Resolving the Structure of Viral Genomes with Atomic Resolution - PowerPoint PPT Presentation

Resolving the Structure of Viral Genomes with Atomic Resolution Aleksei Aksimentiev Department of Physics University of Illinois at Urbana-Champaign

I use Blue Waters to … … understand molecular underpinnings of life … build biologically inspired systems

DNA, the blueprint

Viral genome, the program of infection DNA is a highly charged polymer! Herpes virus (HSV) Cryoem reconstruction with concentric rings (Evilevitch et al, UIUC) Open questions: - What is the 3D structure of the genome? - How genome ejection is triggered and sustained? - Can it be used as a drug target? http://darwin.bio.uci.edu/~faculty/wagner/hsv2f.html

Same sign charges …. F F F F - - - - Same sign charges can Same sign charges repel a0ract (in a medium) (in vacuum) DNA is surrounded +1e, sodium or potassium by counter ions +2e, magnesium or calcium +3e,spermidine +4e,spermine EffecBve a0racBon between DNA is observed when counterions have charge ≥ 2e

All-Atom Molecular Dynamics Simulation of DNA Condensates Add 64 DNA helices Add polyamine cations (+4) Add 150 mM NaCl Add explicit water Apply a half- harmonic wall potential only to DNA DNA-confining wall of radius R Solve the equation of motion (F= ma) under periodic boundary condition in all directions Classical Force Field } Partial charges θ q i q j from quantum X b + 4 ⇡✏ 0 r ij mechanics k b ( b − b 0 ) 2 + non-bonded pairs i,j X X k θ ( θ − θ 0 ) 2 U ( r ) = ◆ 6 ## LJ parameters } "✓ � ij ◆ 12 bonds angles ✓ � ij φ X from + 4 ✏ ij ) 2 + X − k φ (1 + cos ( n φ − φ 0 )) r ij r ij experiments non-bonded pairs i,j dihedrals Bonded parameters from quantum mechanics #

Standard CHARMM & AMBER Force Fields Are Not Perfect for the Simulation of DNA Condensates Na 15-ns MD Cross-sectional view of MD using CHARMM27 Harmonic wall y (nm) Long-lasting contact ion pairs (CIP) between [Na] = 250 mM outside Na + and phosphate stabilize contact DNA pairs. x (nm) [Na] = 250 mM [Mg] = 20 mM [spermine] = 2 mM 100 100 100 6 6 Todd et al. 4 Due to excessive CIP 4 4 Rau et al. Pressure (bar) Pressure (bar) Pressure (bar) 2 Rau et al. AMBER99 2 2 10 formation, the simulations 10 10 4 AMBER99 underestimate both inter-DNA 6 6 2 CHARMM27 4 4 distance and pressure in DNA 1 AMBER99 2 2 array systems. CHARMM27 4 C H A R M M 2 7 1 1 2 22 24 26 28 30 32 22 24 26 28 30 25 30 DNA-DNA distance (Å) DNA-DNA distance (Å) DNA-DNA distance (Å) Rau et al, PNAS 1984 Todd et al, BJ 2008 Yoo & Aksimentiev, JPCL 2012

Champaign-Urbana Non-Bonded FIX (CUFIX): Improved Lennard-Jones Parameters for CHARMM & AMBER • “Much of what is known about association and Standard r min = 3.11 Å Exp. from Robinson 1959 dissociation of solutes and ions comes from 0.20 0.20 200 200 measurements of colligative properties ” — OSM pressure (bar) 0.15 0.15 150 150 NBFIX U LJ (kcal/mol) Molecular driving forces by Dill & Bromberg. r min = 3.20 Å 0.10 0.10 0.05 0.05 100 100 Dimethylphosphate Acetate 0.00 0.00 50 50 Standard -0.05 -0.05 ≈ Na -0.10 -0.10 0 0 2.8 2.8 3.2 3.2 3.6 3.6 4.0 4.0 0 0 1 1 2 2 3 3 4 4 Na–O distance (Å) molal conc (m) http://bionano.physics.illinois.edu/CUFIX Effectively infinite slab under PBC CUFIX for CHARMM36 & AMBER99 -4 -2 0 2 4 1.2 Density (g/cm 1.0 0.8 Na Acetate 0.6 Water Total 0.4 0.2 0.0 3 ) Yoo & Aksimentiev, JPCL 2012 Murad & Powles, JCP 1993 Yoo & Aksimentiev, JCTC 2016 Yoo & Aksimentiev, JPCL 2012 Yoo, Wilson & Aksimentiev, Biopolymers 2016 Luo & Roux, JPCL 2010

