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COMBINING MOLECULAR DOCKING
WITH RECEPTOR DOMAIN MOTIONS:
SIMULATIONS OF BINDING OF ATP TO CA-ATPase
Pyrkov T.V.*1, 2, Kosinsky Yu.A.1, Arseniev A.S.1, Priestle J.P.3,
Jacoby E.3, Efremov R.G.1
1
M.M. Shemyakin and Yu. A. Ovchinnikov Institute of Bioorganic Chemistry, RAS, Moscow, Russia;
Moscow Institute of Physics and Technology, Moscow, Russia; 3 Novartis Institutes for Biomedical
Research, Basel, Swizerland
*
Corresponding author: e-mail: [email protected]
2
Key words:
receptor flexibility, molecular dynamics, ATP-binding
SUMMARY
Motivation: Most molecular docking algorithms consider only flexibility of ligand
molecules while receptor is held rigid. At the same time it becomes evident that receptor’s
flexibility is indispensable for obtaining correct structures of protein-ligand complexes via
docking simulations.
Results: To study the effect of receptor flexibility we used the ATP – Ca-ATPase
complex. In experimental structures of the complex ATP-analogues simultaneously
interact with two active sites which are situated in different protein domains. In the apoform the distance between these sites is too large and standard docking protocols fail to
reproduce the experimental structure. We used molecular dynamics simulations of
nucleotide-binding part of Ca-ATPase to study its domain motions and generate a
representative ensemble of the receptor conformations. This approach allowed correct
prediction of the structure of the complex based on the apo-structure of the Ca-ATPase.
Availability: http://model.nmr.ru.
INTRODUCTION
Most docking algorithms incorporate ligand flexibility while keeping the protein rigid.
But structural studies of ligand-protein associations indicate that these processes are often
accompanied by conformational changes in protein structure. In many cases these changes
are related to the protein side-chain conformations although main-chain motions of
flexible loops are also observed. Moreover, for some proteins (e.g. ATP – Ca-ATPase
complex (Toyoshima et al., 2004)) changes in receptor structure involve even relative
domain motions. Such effects indicate the necessity of modeling the receptor’s flexibility
in the docking procedure.
Nowadays a lot of various approaches are applied to incorporate the flexibility of
receptor into the docking algorithm. As a rule, all of them are aimed at collecting a
representative ensemble of protein conformations. After this is done the standard docking
algorithm of a flexible ligand to rigid receptor is performed for each of them. The ways of
generating such ensemble may vary – from collecting different X-ray structures of the
target protein and stochastic exploring its conformational space (sampling method based
on bonds network, “rotamer libraries”) to taking snapshots of molecular dynamics (MD)
simulations. However, the latter approach yet has very few examples (Cavasotto et al.,
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2005) of applications to investigate the influence of global domain motions on docking
simulations. Our work is dedicated to implementation of MD simulations to sample
global domain rearrangements and local side-chain motions of ATP-binding part of CaATPase as a target for ATP docking.
METHODS
MD of ATP-binding domain of Ca-ATPase was performed for 3 experimental models:
1EUL (“open” apo-form), 1IWO (another “open” apo-form), and 1T5S (“closed” holo-form).
The GROMACS program and the GROMOS96 force field (Berendsen et al., 1995) were
used. Molecules with uncharged N- and C-termini were placed in rectangular boxes of a
simple point charge (SPC) model of water molecules with edges of 10 Å. The threedimensional periodic boundary conditions were imposed. To relax the system we performed
300 steepest descent iterations with fixed protein atoms followed by 300 conjugate-gradients
steps with fixed backbone, and finally 300 conjugate gradients steps without constraints. Then
the system was subjected to a 10 ps MD run in an NPT (constant pressure and temperature)
ensemble with fixed protein atoms. Finally, it was heated from 5 to 300 K for 60 ps in an NVT
(constant volume and temperature) ensemble. After that the production run was performed in
an NPT ensemble at 300 K. Non-bonded interactions were truncated with the twin-range
cutoff of 10/18 Å. The length of each MD trajectory exceeded 2 ns so as to allow detection of
domain rearrangements. To characterize the relative domain orientation we introduced the
interdomain angle Θ, determined as the angle between two vectors originating from the centre
of the hinge region and pointing to the centres of the N- and P-domains. For the ATP docking
experiments the snapshots of protein structure from the first 2 ns of each MD run were taken
at 8 ps interval.
