SAXS reveals the molecular basis underlying pH-driven G3BP1 conformational dynamics: implications for stress granule formation

SASDWK8 – G3BP1ΔRGG (residues 1-413) at pH 7.5, 150 mM NaCl

RGG deletion mutant of Ras GTPase-activating protein-binding protein 1
MWexperimental 109 kDa
MWexpected 94 kDa
VPorod 226 nm3
log I(s) 2.01×10-1 2.01×10-2 2.01×10-3 2.01×10-4
RGG deletion mutant of Ras GTPase-activating protein-binding protein 1 small angle scattering data  s, nm-1
ln I(s)
RGG deletion mutant of Ras GTPase-activating protein-binding protein 1 Guinier plot ln 2.01×10-1 Rg: 6.6 nm 0 (6.6 nm)-2 s2
(sRg)2I(s)/I(0)
RGG deletion mutant of Ras GTPase-activating protein-binding protein 1 Kratky plot 1.104 0 √3 sRg
p(r)
RGG deletion mutant of Ras GTPase-activating protein-binding protein 1 pair distance distribution function Rg: 7.4 nm 0 Dmax: 32 nm

Data validation

Fits and models

log I(s)
 s, nm-1
RGG deletion mutant of Ras GTPase-activating protein-binding protein 1 DAMMIN model
log I(s)
 s, nm-1
RGG deletion mutant of Ras GTPase-activating protein-binding protein 1 CORAL model
Synchrotron SAXS data from solutions of G3BP1ΔRGG (residues 1-413) at pH 7.5, 150 mM NaCl in 20 mM Tris, 150 mM NaCl,, pH 7.5 were collected on the B21 beam line at the Diamond Light Source storage ring (Didcot, UK) using a Eiger 4M detector at a sample-detector distance of 3.7 m and at a wavelength of λ = 0.09464 nm (I(s) vs s, where s = 4πsinθ/λ, and 2θ is the scattering angle). In-line size-exclusion chromatography (SEC) SAS was employed. The SEC parameters were as follows: A 60.00 μl sample at 10 mg/ml was injected at a 0.08 ml/min flow rate onto a Cytiva Superdex 200 Increase 3.2/300 column at 15°C. 600 successive 3 second frames were collected. The data were normalized to the intensity of the transmitted beam and radially averaged; the scattering of the solvent-blank was subtracted.

RGG deletion mutant of Ras GTPase-activating protein-binding protein 1 (G3BP1(ΔRGG))
Mol. type   Protein
Organism   Homo sapiens
Olig. state   Dimer
Mon. MW   47.0 kDa
 
UniProt   Q13283 (1-413)
Sequence   FASTA