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Journal of Virology, April 2007, p. 3583-3596, Vol. 81, No. 7
0022-538X/07/$08.00+0     doi:10.1128/JVI.02306-06
Copyright © 2007, American Society for Microbiology. All Rights Reserved.

Crystal Structure of Poliovirus 3CD Protein: Virally Encoded Protease and Precursor to the RNA-Dependent RNA Polymerase{triangledown}

Laura L. Marcotte,1 Amanda B. Wass,1 David W. Gohara,1,{dagger} Harsh B. Pathak,2,{ddagger} Jamie J. Arnold,2 David J. Filman,1 Craig E. Cameron,2 and James M. Hogle1*

Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, Massachusetts 02115,1 Department of Biochemistry and Molecular Biology, Pennsylvania State University, University Park, Pennsylvania 168022

Received 20 October 2006/ Accepted 19 January 2007

Poliovirus 3CD is a multifunctional protein that serves as a precursor to the protease 3Cpro and the viral polymerase 3Dpol and also plays a role in the control of viral replication. Although 3CD is a fully functional protease, it lacks polymerase activity. We have solved the crystal structures of 3CD at a 3.4-Å resolution and the G64S fidelity mutant of 3Dpol at a 3.0-Å resolution. In the 3CD structure, the 3C and 3D domains are joined by a poorly ordered polypeptide linker, possibly to facilitate its cleavage, in an arrangement that precludes intramolecular proteolysis. The polymerase active site is intact in both the 3CD and the 3Dpol G64S structures, despite the disruption of a network proposed to position key residues in the active site. Therefore, changes in molecular flexibility may be responsible for the differences in fidelity and polymerase activities. Extensive packing contacts between symmetry-related 3CD molecules and the approach of the 3C domain's N terminus to the VPg binding site suggest how 3Dpol makes biologically relevant interactions with the 3C, 3CD, and 3BCD proteins that control the uridylylation of VPg during the initiation of viral replication. Indeed, mutations designed to disrupt these interfaces have pronounced effects on the uridylylation reaction in vitro.

* Corresponding author. Mailing address: Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, 240 Longwood Avenue, Boston, MA 02115. Phone: (617) 432-3918. Fax: (617) 432-4360. E-mail: jhogle{at}hms.harvard.edu.

{triangledown} Published ahead of print on 24 January 2007.

{dagger} Present address: Center for Computational Biology, Washington University School of Medicine, St. Louis, MO 63110.

{ddagger} Present address: Department of Medical Oncology, Fox Chase Cancer Center, Philadelphia, PA 19111.

Journal of Virology, April 2007, p. 3583-3596, Vol. 81, No. 7
0022-538X/07/$08.00+0     doi:10.1128/JVI.02306-06
Copyright © 2007, American Society for Microbiology. All Rights Reserved.



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