Elastic Image Registration To Fully Explore Macromolecular Dynamics By Electron Microscopy

Qiyu Jin 1, Carlos Oscar Sanchez Sorzano 2, Isabelle Callebaut 1, Florence Tama 3, and Slavica Jonic 1

1 IMPMC, Sorbonne Universités - CNRS UMR 7590, UPMC Univ Paris 6, MNHN, IRD UMR 206, 75005 Paris, France. 2 Biocomputing Unit, Centro Nacional de Biotecnología – CSIC, 28049 Madrid, Spain.

3 RIKEN, Advanced Institute for Computational Sciences, Kobe, Hyogo, 650-0047, Japan.

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Normal mode analysis (NMA)

High frequenciesLow frequencies

Dynamics is described as a linear combination of independent harmonic oscillators

Mode 28 Mode 80 Mode 107

Tomato Bushy Stunt Virus (TBSV)

Deformation (displacement) amplitude :

N atoms, 3N coordinates : Nn 3,...,1

Initialstructure

Mlaaa Nllll ,...,1,,...,, 321 aModes :

Frequency : llq

3

0

022

21

L

Eigenvalues problem

Hessian: 2nd derivative of the potential

Eigenvector = normal mode Eigenvalue = frequency^2

21 aaA

Size of the system (3Nx3N)

LHAAt

ji

p

rr

E2H

Harmonic dynamics of the potential energy function around a minimum energy conformation

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, ,, 2a b a b a b

CE r r r r

E p E ra,rb ra,b

0 Rc

Elastic Network Model : Atoms connected via elastic springs

Computation of normal modes

Cowpea chlorotic mottle virus (CCMV)

Tama et al., 2002, JMB 318

TBSV

Flexible fitting: Flexible 3D-to-3D alignment of two different conformations

Study of conformational changes for unveiling biological functions:Use of normal modes to explore different conformations

A

B

C

5TBSV experiments: pseudo-atomic structure as reference

Flexible 2D-to-3D alignment

Hybrid Electron Microscopy Normal Mode Analysis (HEMNMA)Structure 22:496-506 , 2014

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• Normal modes are used within a flexible 2D-to-3D alignment of images with a reference EM or atomic structure

• Hybrid approach

• Possible movements of unknown amplitudes are determined by NMA

• Movements computed by NMA are verified experimentally by determining their

amplitudes through image analysis (flexible 2D-to-3D alignment)

• HEMNMA allows identifying intermediate conformations and continuous conformational changes of complexes in the sample

• Each particle image may represent a unique conformation

• Image classification and 3D reconstruction are not mandatory but can be done after image analysis

HEMNMA highlights

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TBSV experiments: atomic structure as reference

24%23%11% 19% 23%

RMSD=2.1 nm

JEOL JEM 2100F,ultra high-resolutionpole piece (Cs=0.5 mm)

4000 images analyzed with combined modes 28, 80 and 107

8Normal Mode Analysis and Flexible Alignment modules

HEMNMA integral graphical interface and protocol J Struct Biol, Sep 27, 2014

xmipp.cnb.csic.es

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Advanced Results Analysis module

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• HEMNMA: Combination of single-particle electron microscopy, normal mode analysis, and image analysis

• Elastic registration of images with a reference high- or low-resolution structure

• Full dynamics (distribution of conformations)• Modeling of transition pathways• Continuous conformational changes

• User-friendly integral graphical interface in Xmipp• Access to other Xmipp methods for single-particle analysis

• Study of TBSV conformational changes – work in progress• Analysis of larger number of images to better quantify the dynamics at

atomic level

Conclusions and Further work

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Thank you

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Iterative 3D-to-2D elastic registration