SREFLEX: S AXS REF inement through FLEX ibility Alejandro Panjkovich - - PowerPoint PPT Presentation

▶
sreflex s axs ref inement through flex ibility
SMART_READER_LITE
LIVE PREVIEW

SREFLEX: S AXS REF inement through FLEX ibility Alejandro Panjkovich - - PowerPoint PPT Presentation

Nov 02, 2022 12 likes •285 views

SREFLEX: S AXS REF inement through FLEX ibility Alejandro Panjkovich EMBL Hamburg 20.10.2016 A. Panjkovich (EMBL) SAXS + NMA 20.10.2016 1 / 27 Outline Motivation for flexible refinement SREFLEX method Example from user Running the

slide-1
SLIDE 1

SREFLEX: SAXS REFinement through FLEXibility

Alejandro Panjkovich EMBL Hamburg 20.10.2016

  • A. Panjkovich (EMBL)

SAXS + NMA 20.10.2016 1 / 27

slide-2
SLIDE 2

Outline

Motivation for flexible refinement SREFLEX method Example from user Running the program

◮ Standard run ◮ Advanced domain definition

Interpreting results

  • A. Panjkovich (EMBL)

SAXS + NMA 20.10.2016 2 / 27

slide-3
SLIDE 3

SAXS and conformational change

Crystalline and solution conformation may differ SAXS can provide insight into conformational transition

  • A. Panjkovich (EMBL)

SAXS + NMA 20.10.2016 3 / 27

slide-4
SLIDE 4

SAS modeling principle

  • A. Panjkovich (EMBL)

SAXS + NMA 20.10.2016 4 / 27

slide-5
SLIDE 5

Estimating protein flexibility: normal mode analysis (NMA)

Delarue M, Sanejouand YH (2002) Simplified normal mode analysis of conformational transitions in dna-dependent polymerases: the elastic network model. J Mol Biol 320: 1011-1024.

  • A. Panjkovich (EMBL)

SAXS + NMA 20.10.2016 5 / 27

slide-6
SLIDE 6

Time for some movement

  • A. Panjkovich (EMBL)

SAXS + NMA 20.10.2016 6 / 27

slide-7
SLIDE 7

SREFLEX: SAXS REFinement through FLEXibility

  • A. Panjkovich (EMBL)

SAXS + NMA 20.10.2016 7 / 27

slide-8
SLIDE 8

SREFLEX: SAXS REFinement through FLEXibility

Input: SAXS data PDB coordinates Program stages:

1 Structure partition 2 Domain level refinement 3 Residue level refinement

  • A. Panjkovich (EMBL)

SAXS + NMA 20.10.2016 8 / 27

slide-9
SLIDE 9

Automatic domain assignment based on dynamics

  • A. Panjkovich (EMBL)

SAXS + NMA 20.10.2016 9 / 27

slide-10
SLIDE 10

Assignment comparison, SCOP vs. auto

  • A. Panjkovich (EMBL)

SAXS + NMA 20.10.2016 10 / 27

slide-11
SLIDE 11

Experimental example: Josephin domain of ataxin-3

Nicastro et al. (2006) J. Biomol. NMR 36, 267-77

  • A. Panjkovich (EMBL)

SAXS + NMA 20.10.2016 11 / 27

slide-12
SLIDE 12

Experimental example: MurA

Sch¨

  • nbrunn et al. (1998) Eur. J. Biochem. 253, 406-412
  • A. Panjkovich (EMBL)

SAXS + NMA 20.10.2016 12 / 27

slide-13
SLIDE 13

SREFLEX: examples from benchmark

  • A. Panjkovich (EMBL)

SAXS + NMA 20.10.2016 13 / 27

slide-14
SLIDE 14

SREFLEX benchmark

10 20 30 40 50 60 70 scop auto ffs % benchmark cases RMSDtarget < 5.0 Å closing

  • pening

total 20 40 60 80 100 scop auto ffs % benchmark cases χ2 < 2.0 closing

  • pening

total 20 40 60 80 100 scop auto fs % benchmark cases ∆ RMSDtarget > 1.0 Å closing

  • pening

total

  • A. Panjkovich (EMBL)

SAXS + NMA 20.10.2016 14 / 27

slide-15
SLIDE 15

Panjkovich A & Svergun DI (2016) Phys. Chem. Chem. Phys. 18, 5707-19

  • A. Panjkovich (EMBL)

SAXS + NMA 20.10.2016 15 / 27

slide-16
SLIDE 16

SREFLEX, user example

  • A. Panjkovich (EMBL)

SAXS + NMA 20.10.2016 16 / 27

slide-17
SLIDE 17

SREFLEX, ab-initio model mismatch

  • A. Panjkovich (EMBL)

SAXS + NMA 20.10.2016 17 / 27

slide-18
SLIDE 18

SREFLEX, conformational change

  • A. Panjkovich (EMBL)

SAXS + NMA 20.10.2016 18 / 27

slide-19
SLIDE 19

Conformational change fits ligand binding

  • A. Panjkovich (EMBL)

SAXS + NMA 20.10.2016 19 / 27

slide-20
SLIDE 20

SREFLEX, web interface (ATSAS online)

  • A. Panjkovich (EMBL)

SAXS + NMA 20.10.2016 20 / 27

slide-21
SLIDE 21

SREFLEX, command line interface

  • A. Panjkovich (EMBL)

SAXS + NMA 20.10.2016 21 / 27

slide-22
SLIDE 22

Divide et impera - split your structure into pieces

The new ATSAS binary partistr may help Check Pfam or SCOP domains Use your intuition (particularly for opening structures) Feed the structure as

◮ d1.pdb,d2.pdb,d3.pdb (command line) ◮ Or as a .zip file to the ATSAS-online version

  • A. Panjkovich (EMBL)

SAXS + NMA 20.10.2016 22 / 27

slide-23
SLIDE 23

Interpretation of SREFLEX results

Restrained models (rc), higher χ2 but better stereochemistry Unrestrained models (uc), lower χ2, more structural distortion Prioritize lower RMSD against initial structure Check degree and relevance of clashes and breaks Make an informed decision on which model is the better choice

  • A. Panjkovich (EMBL)

SAXS + NMA 20.10.2016 23 / 27

slide-24
SLIDE 24

Use the proper tool for your question

MW or oligomeric state are way off (better SASREF) There are long linkers or unstructured regions (better use EOM) Unrealistic expectations (folding problem not yet solved)

  • A. Panjkovich (EMBL)

SAXS + NMA 20.10.2016 24 / 27

slide-25
SLIDE 25

Summary

Motivation Understanding the approach Running the program

◮ Standard run ◮ Custom domain definition

Interpreting results

  • A. Panjkovich (EMBL)

SAXS + NMA 20.10.2016 25 / 27

slide-26
SLIDE 26

Acknowledgements

Dmitri Svergun and the rest of the SAXS team at EMBL Hamburg (https://www.embl-hamburg.de/biosaxs/)

Funding:

◮ EMBL and Marie Curie Actions for postdoc fellowship (EIPOD)

  • A. Panjkovich (EMBL)

SAXS + NMA 20.10.2016 26 / 27

slide-27
SLIDE 27

SAS in structural biology

ab-initio shape determination missing fragments rigid-body hybrid modeling mixtures ensemble approach

  • A. Panjkovich (EMBL)

SAXS + NMA 20.10.2016 27 / 27