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Publication Abstract from PubMed

Encephalomyocarditis virus (EMCV) is a member of the Cardiovirus genus within the large Picornaviridae family that includes a number of important human and animal pathogens. The RNA-dependent RNA polymerase (RdRP) 3Dpol is a key enzyme for viral genome replication. In this study, we report the X-ray structures of two different crystal forms of the EMCV RdRP determined at 2.8 and 2.15 A resolution. The in vitro elongation and VPg uridylylation activities of the purified enzyme have also been demonstrated. Although the overall structure of EMCV 3Dpol is shown to be similar to the known RdRPs of other members of the Picornaviridae family, structural comparisons show a large reorganization of the active site cavity in one of the crystal forms. The rearrangement affects mainly the motif A, where the conserved residue Asp240, involved in rNTP selection and its neighbor residue, Phe239, move about 10 A from its expected position, within the ribose binding pocket, towards the entrance of the rNTP tunnel. This altered conformation of motif A is stabilized by a cation-pi interaction established between the aromatic ring of Phe239 and the side chain of Lys56, within the fingers domain. Other contacts, involving Phe239 and different residues of motif F are also observed. The movement of motif A is connected with important conformational changes in the fingers region flanked by residues 54 to 63, harboring Lys56, and in the polymerase N-terminus. The structures determined in this work provide essential information for studies on the Cardiovirus RNA replication process and may have important implications for the development of new antivirals targeting the altered conformation of motif A. IMPORTANCE: The Picornaviridae family is one of the largest virus families known, including many important human and animal pathogens. The RNA-dependent RNA polymerase (RdRP) 3Dpol is a key enzyme for picornavirus genome replication and a validated target for the development of antiviral therapies. Solving the X-ray structure of the first cardiovirus RdRP, the EMCV 3Dpol, we captured an altered conformation of a conserved motif in the polymerase active site (motif A), containing the aspartic acid residue involved in rNTP selection and binding. This altered conformation of motif A, interfering with the correct positioning of the rNTP substrate in the active site, is stabilized by a number of residues strictly conserved among picornaviruses. The rearrangements observed suggest that this motif A segment is a dynamic element that can be modulated by external effectors, either activating or inhibiting the enzyme activity and this type of modulation appears to be general to all picornaviruses.

The crystal structure of a Cardiovirus RNA-dependent RNA polymerase reveals an unusual conformation of the polymerase active site.,Vives-Adrian L, Lujan C, Oliva B, van der Linden L, Selisko B, Coutard B, Canard B, van Kuppeveld FJ, Ferrer-Orta C, Verdaguer N J Virol. 2014 Mar 5. PMID:24600002^[1]

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.