To perform docking of ATP we used 8 high-resolution experimental structures of CaATPase available from the Protein Data Bank and a set of conformations from MD
trajectories. To dock ATP into the ATP-binding domain of Ca-ATPase we used the GOLD
2.0 (Jones et al., 1997) software with the scoring function “goldscore”. The docking sphere
radius was 21 Å, the origin of this sphere was atom CD1 of Phe482. 60 runs of the docking
procedure were performed for each receptor conformation. All ligands and water molecules
were removed from the receptor structure prior to docking. To retrieve a correct
conformation of ATP from a diverse set produced by docking we used our special ATPoriented ranking criterion. It is based on the analysis of ATP-protein interactions in a set of
experimental structures of ATP-protein complexes. This criterion is based mainly on
hydrophobic interactions of the adenine moiety of ATP with its protein environment.
RESULTS AND DISCUSSION
The conformation of the ligand in the N-domain was reproduced in all receptor structures
crystallized with ATP-analogs (Table 1). Also, ATP conformation close to the experimental
one was found for the apo-form 1IWO. However, docking failed to predict ATP poses in the
N-domain for other apo-forms of the receptor. This indicates that docking may yield
unsatisfactory results if the site geometry is not optimized for a particular ligand. Moreover, in
the experimental models bound ATP-analogues interact simultaneously with active sites in N
and P-domains. Mutual disposition of the latter ones corresponds to the interdomain angle Θ
~111°. This was reproduced by docking for the receptor structures taken from complexes with
ATP-analogues. But in the “open” forms of Ca-ATPase (corresponding to the values of Θ
~125°-165°) the distance between these sites is approximately twice as large as the size of
ATP molecule. That means that in such cases standard docking algorithms which consider
receptor rigid do not allow successful prediction of ligand-receptor complex structure.
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Table 1. Results ATP docking into experimental models of Ca-ATPase
PDB
Rank of correct docking solution
Ligand
Angle Θ
code
“Goldscore”
ATP-criterion
1VFP
ACP
112°
1
1
1T5T
ADP
111°
1
1
1T5S*
ACP
111°
7
1
1WPE
ADP
111°
1
1
1WPG
ADP
157°
20
1
1IWO*
–
125°
33
11
1XP5
–
155°
–
–
1EUL*
–
165°
–
–
* For these models MD simulations were performed.
To take into account flexibility of the receptor we used MD simulations to generate an
ensemble of its conformational states starting from 3 different experimental models. In
the calculated MD trajectories high-amplitude relative domain motions of type “closure”
were detected which corresponds to available experimental structures of Ca-ATPase.
For each trajectory it was shown that it is possible to obtain correct docking solutions
for the N-domain site for nearly a half of the receptor conformations. Also it is possible to
dock ATP to both active sites when the relative domain orientation is appropriate, namely
it was found that such docking solutions can be obtained for MD-conformations, with the
angle Θ < 120°. Such “closed” form was observed in each trajectory independently of the
starting conformation.
Proper scoring of the docking solutions needs a special consideration. Thus, docking
yielded correct solutions for 6 experimental Ca-ATPase structures, but ranked them top
only in 3 cases. To improve ranking we applied our ATP-criterion which ranked correct
solution top in 5 cases and improved the rank of such solution in the remaining case. The
results over all ATP poses for each MD trajectory were similar. Comparison with the
experimental structure of the ATP – Ca-ATPase complex showed that among ~14000
solutions only ~450 were correct predictions. The distribution of these correct solutions
when ranked by ATP-criterion is considerably better than that ranked by the “goldscore”
function, which is close to a random distribution (Fig. 1).
Figure 1. Ranking of docking solutions by the “goldscore” function (gray) and by ATP-criterion (black)
over all MD-conformations of the receptor (here are shown the results for 1EUL). The fraction of correct
docking solutions (~ 450 in totals) is shown.
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ACKNOWLEDGEMENTS
This work was supported by the Russian Foundation for Basic Research (grants
Nos 04-04-48875-a, 05-04-49283-а, 06-04-49194-а) and by the Russian Federation
Federal Agency for Science and Innovations (The State contract 02.467.11.3003 of
20.04.2005, grant SS-4728.2006.4).
REFERENCES
Berendsen H.J.C., van der Spoel D., Drunen R. (1995) GROMACS. Comp. Phys. Comm., 91, 43–56.
Cavasotto C.N., Orry A.J.W., Abagyan R.A. (2005) The challenge of considering receptor flexibility in
ligand docking and virtual screening. Curr. Comput. Aided Drug Des., 1, 423–440.
Jones G., Willet P., Glen R.C., Leach A.R., Taylor R. (1997) Development and validation of a genetic
algorithm for flexible docking. J. Mol. Biol., 267, 727–748.
Toyoshima C., Nomura H., Tsuda T. (2004) Lumenal gating mechanism revealed in calcium pump
crystal structures with phosphate analogues. Nature, 432, 361–368.
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