2RHB

Crystal structure of Nsp15-H234A mutant- Hexamer in asymmetric unit

Summary for 2RHB

• Entry DOI: 10.2210/pdb2rhb/pdb
• Related: 2gth 2h85
• Descriptor: Uridylate-specific endoribonuclease (2 entities in total)
• Functional Keywords: endoribonuclease, sars, hexamer, rna, nsp, viral protein
• Biological source: SARS coronavirus
• Cellular location: Non-structural protein 3: Host membrane; Multi-pass membrane protein (Potential). Non-structural protein 4: Host membrane; Multi-pass membrane protein (Potential). Non-structural protein 6: Host membrane; Multi-pass membrane protein (Potential). Non-structural protein 7: Host cytoplasm, host perinuclear region (By similarity). Non-structural protein 8: Host cytoplasm, host perinuclear region (By similarity). Non-structural protein 9: Host cytoplasm, host perinuclear region (By similarity). Non-structural protein 10: Host cytoplasm, host perinuclear region (By similarity). Helicase: Host endoplasmic reticulum-Golgi intermediate compartment (Potential). Uridylate-specific endoribonuclease: Host cytoplasm, host perinuclear region (By similarity): P59641
• Total number of polymer chains: 6
• Total formula weight: 236554.76

Authors

Palaninathan, S.,Bhardwaj, K.,Alcantara, J.M.O.,Guarino, L.,Yi, L.L.,Kao, C.C.,Sacchettini, J. (deposition date: 2007-10-08, release date: 2007-11-27, Last modification date: 2023-08-30)

Primary citation

Bhardwaj, K.,Palaninathan, S.,Alcantara, J.M.,Yi, L.L.,Guarino, L.,Sacchettini, J.C.,Kao, C.C.
Structural and functional analyses of the severe acute respiratory syndrome coronavirus endoribonuclease Nsp15.
J.Biol.Chem., 283:3655-3664, 2008

PubMed Abstract: The severe acute respiratory syndrome (SARS) coronavirus encodes several RNA-processing enzymes that are unusual for RNA viruses, including Nsp15 (nonstructural protein 15), a hexameric endoribonuclease that preferentially cleaves 3' of uridines. We solved the structure of a catalytically inactive mutant version of Nsp15, which was crystallized as a hexamer. The structure contains unreported flexibility in the active site of each subunit. Substitutions in the active site residues serine 293 and proline 343 allowed Nsp15 to cleave at cytidylate, whereas mutation of leucine 345 rendered Nsp15 able to cleave at purines as well as pyrimidines. Mutations that targeted the residues involved in subunit interactions generally resulted in the formation of catalytically inactive monomers. The RNA-binding residues were mapped by a method linking reversible cross-linking, RNA affinity purification, and peptide fingerprinting. Alanine substitution of several residues in the RNA-contacting portion of Nsp15 did not affect hexamer formation but decreased the affinity of RNA binding and reduced endonuclease activity. This suggests a model for Nsp15 hexamer interaction with RNA.

PubMed: 18045871
DOI: 10.1074/jbc.M708375200

Experimental method

X-RAY DIFFRACTION (2.8 Å)