Assignment of Ala, Ile, LeuproS, Met, and ValproS methyl groups of the protruding domain of murine norovirus capsid protein VP1 using methyl–methyl NOEs, site directed mutagenesis, and pseudocontact shifts

Biomolecular NMR Assignments - Tập 16 - Trang 97-107 - 2022
Thorben Maass1, Leon Torben Westermann1, Robert Creutznacher1, Alvaro Mallagaray1, Jasmin Dülfer2, Charlotte Uetrecht2,3, Thomas Peters1
1Center of Structural and Cell Biology in Medicine (CSCM), Institute of Chemistry and Metabolomics, University of Luebeck, Luebeck, Germany
2Leibniz Institute for Experimental Virology (HPI), Hamburg, Germany
3School of Life Sciences, University of Siegen, 57076 Siegen & Centre for Structural Systems Biology (CSSB), & Deutsches Elektronensynchrotron (DESY), Schenefeld, Germany

Tóm tắt

The protruding domain (P-domain) of the murine norovirus (MNV) capsid protein VP1 is essential for infection. It mediates receptor binding and attachment of neutralizing antibodies. Protein NMR studies into interactions of the P-domain with ligands will yield insights not easily available from other biophysical techniques and will extend our understanding of MNV attachment to host cells. Such studies require at least partial NMR assignments. Here, we describe the assignment of about 70% of the Ala, Ile, LeuproS, Met, and ValproS methyl groups. An unfavorable distribution of methyl group resonance signals prevents complete assignment based exclusively on 4D HMQC-NOESY-HMQC experiments, yielding assignment of only 55 out of 100 methyl groups. Therefore, we created point mutants and measured pseudo contact shifts, extending and validating assignments based on methyl-methyl NOEs. Of note, the P-domains are present in two different forms caused by an approximate equal distribution of trans- and cis-configured proline residues in position 361.

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