4X2Y

Crystal structure of a chimeric Murine Norovirus NS6 protease (inactive C139A mutant) in which the P4-P4 prime residues of the cleavage junction in the extended C-terminus have been replaced by the corresponding residues from the NS2-3 junction.

Summary for 4X2Y

• Entry DOI: 10.2210/pdb4x2y/pdb
• Related: 4ASH 4X2V 4X2W 4X2X
• Descriptor: NS6 Protease,NS6 Protease (2 entities in total)
• Functional Keywords: murine norovirus, protease, viral protein
• Biological source: Murine norovirus 1; More
• Total number of polymer chains: 2
• Total formula weight: 37649.03

Authors

Leen, E.N.,Pfeil, M.-P.,Fernandes, H.,Curry, S. (deposition date: 2014-11-27, release date: 2015-02-18, Last modification date: 2024-01-10)

Primary citation

Fernandes, H.,Leen, E.N.,Cromwell, H.,Pfeil, M.P.,Curry, S.
Structure determination of Murine Norovirus NS6 proteases with C-terminal extensions designed to probe protease-substrate interactions.
Peerj, 3:e798-e798, 2015

PubMed Abstract: Noroviruses are positive-sense single-stranded RNA viruses. They encode an NS6 protease that cleaves a viral polyprotein at specific sites to produce mature viral proteins. In an earlier study we obtained crystals of murine norovirus (MNV) NS6 protease in which crystal contacts were mediated by specific insertion of the C-terminus of one protein (which contains residues P5-P1 of the NS6-7 cleavage junction) into the peptide binding site of an adjacent molecule, forming an adventitious protease-product complex. We sought to reproduce this crystal form to investigate protease-substrate complexes by extending the C-terminus of NS6 construct to include residues on the C-terminal (P') side of the cleavage junction. We report the crystallization and crystal structure determination of inactive mutants of murine norovirus NS6 protease with C-terminal extensions of one, two and four residues from the N-terminus of the adjacent NS7 protein (NS6 1', NS6 2', NS6 4'). We also determined the structure of a chimeric extended NS6 protease in which the P4-P4' sequence of the NS6-7 cleavage site was replaced with the corresponding sequence from the NS2-3 cleavage junction (NS6 4' 2|3).The constructs NS6 1' and NS6 2' yielded crystals that diffracted anisotropically. We found that, although the uncorrected data could be phased by molecular replacement, refinement of the structures stalled unless the data were ellipsoidally truncated and corrected with anisotropic B-factors. These corrections significantly improved phasing by molecular replacement and subsequent refinement.The refined structures of all four extended NS6 proteases are very similar in structure to the mature MNV NS6-and in one case reveal additional details of a surface loop. Although the packing arrangement observed showed some similarities to those observed in the adventitious protease-product crystals reported previously, in no case were specific protease-substrate interactions observed.

PubMed: 25755927
DOI: 10.7717/peerj.798

Experimental method

X-RAY DIFFRACTION (2.417 Å)