Dynamic RNA stem-loops Drive dimerization-assisted higher-order assemblies of SARS-CoV-2 Nsp9

Jill Trewhella.

SASDYQ6 – SARS-CoV-2 Non-structural protein 9 (Nsp9) at 6 mg/mL

SARS-CoV-2 non-structural protein 9 (Nsp9)
MWexperimental 26 kDa
MWexpected 25 kDa
VPorod 36 nm3
log I(s) 2.47×10-2 2.47×10-3 2.47×10-4 2.47×10-5
SARS-CoV-2 non-structural protein 9 (Nsp9) small angle scattering data  s, nm-1
ln I(s)
SARS-CoV-2 non-structural protein 9 (Nsp9) Guinier plot ln 2.48×10-2 Rg: 2.1 nm 0 (2.1 nm)-2 s2
(sRg)2I(s)/I(0)
SARS-CoV-2 non-structural protein 9 (Nsp9) Kratky plot 1.104 0 √3 sRg
p(r)
SARS-CoV-2 non-structural protein 9 (Nsp9) pair distance distribution function Rg: 2.1 nm 0 Dmax: 8.1 nm

Data validation


Fits and models


log I(s)
 s, nm-1
SARS-CoV-2 non-structural protein 9 (Nsp9) DAMMIN model
SARS-CoV-2 non-structural protein 9 (Nsp9) DAMMIN model

SAXS data for non-structural protein 9 (Nsp9) from SARS-CoV-2, in 25 mM sodium phosphate pH 6.0, 150 mM NaCl, 1 mM DTT with 5% glycerol as a stabilizer, were collected at the BioSAXS beamline of the Australian Synchrotron. The X-ray wavelength was 1.002 Å; the detector was a Dectris Pilatus3X-2M with in-vacuum movable detector system, 172 µm2 pixel size, 25 Hz frame rate, count rate per detector pixel was 39.95 counts/s; the sample-to-detector distance was 3150 mm to yield a measured q-range 0.0051–0.4768 Å-1; exposures were 1 second for 16 exposures and absolute scaling used water as a secondary standard. Samples were spun at 21,000 x g for 5 min and degassed in a vacuum chamber for 5 min just prior to loading onto a 96-well plate for measurement. Concentration values are for the nominal Nsp9 component prior to that procedure according to absorbance at 280 nm. Shown here are the 6 mg/mL data, where the structural parameters, including the Porod volume, and DAMMIN envelops indicate the protein is fully dimeric. The very small deviations in the low-q residual difference plots are consistent with a very small amount of aggregate present in the solution. A full concentration series (0.05–8 mg/mL) was performed for Nsp9 and data for all concentrations are available in zipped folders with this deposition.

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SARS-CoV-2 non-structural protein 9 (Nsp9) (Nsp9)
Mol. type   Protein
Organism   Severe acute respiratory syndrome coronavirus 2
Olig. state   Dimer
Mon. MW   12.4 kDa
 
UniProt   P0DTD1 (4141-4253)
Sequence   FASTA