nsp8_kurauskas
Description
NMR spectra of SARS-CoV-2 full-length nsp8 protein: in complex with different ratios of interaction partner nsp7, as well as free nsp8 spectra at different protein and salt concentrations. Spectra include 4D methyl NOESY spectra used for the assignment of nsp8 (in nsp7-nsp8 complex). Spectra show large contrast between nsp7 bound nsp8 spectra and free nsp8 spectra. While nsp7 bound nsp8 spectra displays sharp peaks and a good correspondence with the X-ray structures, free nsp8 spectra is broad suggesting dynamic nature of nsp8. We have studied what biophysical/biological process leads to spectral broadening of nsp8, and based on this and additional data came to conclusion that nsp8 is a molten globule in its free state.
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Sample conditions in these experiments were 10mM MOPS or Na-phosphate buffer pH 7, 2-3mM TCEP as reducing agent, 10% D2O (4D NOESY spectrum recorded in 100% D2O) and at least 150mM NaCl (at 50mM NaCl severe signal loss is observed over the period of several days). Spectra were processed conventionally. The 3D and 4D spectra were recorded with non-uniform sampling, and were reconstructed using SMILE.
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Funding
National Institutes of Health
R35GM141748
National Institutes of Health
R01AI158463
National Institutes of Health
R24GM141526
U.S. National Science Foundation
MCB-2031269