Introduction to Molecular Dynamics Simulations

Roland H. Stote Institut de Chimie LC3-UMR 7177 Universit Louis Pasteur Strasbourg France

1EA5

Title Native Acetylcholinesterase (E.C. 3.1.1.7) From Torpedo Californica At 1.8A Resolution Classification Cholinesterase Compound Mol_Id: 1; Molecule: Acetylcholinesterase; Chain: A; Ec: 3.1.1.7 Exp. Method X-ray Diffraction

Macromolecules in motion
Local motions
(0.01 5 , 10-15 10-1 s) Atomic Fluctuations Sidechain motions Loop motions (1 10 , 10-9 1 s) Helix motions Domain motions Subunit motions (> 5 , 10-7 104 s) helix-coil Transitions Dissociation/Association Folding and unfolding

Rigid body motions

Large scale motions

Biological function requires exibility (dynamics)

Energy Minimization
"E a # b # "E b # c # "E c ### "E MIN $ 0 # fin

c b

Central idea of Molecular Dynamics simulations

Biological activity is the result of time dependent interactions between molecules and these interactions occur at the interfaces such as protein-protein, protein-NA, protein-ligand. Macroscopic observables (laboratory) are related to microscopic behavior (atomic level). Time dependent (and independent) microscopic behavior of a molecule can be calculated by molecular dynamics simulations.

Molecular Dynamics Simulations

One of the principal tools for modeling proteins, nucleic acids and their complexes. Stability of proteins Folding of proteins Molecular recognition by:proteins, DNA, RNA, lipids, hormones STP, etc. Enzyme reactions Rational design of biologically active molecules (drug design) Small and large-scale conformational changes. determination and construction of 3D structures (homology, Xray diffraction, NMR) Dynamic processes such as ion transport in biological systems.

Molecular dynamics simulations

Approximate the interactions in the system using simplied models (fast calculations). Include in the model only those features that are necessary to describe the system. In the case of molecular dynamics simulations, this means a potential energy function that models the basic interactions. Allows one to gain insight into situations that are impossible to study experimentally Run computer experiments. Ask the question What if? The method allows the prediction of the static and dynamic properties of molecules directly from the underling interactions between the molecules.

Classical Dynamics
Newtons Equations of motion

dvi d 2ri Fi = mi ! ai = mi ! = m! 2 dt dt

Fi = !"i E
Position, speed and acceleration are functions of time ri(t); vi(t); ai(t) The force is related to the acceleration and, in turn, to the potential energy Integration of the equations of motion => initial structure : ri(t=0); initial distribution of velocities: vi(t=0)

Dynamics: calculating trajectories

Trajectory: positions as function of time: ri (t) How does one determine ri (t) from Fi = mi ai ?

Simple case where acceleration is constant

dvi d 2ri Fi = mi ! ai = mi ! = m! 2 dt dt
v = at + v0

a=

dv dt

Simple case: motion of a particle in one dimension

Acceleration: If a is constant af(t) Speed:

a=

dv dt

v(t) = at + v0
v(t) = dx(t) dt

Position: The trajectory x(t) obtained by integration taking into account the initial positions and velocities (x0 et v0)

t2 x(t) = v ! t + x0 = a ! + v0t + x0 2

Balistic trajectory
Initial conditions are x(0) = z(0) = 0 vx (0) = vo cos vz (0) = vo sin In the x direction ax = 0 vx (t) = vo cos x(t) = vo cos t
Z

V0 X

In the z direction, one has to take into account gravity az = g vz (t) = vo sin - gt z(t)= vo sin t g t2 /2 z = ax -b x2 : the trajectory in the (x,z) plane is parabolic

Potential Energy
E(R) = 1 2 1 2 Kb ( b " b0 ) + ! K# (# " # 0 ) + ! K$ (1 + cos(n$ " % )) 1, 2 pairs 2 angles 2 dihedrals

4 8 .( + 1 2 ( + 6 1 6 ' ij - 3 qi q j 6 0* ' ij 9 + ! 54& ij 0* - " * - 3 + * r i, j 6 0) rij , ) ij , 3 &Drij 6 / 2 7 :

The energy is a function of the positions ri Therefore the acceleration is a function of the positions Since the positions vary as a function of time ri(t), so does the acceleration, ai(t)

Numerical Integration
Taylor series development

3 t2 ' t + O(t 4 ) x(t) = x0 + v0 t + a0 + a 0 2 3!

If we know x at time t, after passage of a certain time, t, we can nd x(t+t)

F(t) !t 2 F ' (t) !t 3 x(t + !t) = x(t) + v(t)!t + + + O(!t 4 ) m 2 m 3!

