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PDBsum entry 2f43
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480 a.a.
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471 a.a.
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124 a.a.
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References listed in PDB file
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Key reference
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Title
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Mitochondrial ATP synthase. Crystal structure of the catalytic f1 unit in a vanadate-Induced transition-Like state and implications for mechanism.
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Authors
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C.Chen,
A.K.Saxena,
W.N.Simcoke,
D.N.Garboczi,
P.L.Pedersen,
Y.H.Ko.
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Ref.
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J Biol Chem, 2006,
281,
13777-13783.
[DOI no: ]
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PubMed id
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Abstract
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ATP synthesis from ADP, P(i), and Mg2+ takes place in mitochondria on the
catalytic F1 unit (alpha3beta3gammedeltaepsilon) of the ATP synthase complex
(F0F1), a remarkable nanomachine that interconverts electrochemical and
mechanical energy, producing the high energy terminal bond of ATP. In currently
available structural models of F1, the P-loop (amino acid residues
156GGAGVGKT163) contributes to substrate binding at the subunit catalytic sites.
Here, we report the first transition state-like structure of F1 (ADP.V(i).Mg.F1)
from rat liver that was crystallized with the phosphate (P(i)) analog vanadate
(VO(3-)4 or V(i)). Compared with earlier "ground state" structures,
this new F1 structure reveals that the active site region has undergone
significant remodeling. P-loop residue alanine 158 is located much closer to
V(i) than it is to P(i) in a previous structural model. No significant movements
of P-loop residues of the subunit were observed at its analogous but
noncatalytic sites. Under physiological conditions, such active site remodeling
involving the small hydrophobic alanine residue may promote ATP synthesis by
lowering the local dielectric constant, thus facilitating the dehydration of ADP
and P(i). This new crystallographic study provides strong support for the
catalytic mechanism of ATP synthesis deduced from earlier biochemical studies of
liver F1 conducted in the presence of V(i) (Ko, Y. H., Bianchet, M., Amzel, L.
M., and Pedersen, P. L. (1997) J. Biol. Chem. 272, 18875-18881; Ko, Y. H., Hong,
S., and Pedersen, P. L. (1999) J. Biol. Chem. 274, 28853-28856).
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Figure 4.
FIGURE 4. A, plot of the difference in distance between
 -subunit atoms in the
ADP·V[i]·Mg·F[1] transition state-like
structure reported here aligned with the corresponding -subunit
atoms of the ADP·P[i]·F[1] structure (10). The two
-subunit structures were
aligned and the average distance between corresponding amino
acid atoms at each position throughout the sequences (residues
1-399 and 406-477) were calculated and plotted against residue
number. The average distance that includes all difference
calculations between all corresponding atoms in the two
structures is only 0.36 Å. In contrast, difference
calculations between corresponding atoms in the two structures
that include the P-loop (^156GGAGVGKT^163) gave an average value
of 1.0 Å (red line). B, overlay of a stick representation
of the P-loop region of the subunit of the
ADP·V[i]·Mg·F[1] transition state-like
structure reported here with that of the subunit of the
ADP·P[i]·F[1] structure (10). The conformational
differences in the P-loops of the two structures are clearly
delineated as are the relative positions of the -carbon
atom of Ala^158. In addition, the overlay shows that V[i] (red)
is much nearer the P-loop in the
ADP·V[i]·Mg·F[1] structure than is P[i]
(green)in the ADP·P[i]·F[1] structure.
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Figure 7.
FIGURE 7. Proposed critical events in the formation of ATP
catalyzed by the F[1] moiety of mitochondrial ATP synthase based
on structural work reported here, our earlier collaborative
structural studies (10), and our earlier biochemical studies
(15, 16). In the top, ADP and P[i] are both able to bind to the
catalytic F[1] moiety of rat liver ATP synthase in the absence
of Mg^2+ (10). In the middle, when Mg^2+ enters it binds to the
bound P[i], facilitating the formation of the transition state
(16). ADP and MgP[i] are then brought closer together, whereas
the methyl group of P-loop alanine 158 is brought into the
active site. The lower dielectric environment facilitates the
release of water as the ADP and MgP[i] are dehydrated to form
ATPMg (bottom).
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The above figures are
reprinted
by permission from the ASBMB:
J Biol Chem
(2006,
281,
13777-13783)
copyright 2006.
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Secondary reference #1
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Title
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The 2.8-A structure of rat liver f1-Atpase: configuration of a critical intermediate in ATP synthesis/hydrolysis.
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Authors
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M.A.Bianchet,
J.Hullihen,
P.L.Pedersen,
L.M.Amzel.
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Ref.
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Proc Natl Acad Sci U S A, 1998,
95,
11065-11070.
[DOI no: ]
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PubMed id
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Figure 1.
Fig. 1. Schematic representation of the crystallization
conditions and the bound nucleotides in the two-nucleotide
structure (Left, bovine heart, ref. 14) and the
"three-nucleotide structure" (Right, rat liver, this work).
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Figure 2.
Fig. 2. (A) Diffraction quality crystals of rat liver
F[1]. (B) Scan of an SDS/PAGE gel of redissolved F[1] crystals.
Crystals of F[1] were dissolved in 50 µl of 250 mM KP[i]
plus 5 mM EDTA (pH 7.5), and 15 µg were subjected to
SDS/PAGE in cylindrical gels. (C) Capacity of redissolved
crystals to rebind to F[1]-depleted inner membrane vesicles of
mitochondria and regain sensitivity to oligomycin, an inhibitor
of oxidative phosphorylation. Where indicated, 1 µg
oligomycin was present. (D) Relative capacity of ATP and MgCl[2]
to initiate the F[1] ATPase reaction. The decrease in optical
density of NADH at 340 nm was monitored. Reaction 1 was
initiated with 4 mM ATP, and reaction 2 was initiated with 4.8
mM MgCl[2].
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Secondary reference #2
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Title
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The three-Dimensional structure of rat liver mitochodria f1-Atpase: X-Ray diffraction studies.
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Authors
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M.Bianchet,
D.Medjahed,
J.Hulihen,
P.L.Pedersen,
L.M.Amzel.
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Ref.
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Biochim Biophys Acta, 1994,
1187,
163-164.
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PubMed id
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Secondary reference #3
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Title
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Quaternary structure of ATP synthases: symmetry and asymmetry in the f1 moiety.
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Authors
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L.M.Amzel,
M.A.Bianchet,
P.L.Pedersen.
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Ref.
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J Bioenerg Biomembr, 1992,
24,
429-433.
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PubMed id
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Secondary reference #4
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Title
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Mitochondrial ATP synthase. Quaternary structure of the f1 moiety at 3.6 a determined by X-Ray diffraction analysis.
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Authors
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M.Bianchet,
X.Ysern,
J.Hullihen,
P.L.Pedersen,
L.M.Amzel.
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Ref.
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J Biol Chem, 1991,
266,
21197-21201.
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PubMed id
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