PhD student: Søren Enemark

Supervisors: Prof. A. Redaelli Ing. S. Monica

Tubulin Monomer Mechanical Properties Obtained by Simulating Atomic Force Microscopy Experiments Using Molecular Dynamics

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Rot. + trans. symmetry

Introduction: What am I talking about?

αβ-tubulin dimer

Hetero dimer

α-Tubulin β-Tubulin

2 x ~450 residues

Microtubules (MTs)

Length ~ 1-10μm

Cylinder-shaped

Subunit in MTs

30 nm

18 nm

Lattice structure

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slideIntroduction: What are microtubules good for?

MTs’ main functions:

• Structural elements

• Intracellular transport

• Cell division

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slideModeling: Stress-strain directions

Lateral

Longitudinal

Compression Elongation

4 tests for each monomer

Single monomer mechanical properties

MT mechanical properties

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Atomic structure by K. Downing

Basic structure 1JFF.pdb

Fitted to MT structure data

Modeling: Stress-strain directions (cont’d)

Lateral interactions

End view on MTMonomer centre-of-mass (CM)

13.8°

Longitudinal interactions

Along straight line

Directions depend on the MT structure:

Along lines towards CM of lateral monomers

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slideProcedure: Preparing the structure

Monomer structure extracted

MD Pull groups position restraints

1.

EM Steepest descent (1000 steps)3.

4.

Arrange in box w/ SPC water2.

Dodecahedron box

Twin-range cut-off

All-bonds constraints, Lincs

Berendsen thermostat Two groups: SOL monomer

tau= 0.1 psTref = 300 K

rvdw = 1.4 rcoulomb=1.4Parametersrlist= 0.8 nstlist = 5

System size ~ 37,000 (SOL)

+ 4,500 (monomer)

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slidePreparing AFM-like MD

• residues < 8 Ǻ from interacting monomer

• In longitudinally tests: 731 and 675 atoms

• In laterally tests: 188 and 198 atoms

Pull groups:

Retain interface structure, butgenerate new configurations

0 ps 1000 ps

Position restraint pull groups

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slideAFM-like MD – how to measure the stiffness

Pull group

P1

Pull group

P2

Spring S2Spring S1

v=5 10-3 nm/ps

0

0.1

0.2

0.3

-0.1

x (nm)

t (ps)

-0.2

-0.3

100 600500400300200

S

S’

P’

P

AFM-like Method Typical results (pulling)

S1

S2

P2

P1

Spring stiffness

Spring velocity

103 kJ/(nm2 mol) = 1.67 nN/nm

(7-11 simulations)

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slideSingle monomer - MD results

F (nN)

l (nm)

v=5 10-3 nm/ps

Linear fit y(x) = a x + b

a= 4.6 nN/nm b= 0.2 nN

-tubulin – pulling - longitudinally

F (nN)

l (nm)

a= 5.2±0.4 nN/nm b= 0.4±0.1 nNv=5 10-3 nm/ps

1.25

1.00

0.75

0.1 0.2 0.3-0.1 0

0.1 0.2 0.3-0.1 0

1.25

1.00

0.75

0.50

0.25

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slideSingle monomer - MD results

α-tubulin

β-tubulin

Longitudinally Laterally

Elongation Compression Elongation Compression

b -1.1± 0.5

a 5.8± 0.6

b 0.4± 0.1

a 5.2± 0.4

b 0.9± 0.1

a 6.6± 0.3

v=5 10-3 nm/ps

b 0.0± 0.1

a 6.4± 0.4

b -0.5± 0.3

a 5.4± 1.3

b 1.0± 0.3

a 3.8± 0.9

b 1.6± 0.2

a 3.2± 1.0

b -0.3± 0.1

a 3.7± 0.5

Monomer might be less rigid under elongation than compression

α-tubulin might be less rigid then β-tubulin longitudinally, but more rigid laterally

Monomers are more rigid longitudinally than laterally

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k

k

2k

CM

CM

k

k

CM

CM

k~

k~

k~

Simplified MT model – a bed of springs

Elastic constants

EST Marie Curie programme contract No. MEST-CT-2004-504465

Active BIOMICS STREP project contract No. NMP4-CT-2004-516989Funded

“Tubulin Monomer Interaction Study by Molecular Dynamics Simulation” Marco Deriu et al.

α-, and β-tubulin stiffness &

monomer-to-monomer interactions

Axial tests on 1 μm MT

MD

FEM

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