Structural plasticity of the coiled-coil interactions in human SFPQ.

Koning HJ, Lai JY, Marshall AC, Stroeher E, Monahan G, Pullakhandam A, Knott GJ, Ryan TM, Fox AH, Whitten A, Lee M, Bond CS, Nucleic Acids Res (2024) Europe PMC

SASDMW8 – The disulphide-linked tetramer of splicing factor, proline- and glutamine-rich and Non-POU domain-containing octamer-binding protein (SFPQ276-598(R542C)/NONO53-312) at 0.78mg/ml

Non-POU domain-containing octamer-binding protein
Splicing factor, proline- and glutamine-rich SFPQ276-598(R542C)/NONO53-312 dimer)
MWexperimental 186 kDa
MWexpected 137 kDa
VPorod 257 nm3
log I(s) 8.02×10-2 8.02×10-3 8.02×10-4 8.02×10-5
Non-POU domain-containing octamer-binding protein Splicing factor, proline- and glutamine-rich SFPQ276-598(R542C)/NONO53-312 dimer) small angle scattering data  s, nm-1
ln I(s)
Non-POU domain-containing octamer-binding protein Splicing factor, proline- and glutamine-rich SFPQ276-598(R542C)/NONO53-312 dimer) Guinier plot ln 8.03×10-2 Rg: 5.2 nm 0 (5.2 nm)-2 s2
(sRg)2I(s)/I(0)
Non-POU domain-containing octamer-binding protein Splicing factor, proline- and glutamine-rich SFPQ276-598(R542C)/NONO53-312 dimer) Kratky plot 1.104 0 √3 sRg
p(r)
Non-POU domain-containing octamer-binding protein Splicing factor, proline- and glutamine-rich SFPQ276-598(R542C)/NONO53-312 dimer) pair distance distribution function Rg: 5.4 nm 0 Dmax: 21 nm

Data validation

Fits and models

log I(s)
 s, nm-1
Non-POU domain-containing octamer-binding protein Splicing factor, proline- and glutamine-rich SFPQ276-598(R542C)/NONO53-312 dimer) DAMMIF model
Synchrotron SAXS data from solutions of SFPQ276-598(R542C)/NONO53-312 at 0.78mg/ml in 20 mM Tris pH 7.5, 250 mM NaCl, were collected on the SAXS/WAXS beam line at the Australian Synchrotron (Melbourne, Australia) using a Pilatus3 S 2M detector at a wavelength of λ = 0.10781 nm (I(s) vs s, where s = 4πsinθ/λ, and 2θ is the scattering angle). One solute concentration of 0.78 mg/ml was measured at 25°C. The data were normalized to the intensity of the transmitted beam and radially averaged; the scattering of the solvent-blank was subtracted.

A tetramer of SFPQ276-598(R542C)/NONO53-312 at 0.78mg/ml which forms when dimers of SFPQ276-598(R542C)/NONO53-312 bind to each other and become cross-linked by the formation of disulfide bond between the coiled-coil domains of SFPQ. The P(r) function and other relevant pieces of data are in agreement with those of a similar construct (SFPQ214-598(R542C)/NONO53-312) resolved by SEC-SAXS with a linear Guinier region (deposited). Additionally, the DAMMIF model of our data overlays reasonably well against a crystal structure of the protein formatted as a tetramer (6WMZ). The rough shape of the molecule appears unchanged as a function of the measured concentration range. We interpret this as the disulfide-linked tetramer persisting over a range of concentrations. Additional data are made available in the full entry zip archive with scattering for the protein at 1.6mg/ml and the corresponding p(r) distribution.

Non-POU domain-containing octamer-binding protein (NONO dimer)
Mol. type   Protein
Organism   Homo sapiens
Olig. state   Dimer
Mon. MW   30.1 kDa
 
UniProt   Q15233 (53-312)
Sequence   FASTA
 
Splicing factor, proline- and glutamine-rich SFPQ276-598(R542C)/NONO53-312 dimer) (SFPQ cysteine-linked)
Mol. type   Protein
Organism   Homo sapiens
Olig. state   Dimer
Mon. MW   38.2 kDa
 
UniProt   P23246 (276-598)
Sequence   FASTA