Dodecamer Structure of Severe Acute Respiratory Syndrome Coronavirus Nonstructural Protein nsp10

ABSTRACT The severe acute respiratory syndrome coronavirus (SARS-CoV) nonstructural proteins nsp1 to nsp16 have been implicated by genetic analysis in the assembly of a functional replication/transcription complex. We report the crystal structure of nsp10 from SARS-CoV at 2.1-Å resolution. The nsp10 structure has a novel fold, and 12 identical subunits assemble to form a unique spherical dodecameric architecture. Two zinc fingers have been identified from the nsp10 monomer structure with the sequence motifs C-(X)2-C-(X)5-H-(X)6-C and C-(X)2-C-(X)7-C-(X)-C. The nsp10 crystal structure is the first of a new class of zinc finger protein three-dimensional structures to be revealed experimentally. The zinc finger sequence motifs are conserved among all three coronavirus antigenic groups, implicating an essential function for nsp10 in all coronaviruses. Based on the structure, we propose that nsp10 is a transcription factor for coronavirus replication/transcription.

[1]  A. Vagin,et al.  MOLREP: an Automated Program for Molecular Replacement , 1997 .

[2]  P. Kraulis A program to produce both detailed and schematic plots of protein structures , 1991 .

[3]  M. Denison,et al.  Four Proteins Processed from the Replicase Gene Polyprotein of Mouse Hepatitis Virus Colocalize in the Cell Periphery and Adjacent to Sites of Virion Assembly , 2000, Journal of Virology.

[4]  Zihe Rao,et al.  Structural insights into SARS coronavirus proteins , 2005, Current Opinion in Structural Biology.

[5]  Zihe Rao,et al.  Insights into SARS-CoV transcription and replication from the structure of the nsp7–nsp8 hexadecamer , 2005, Nature Structural &Molecular Biology.

[6]  J. Thompson,et al.  The CLUSTAL_X windows interface: flexible strategies for multiple sequence alignment aided by quality analysis tools. , 1997, Nucleic acids research.

[7]  J. Thornton,et al.  PROCHECK: a program to check the stereochemical quality of protein structures , 1993 .

[8]  George M Sheldrick,et al.  Substructure solution with SHELXD. , 2002, Acta crystallographica. Section D, Biological crystallography.

[9]  Sonia Longhi,et al.  The severe acute respiratory syndrome-coronavirus replicative protein nsp9 is a single-stranded RNA-binding subunit unique in the RNA virus world. , 2004, Proceedings of the National Academy of Sciences of the United States of America.

[10]  Eugene Krissinel,et al.  The CCP4 molecular-graphics project. , 2002, Acta crystallographica. Section D, Biological crystallography.

[11]  J. Zou,et al.  Improved methods for building protein models in electron density maps and the location of errors in these models. , 1991, Acta crystallographica. Section A, Foundations of crystallography.

[12]  R J Read,et al.  Crystallography & NMR system: A new software suite for macromolecular structure determination. , 1998, Acta crystallographica. Section D, Biological crystallography.

[13]  G. Gao,et al.  The crystal structures of severe acute respiratory syndrome virus main protease and its complex with an inhibitor , 2003, Proceedings of the National Academy of Sciences of the United States of America.

[14]  Thomas C. Terwilliger,et al.  Automated MAD and MIR structure solution , 1999, Acta crystallographica. Section D, Biological crystallography.

[15]  E A Merritt,et al.  Raster3D: photorealistic molecular graphics. , 1997, Methods in enzymology.

[16]  Kin Moy,et al.  Structural Basis of Severe Acute Respiratory Syndrome Coronavirus ADP-Ribose-1″-Phosphate Dephosphorylation by a Conserved Domain of nsP3 , 2005, Structure.

[17]  Xavier Robert,et al.  ESPript/ENDscript: extracting and rendering sequence and 3D information from atomic structures of proteins , 2003, Nucleic Acids Res..

[18]  David I Stuart,et al.  The nsp9 Replicase Protein of SARS-Coronavirus, Structure and Functional Insights , 2004, Structure.

[19]  K. Yuen,et al.  Correction: Design of Wide-Spectrum Inhibitors Targeting Coronavirus Main Proteases , 2005, PLoS Biology.

[20]  Stuart G. Siddell,et al.  Viral Replicase Gene Products Suffice for Coronavirus Discontinuous Transcription , 2001, Journal of Virology.

[21]  G. Sheldrick,et al.  Advances in direct methods for protein crystallography. , 1999, Current opinion in structural biology.

[22]  R M Esnouf,et al.  An extensively modified version of MolScript that includes greatly enhanced coloring capabilities. , 1997, Journal of molecular graphics & modelling.

[23]  Z. Otwinowski,et al.  Processing of X-ray diffraction data collected in oscillation mode. , 1997, Methods in enzymology.

[24]  Anastassis Perrakis,et al.  Developments in the CCP4 molecular-graphics project. , 2004, Acta crystallographica. Section D, Biological crystallography.

[25]  J. Ziebuhr The Coronavirus Replicase , 2005, Current topics in microbiology and immunology.