A General Small-Angle X-ray Scattering-Based Screening Protocol Validated for Protein-RNA Interactions.

Chen PC, Masiewicz P, Rybin V, Svergun D, Hennig J, ACS Comb Sci 20(4):197-202 (2018) Europe PMC

SASDDZ4 – 1:1 Mixture between Protein sex-lethal mutant (Sxl10GS) and RNA decaneucleotide U8GU

Protein sex-lethal mutant
RNA decaneucleotide U8GU
MWexperimental 20 kDa
MWexpected 24 kDa
VPorod 34 nm3
log I(s) 4.19×101 4.19×100 4.19×10-1 4.19×10-2
Protein sex-lethal mutant RNA decaneucleotide U8GU small angle scattering data  s, nm-1
ln I(s)
Protein sex-lethal mutant RNA decaneucleotide U8GU Guinier plot ln 4.20×101 Rg: 2.2 nm 0 (2.2 nm)-2 s2
(sRg)2I(s)/I(0)
Protein sex-lethal mutant RNA decaneucleotide U8GU Kratky plot 1.104 0 3 sRg
p(r)
Protein sex-lethal mutant RNA decaneucleotide U8GU pair distance distribution function Rg: 2.3 nm 0 Dmax: 7.8 nm

Data validation


Fits and models


log I(s)
 s, nm-1
1:1 Mixture between Protein sex-lethal mutant (Sxl10GS) and RNA decaneucleotide U8GU Rg histogram Rg, nm
Protein sex-lethal mutant RNA decaneucleotide U8GU EOM/RANCH model
Protein sex-lethal mutant RNA decaneucleotide U8GU EOM/RANCH model
Protein sex-lethal mutant RNA decaneucleotide U8GU EOM/RANCH model
Protein sex-lethal mutant RNA decaneucleotide U8GU EOM/RANCH model
Protein sex-lethal mutant RNA decaneucleotide U8GU EOM/RANCH model
Protein sex-lethal mutant RNA decaneucleotide U8GU EOM/RANCH model

Synchrotron SAXS data from solutions of a 1:1 mixture between Protein sex-lethal mutant (Sxl10GS) and RNA decaneucleotide U8GU in 50% dilution of protein buffer (10 mM KH2PO4, 50 mM NaCl, 10 mM DTT), pH 6, were collected on the BM29 beam line at the ESRF storage ring (Grenoble, France) using a Pilatus 1M detector at a sample-detector distance of 2.9 m and at a wavelength of λ = 0.09919 nm (l(s) vs s, where s = 4πsinθ/λ, and 2θ is the scattering angle). One solute concentration of 1.00 mg/ml was measured at 20°C. 10 successive 1 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.

Note that the uploaded EOM ensemble is chosen as the 1:1-complex with U8GU positioned in the canonical RRM2 at 5' position from the Protein data bank (PDB) entry 1B7F.pdb. There is no observable discrimination between whether RRM1 or RRM2 is bound or its position along the RNA sequence.

Protein sex-lethal mutant (Sxl10GS)
Mol. type   Protein
Organism   Drosophila melanogaster
Olig. state   Monomer
Mon. MW   20.4 kDa
 
UniProt   P19339 (123-294)
Sequence   FASTA
 
RNA decaneucleotide U8GU (U8GU)
Mol. type   RNA
Organism   synthetic construct
Olig. state   Monomer
Mon. MW   3.1 kDa
Sequence   FASTA