We restart from the coordinates x(t+t) to get x(t+2t) To pass from x(t) to x(t+t) is to carry out 1 step of dynamics The change in velocity v(t) to v(t+t) can be calculated in the same manner The acceleration is recalculate from E(r) at each step

Acceleration as a function of time

Acceleration: calculated from the force, that is, from the derivative of the potential energy, including at t=0

Potential Energy
E(RN ) =
1,2 pairs 2

1 dE(RN ) ai (t) = ! m dri (t)

1 Kb (b " b0 ) +
2 angles 2

1 2 K# (# " # 0 ) +

dihedrals

! K$ (1 + cos(n$ " % ))

4 .( ' + 12 ( ' + 6 1 q q 8 6 ij ij i j 6 + ! 54& ij 0* - " * - 3 + * r - 3 &Dr 9 0* rij ) ij , 2 ij 6 i, j 6 /) , 7 :

Principle of the trajectory

t0+4 t t0+2t t0+t t0 t0+7t

Integration algorithms Verlet, Velocity Verlet LeapFrog, Beeman

Choice of the algorithm:
Energy conservation Calculation time (least expensive) Integration time step as large as possible

Trajectory of a macromolecule
Initial positions x0
PDB le Xray NMR Model

Initial velocities v0

Coupled to the temperature

3 m v2 NkT = ! i i 2 2 i

Acceleration
Calculated from the force, that is, from the derivative of the potential energy.

a=!

1 dE m dr

Relationship between velocities and temperature

Temperature specifies the thermodynamic state of the system Important concept in dynamics simulations. Temperature is related to the microscopic description of simulations through the kinetic energy Kinetic energy is calculated from the atomic velocities.

3 m v2 NkT = ! i i 2 2 i

Molecular Dynamics Simulation programs AMBER CHARMM NAMD POLY-MD etc Potential energy function parameter les contain the numerical constants needed to evaluate forces and energies

http://www.pharmacy.umaryland.edu/faculty/amackere/research.html

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Molecular Dynamics
Calculation of forces Displacement t=t New set of coordinates

Practical Aspects
Choice of integration timestep t
> As long as possible compatible with a correct numerical integration > 1 to 2 fs (10-15 s)

Calculating nonbonded Interactions: consumes the most CPU time

> The cost (CPU) is proportional to N2 (N number of atoms) > Truncation

0 4 *# & 1 2 # & 6 2 ,% " ij ( ) % " ij ( / + qi q j 2 7 14! ij ,% r ( % r ( / !r 5 i, j 2 $ ij ' . ij 2 +$ ij ' 3 6

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Nonbonded Energy Terms

Electrostatic Forces + r + r +

E(R) =

van der Waals Forces 1

1, 2 pairs

! r

Kb ( b " b0 ) +

angles

1 ! 2 K (# " # )
#
0

dihedrals

K$ (1 + cos(n$ " % ))

4 8 .( + 1 2 ( + 6 1 6 0* ' ij - " * ' ij - 3 + qi q j 6 + ! 54& ij 0* * r - 3 &Dr 9 rij , i, j 6 0 ) ij , 2 3 ij 6 /) 7 :

Truncation
Switch
Bring the potential to zero between ron and roff. The potential is not modied for r < ron and equals zero for r > roff

Shift
Modify the potential over the entire range of distances in order to bring the potential to zero for r > rcut

Long-range electrostatic interactions

Ewald summation Multipole methods (Extended electrostatics model)

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Treatment of solvent
Implicit: The macromolecule interacts only with itself, but the electrostatic interactions are modied to account for the solvent

E elec (r ) = A

qi q j

!r

All solvent effects are contained in the dielectric constant

Vacuum =1 Proteins = 2-20 Water = 80

Treatment of solvent
Explicit representation The macromolecule is surrounded by solvent molecules (water, ions) with which the macromolecule interacts. Specic nonbond interactions are calculated

0 4 *# & 1 2 # & 6 2 ,% " ij ( ) % " ij ( / + qi q j 2 7 14! ij ,% r ( % r ( / r 5 i, j 2 $ ij ' . ij 2 +$ ij ' 3 6

In this case, one must use =1. More correct (fewer approximations) but more expensive

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Periodic boundary conditions

For explicit representation of solvent The boundaries of the system must be represented

For periodic system

Permits the modeling of very large systems, but introduces a level of periodicity not present in nature.

Boundary Conditions
Solvation sphere: nite system Around the entire macromolecule Around the active site

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Some properties that can be calculated from a trajectory

Average Energie moyenne RMS between 2 structures (ex : initial structure)

Fluctuations of atomic des positions

Temperature Fators

Radius of gyration

Copyright " www.ch.embnet.org/MD_tutorial" Reproduction ULP Strasbourg. Autorisation CFC - Paris

Protocol for an MD simulation

Initial Coordinates
X-ray diffraction or NMR coordinates from the Protein Data Bank Coordinates constructed by modeling (homology)

Treatment of non-bonded interactions

Choice of truncation

Treatment of solvent
implicit: choice of dielectric constant Implicit: advanced treatment of solvent: Generalized Born, ACE, EEF1 explicit: solvation protocol

If using explicit treatment of solvent ->boundary condition

Periodic boundary conditions (PBC) Solvation sphere Active site dynamics Time step for integration of equations of motion

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Steps of a molecular dynamics simulation

An application of Molecular Dynamics Simulations The acetylcholinesterase story

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Acetylcholinesterase
Acetylcholinesterase (AChE) is an enzyme that hydrolyzes ACh to acetate and choline to inactivate the neurotransmitter A very fast enzyme, approaching diffusion controlled. Inhibitors are utilized in the treatment of various neurological diseases, including Alzheimers disease. Organophosphorus compounds serve as potent insecticides by selectively inhibiting insect AChE.

