EMBO Practical Course on Solution Scattering from ...€¦ · EMBO Practical Course on Solution...
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EMBO Practical Course on Solution Scattering from Biological Macromolecules
D R id M d lD R id M d lDummy Residues ModelDummy Residues ModelProteins typically consist of folded polypeptide• Proteins typically consist of folded polypeptidechains composed of amino acid residues
• At a resolution of 0 5 nm each amino acid can beAt a resolution of 0.5 nm each amino acid can berepresented as one entity (dummy residue)
• In GASBOR a protein is represented by anbl f K DR thensemble of K DRs those are
– IdenticalHave no ordinal– Have no ordinalnumber
– For simplicity areFor simplicity arecentered at the Cαpositions
EMBO Practical Course on Solution Scattering from Biological Macromolecules
DR M d lli GASBORDR M d lli GASBORDR Modelling: GASBORDR Modelling: GASBOR• Finds coordinates {ri}
of K DRs within the sphericalh lsearch volume
• Scattering is computed usingth D b (1915) f lthe Debye (1915) formula
( ) ( ) ( )∑∑=K K
ijjiDR
srsgsgsI
sin
• Requires polypeptide chain-
( ) ( ) ( )∑∑= =i j ij
jiDR sr1 1
= < … … >q p yp p
compatible arrangement ofDRs
D.I. Svergun, M.V. Petoukhov, & M.H.J. Koch (2001) Biophys. J. 80, 2946-53
EMBO Practical Course on Solution Scattering from Biological Macromolecules
GASBOR R t i tGASBOR R t i tGASBOR RestraintsGASBOR RestraintsE l d d l ff t d l l i t ti l d tExcluded volume effects and local interactions lead to acharacteristic distribution of nearest neighbors around agiven residue in a polypeptide chain
Number of neighbours6
4
5
2
3
0 2 0 4 0 6 0 8 1 00
1
Shell radius, nm
0.2 0.4 0.6 0.8 1.0
EMBO Practical Course on Solution Scattering from Biological Macromolecules
DR modelling program GASBORDR modelling program GASBORDR modelling program GASBORDR modelling program GASBOR
searches for a chain-like arrangement of dummysearches for a chain like arrangement of dummy residues fitting the scattering data by minimizing
f (X) = χ2 + Σα P (X)
I, rel. units 2.0 p(r), rel. units
f (X) = χ + ΣαiPi(X)
100
1000
experiment calc. from p(r)
1.0
1.5
L t ti l ti
0 0 0 5 1 0 1 5 2 01
10
0 5 10 150.0
0.5 DmaxLong computational timefor large proteins0.0 0.5 1.0 1.5 2.0s, nm-1
• Symmetry can be taken into account; groups supported:
0 5 10 15 distance r, nm
y y g p ppPn, Pn2 (n=2..19), P23, P432 and icosahedral symmetry
EMBO Practical Course on Solution Scattering from Biological Macromolecules
GASBOR SGASBOR SGASBOR SummaryGASBOR Summary• Task: Searches for a chain like arrangement of dummy• Task: Searches for a chain-like arrangement of dummy
residues fitting the scattering data.• Parameters: 3D coordinates of DRs describing CαParameters: 3D coordinates of DRs describing Cα
positions.• Objects: Applicable for polypeptide chains (i.e. proteins
and their assemblies) with K ≲ 5000.• Capabilities: Fits the scattering curves at higher angles.
Takes into account symmetry and anisometry ReciprocalTakes into account symmetry and anisometry. Reciprocaland real space fitting options.
• Limitations: Fits only SAXS data and applicable forLimitations: Fits only SAXS data and applicable forproteins only. CPU time is quadratically proportional to K.
• Theoretical intensity: computed using the Debyeformula.
GASBORGASBOR ll
EMBO Practical Course on Solution Scattering from Biological Macromolecules
GASBOR GASBOR examplesexamples
• Neocarzinostatin, 113 AA• Catalase, tetramaer, P222, 501AA per , , , p
monomer
• supcomb 1NOA.PDB test01.pdb -e --superposition=backbone
EMBO Practical Course on Solution Scattering from Biological Macromolecules
n: 0 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15Fib(n): 1 1 2 3 5 8 13 21 34 55 89 144 233 377 610 987
Δρ=0.03 e/A3
Width=3A
ρ=0.334 e/A3
EMBO Practical Course on Solution Scattering from Biological Macromolecules
MONSA (multiphase modelling)MONSA (multiphase modelling)MONSA (multiphase modelling)MONSA (multiphase modelling)
DAMMIN/F shape determination
• One can differentiate between distinct partsbetween distinct parts of the particle
• Several curves are requiredrequired
• Assuming the same arrangement of the
t i diff tparts in different samples
EMBO Practical Course on Solution Scattering from Biological Macromolecules
MONSA (multiphase modelling)MONSA (multiphase modelling)MONSA (multiphase modelling)MONSA (multiphase modelling)
1 h 1 t f l ti l• 1 phase = 1 component of a complex particle• For each phase, Rg, V and its scattering curve can be given• For each curve, contrast of each phase are specified , p p
contrast variation and / or use ofand / or use of partial constructs
C 1C 1 t it i RNARNA ll
EMBO Practical Course on Solution Scattering from Biological Macromolecules
Case1: Case1: proteinprotein--RNA RNA complexcomplex
252 AA protein (two domains)
67 nucleotides RNA
Three curves in total:
Free RNA
Complex
Free protein