van
der Waals surface of the protein. The total unoccupied volume is
 1,300
Å^3. c, Sequence alignment of the human PPAR LBDs. Amino
acids that are conserved between PPARs ,
and
 are
in yellow; the secondary structure that the sequence adopts in
PPAR- is
shown in red boxes for -helices
and blue arrows for -strands.
Residues involved in rosiglitazone binding are underlined. The
sequence alignment predicts that several residues involved in
direct interactions of PPAR- with
ligand are not conserved in the other PPAR subtypes, and
explains the specificity of TZDs for PPAR- :
residue H323 is not found in PPAR- ,
and Q286 is not found in PPAR- or
PPAR- .
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Figure 3.
Figure 3: SRC-1 interactions with PPAR- bold gamma-.
a, A sigma-weighted 2 F [o]– F [c] omit electron-density
map is shown contoured at 1.0  for
the area surrounding the rosiglitazone ligand. b, A ribbons
drawing of the PPAR- LBD
dimer and SRC-1, including the ligand rosiglitazone. The two
PPAR- monomers
are blue and green and the two SRC-1 interacting helices are
yellow. The structure of SRC-1 was determined from amino acids
628–640 and 684–703 and was crystallographically refined.
Very weak electron density from residues 670 to 684 was visible
but was not crystallographically refined and is shown as a
dashed line. SRC-1 amino acids 642–669 were disordered and not
structurally determined. The diagram shows how one SRC-1
molecule, with two interacting domains, forms a complex with a
PPAR- homodimer.
The dashed line connecting the two structurally determined
domains of SRC-1 is the proposed connection between these two
domains. c, The binding of SRC-1 (amino acids 628–642) to the
LXXLL-binding site of PPAR- .
SRC-1 is coloured: yellow, carbon; blue, nitrogen; red, oxygen.
The ribbon backbone of the PPAR- LBD
is in green. PPAR- amino
acids binding to the LXXLL helix are also shown in green. d,
Residues H631–T640 of SRC-1 are coloured as in c, with an
electrostatic surface of PPAR- showing
the coactivator-binding site. E471 and K301 side chains result
in the red (negative) and blue (positive) charges on the surface
of the coactivator-binding site at the N and C termini of the
SRC-1 helix, respectively. e, Residues H687–E696 of SRC-1 are
coloured as in c, with an electrostatic surface of PPAR- showing
the coactivator-binding site. f, Amino acids L465–K474 of the
PPAR- AF-2
helix of one monomer in the apo structure are shown in: green,
carbon; blue, nitrogen; red, oxygen, with an electrostatic
surface of PPAR- showing
the coactivator-binding site. E471 and K301 side chains result
in the red (negative) and blue (positive) charges on the surface
at the N and C terminus of the other PPAR- monomer.
This figure shows how one monomer in the apo crystal structure
orientates its AF-2 helix into the coactivator-binding site of
another crystallographically related monomer.
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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 #2
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Title
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A peroxisome proliferator-Activated receptor gamma ligand inhibits adipocyte differentiation.
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Authors
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J.L.Oberfield,
J.L.Collins,
C.P.Holmes,
D.M.Goreham,
J.P.Cooper,
J.E.Cobb,
J.M.Lenhard,
E.A.Hull-Ryde,
C.P.Mohr,
S.G.Blanchard,
D.J.Parks,
L.B.Moore,
J.M.Lehmann,
K.Plunket,
A.B.Miller,
M.V.Milburn,
S.A.Kliewer,
T.M.Willson.
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Ref.
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Proc Natl Acad Sci U S A, 1999,
96,
6102-6106.
[DOI no: ]
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PubMed id
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Figure 2.
Fig. 2. GW0072 is a PPAR  ligand with
a unique functional profile. (A) Dose response on the PPAR -GAL4
chimera for GW0072 (  circle )
and GW0072 plus 100 nM rosiglitazone (  ).
Reporter activity was expressed as the % of the maximal
activation by 1 µM rosiglitazone. GW0072 demonstrates
competitive antagonism of rosiglitazone but retains weak agonist
activity at µM concentrations. (B) Activity on full-length
PPAR 2 for 100
nM rosiglitazone (TZD), 10 µM GW0072 (GW), and 100 nM
rosiglitazone plus 10 µM GW0072 (TZD + GW). Vehicle was
0.1% DMSO. Reporter activity was expressed as the % of the
maximal activation by 1 µM rosiglitazone. (C-F) The
functional activity of GW0072 is paralleled by its effects on
coactivator recruitment to PPAR 2 in a
mammalian two-hybrid assay. GW0072 (GW) (10 µM)
antagonizes recruitment of the coactivators CBP and SRC1
promoted by 1 µM rosiglitazone (TZD). GW0072 (GW) (10
µM) does not recruit the corepressors NCoR or SMRT.
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Figure 4.
Fig. 4. GW0072 defines an additional class of nuclear
receptor ligands. Agonist ligands (gray) shift the AF-2 helix
(yellow) into a position that stabilizes recruitment of
coactivator (green) to the receptor ligand-binding domain
(white). Antagonist ligands (gray) bind to the receptor by using
the same epitopes, but their larger size shifts the AF-2 helix
into a position that displaces the coactivator. Antagonist
ligands also recruit corepressor (red) to the receptor
ligand-binding domain. The partial agonist GW0072 (gray) binds
to its receptor by using different epitopes, such that it does
not directly interact with the AF-2 helix.
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