CUFIX Improves Simulations of DNA Condensates [Na] = 250 mM [Mg] = 20 mM [spermine] = 2 mM 100 100 100 AMBER99 + CUFIX 6 6 4 A M B E R 9 9 + C U F I X A M B E R 9 9 + C U F I X 4 4 Pressure (bar) Pressure (bar) ( M D i n c l u d e d 2 0 0 m M N a ) 2 Pressure (bar) 2 10 2 10 CHARMM27 10 4 6 2 6 CHARMM27 4 1 4 AMBER99 A M B E R 9 9 AMBER99 2 2 4 CHARMM27 1 2 1 22 24 26 28 30 32 22 24 26 28 30 25 30 DNA-DNA distance (Å) DNA-DNA distance (Å) DNA-DNA distance (Å) 120 120 120 80 80 80 40 40 40 y (Å) y (Å) y (Å) 0 0 0 -40 -40 -40 -80 -80 -80 -120 -120 -120 -120 -80 -40 0 40 80 120 -120 -80 -40 0 40 80 120 -120 -80 -40 0 40 80 120 x (Å) x (Å) x (Å) Yoo & Aksimentiev, NAR 2016

DNA is packaged by a motor Takes about 3 minutes to pack DNA 130 times longer than the capsid ! Max Force: 100pN Movie: Carlos Bustamante Lab Can one simulate the process? Packaging process is slow (~min), At higher forces, DNA will deform all-atom simulation at physiological forces is not possible

Strategy: change resolution for speed and detail

Strategy: change resolution for speed and detail

Strategy: change resolution for speed and detail

Strategy: change resolution for speed and detail

Strategy: change resolution for speed and detail

Strategy: change resolution for speed and detail

500 bp dsDNA fragment modeled at different resolutions 24 bp/2 beads 12 bp/2 beads 6 bp/2 beads 3 bp/2 beads 1 bp/2 beads All-atom, ~100 bp

Interactions in a simple coarse-grained DNA model

Interactions in a simple coarse-grained DNA model Bond potential Force r 0 r 0 = n bp × 3 . 4 ˚ A Elastic constant f 0 = 1000pN k spring = f 0 /r 0 Extension http://www.phys.ens.fr/~cocco/Art/24physworld.pdf

Interactions in a simple coarse-grained DNA model 0 sin θ d θ cos θ e − β 1 2 k spring θ 2 ∫ 𝕐 d 𝕐 cos θ δ ( θ ′ � [ 𝕐 ] − θ ) e − β U [ 𝕐 ] π ∫ e − s / L p = ⟨ cos θ ⟩ = = ∫ 𝕐 d 𝕐 e − β U [ 𝕐 ] 0 sin θ d θ e − β 1 π 2 k spring θ 2 ∫ s Angle potential L p = 50 nm Persistence length

Interactions in a simple coarse-grained DNA model Dihedral angle potential Twist persistence length 90° L tw = 90 nm h cos φ i = e � s/L tw Z π k dihed( φ − φ 0)2 = e � s/L tw d φ cos φ e � 2 kBT φ 0 φ 0 = s ⇥ 10 . 14 � / ˚ A

Interactions in a simple coarse-grained DNA model z ���� Periodic in axial axis P r 4 nm 
 cuto ff �����������������������������

Interactions in a simple coarse-grained DNA model 4 bp/ 80 80 4 nm 
 cuto ff Optimized to reproduce Half-harmonic 
 Rau & Parsegian pressure wall to prevent 
 strand crossing

Mapping between coarse-grained resolutions For each helix, Fit a spline between 
 fit a 3D spline through bead coordinates quaternion representation of at end of simulation rotations coordinate 1D spline

Packaging viruses with ARBD ARBD: Atomic Resolution Brownian Dynamics (multi-resolution) Package DNA (CG) with ARBD, into CryoEM reconstruction of a HK97 bacteriophage capsid. A cryoEM map of the portal is fitted into the original capsid reconstruction, and DNA is packaged through the portal. Smooth, purely repulsive grid-based potential obtained by blurring cryoEM density and adding the portal

MulB-resoluBon packaging dsDNA viruses

Internal pressure during packaging outside capsid, % Percent of DNA Evilevitch et al, PNAS Pressure (atm) �27

Comparison to structural data Cryo-electron microscopy Small Angle X-ray Scattering experiment I ( q ) [a . u.] simulation Simulation Experiment q [Å − 1 ] Experiment: J. Mol. Biol. (2009) 391, 471-483, Hendrix et al Journal of molecular biology , 408: 541 (2011) Simulation SAXS data were generated from CRYSOL, using an atomistic PDB of the protein coat and packaged DNA �28

Conclusions and outlook Obtained first atomic-resolution structure of packaged viral particle Developed accurate multi-resolution representation of DNA—DNA and DNA—protein interactions To do: Extend the model to ssRNA and ssDNA viruses �29

Acknowledgements • Funding through CPLC • Computations David Jejoong Yoo Chris Maffeo Kush Coshic Winogradoff