Neuromuscular junction: motor neurons : muscle cells

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1EA5

Title Native Acetylcholinesterase (E.C. 3.1.1.7) From Torpedo Californica At 1.8A Resolution Classification Cholinesterase Compound Mol_Id: 1; Molecule: Acetylcholinesterase; Chain: A; Ec: 3.1.1.7 Exp. Method X-ray Diffraction

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Access of ligands to the active site is blocked --> requires uctuations

Secondary channels open transiently: Identied by MD simulations

Molecular Dynamics Simulation of Acetylcholinesterase

10 ns simulations Protein obtained from the Protein Data Bank (PDB) Structure solved by x-ray crystallography Solvated in a cubic box of water Ions added to neutralize the system Periodic Boundary Conditions Treatment of Long-Range electrostatic interactions Total of 8289 solute atoms and 75615 solvent atoms

Biophysical Journal Volume 81 715-724 (2001) Acc. Chem. Research 35 332-340 (2002)

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Molecular Dynamics Simulation of Acetylcholinesterase

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Effect of the His44Ala mutation on the Nucleocapsid protein from the HIV virus - NC(35-50) Working at the interface of theory and experiment

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Primary function of NC is to bind nucleic acids

The life cycle of the HIV-1 retrovirus and the multiple roles of the nucleocapsid protein

NC

NC

NC

NC

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Structural determinants for the specicity of NC for DNA The structure of the mutant His44Ala:NC(35-50):an NMR, MM and FL study

NMR and Fluorescence studies demonstrate

Mutant protein binds zinc. Mutant protein maintains some structure Binding to nucleic acids is less strong.

Biochemistry (2004) Stote RH et al, 43,7687-7697

Two-dimensional 1H NMR pH 6.5 at 274K

E. Kellenberger and B. Kieffer, ESBS

Answer the questions left unanswered by experiment How does mutant protein bind zinc ion? If folded, why is the activity diminished? Can simulations can predict the structural effects of point mutations?

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From NMR

From MD
angular S
1.2

rmsd ()
0.8 0.7 0.6

0.8

0.5 0.4 0.3 0.2

0.6

Ensemble of structures from MD

0.4

0.2

0.1 0 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50

residue number

Biochemistry (2004) Stote RH et al, 43,7687-7697

Structural Chemical Shifts :

Shifts sapay & Case, J. Am. Chem. Soc. 113

1991

Free

Complex

Structural Chemical Shift ()

Semi-empirical model for the calculation of

divided into different contributions Magnetic anisotropy Ring Current Electrostatics

() = ()complex - ()Random Coil

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Difference between calculated and experimental !" (ppm)

1 G35 C36 W37 K38 C39 G40 K41 E42 G43 A44 Q45 M46 K47 D48 C49 T50

0.5

-0.5

-1

-1.5

-2

Zinc binding by the mutant protein Reorientation of mainchain carbonyl oxygens stabilizes the ion zinc. In more unfolded protein, water molecules move in to form hydrogen bonds

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Study of the DNA/NC complex. Free energy decomposition.

LYS 47

MET 46

TRP 37

Decomposition of the binding free energy by amino acid for the native protein

Amino acids that contribute signicantly to DNA binding are those most affected by the mutation

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Conclusions
Since molecules are dynamic, experimental structures alone can not give the entire picture. An interdisciplinary approach is required. Molecular simulations are a necessary complement to the experimental studies.

Computer Simulation of Liquids

Edition New ed Allen, M. P., Tildesley, D. J.

Computational Chemistry

Grant, Guy H., Richards, W. Graham

Molecular Modelling: Principles and Applications (2nd Edition) (Paperback)

by Andrew Leach

http://www.ch.embnet.org/MD_tutorial/

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Acknowledgements
Herv Muller Elyette Martin

Prof. Bruno Kieffer (ESBS/IGBMC, Illkirch) Dr. Esther Kellenberger (ULP, Illkirch) Marc-Olivier Sercki (ESBS, Illkirch) Prof. Yves Mly (ULP, Illkirch) Dr. Elisa Bombarda (ULP, Illkirch) Prof. Bernard Roques (INSERM/CNRS, Paris)

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