X-ray Crystallographic Structure of theMurine Norovirus protease at 1.66 Å Resolutionand Functional Studies of the β-ribbon
Independent thesis Advanced level (degree of Master (Two Years)), 20 credits / 30 HE creditsStudent thesis
In humans, noroviruses (NVs) cause acute epidemic and viral gastroenteritis. NVs do not only infect humans; viruseshave also been found in pigs, cows, sheep, mice and dogs. The focus in this project has been on the murine norovirus(MNV). MNV is a member of the viral family Caliciviridae and it consists of a single-stranded, positive sense RNAgenome. The genome includes three open reading frames (ORFs), ORF1 encodes for a polyprotein that consists of theprecursor to the 6-7 non-structural (NS) proteins. The polyprotein is cleaved by the NS6 protease. The NS6 isresponsible for all the cleaving in ORF1 and that makes it an attractive target for antiviral drugs. The NS6 proteinstructure has been determined at 1.66 Å resolution using X-ray diffraction techniques. Surprisingly, the electrondensity map revealed density for a peptide bound in the active site. The peptide had a length of 7 residues andoriginated from the C-terminus of another chain in an adjacent asymmetric unit. The active site triad was composed ofthe conserved residues; histidine 30, aspargine 54 and cysteine 139, however in the structure the cysteine 139 ismutated to an alanine to inactivate the protease. Activity assays were performed to probe the importance of the residuein position 109 in the β-ribbon located close to the active site. The three full-length constructs with the mutations;I109A, I109S and I109T were found to have less activity than the full-length wt (1-183). A truncated protease, lacking9 residues in the C-terminus, also had less activity. This indicates that the terminal residues are also important foractivity.
Place, publisher, year, edition, pages
2011. , 66 p.
virus, polyprotein, Caliciviridae, aggregation, Chymotrypsin-like protease
Medical Biotechnology (with a focus on Cell Biology (including Stem Cell Biology), Molecular Biology, Microbiology, Biochemistry or Biopharmacy)
IdentifiersURN: urn:nbn:se:liu:diva-70426ISRN: LITH-IFM-A-EX--11/2486--SEOAI: oai:DiVA.org:liu-70426DiVA: diva2:439323
Subject / course
2011-06-09, BL-32, Linköping, 09:30 (Swedish)
UppsokPhysics, Chemistry, Mathematics
Curry, Stephen, Professor