Biochemical studies on poliovirus polypeptide 2C: evidence for ATPase activity.
Poliovirus 2C is a nonstructural polypeptide proposed to function in viral RNA replication. Poliovirus 2C is a member of a rapidly expanding family of proteins containing a consensus for nucleotide binding (NTP-B). Site-directed mutagenesis of conserved residues in the consensus A and B sites have suggested a functional role for the NTP-B motif in viral RNA replication and proliferation of poliovirus. We have expressed wildtype 2C and a 2C mutant, carrying a single amino acid exchange in the NTP-B motif A (Lys135Gln) using the baculovirus system. Both wildtype and mutant proteins are membrane associated. Following membrane solubilization, we have purified wildtype and mutant proteins to near homogeneity using conventional chromatography. We present biochemical evidence that wildtype 2C copurifies with an ATPase activity that is absent in the mutant preparation.
Mirzayan
C
C
Department of Molecular Microbiology, State University of New York at Stony Brook 11794.
Wimmer
E
E
D000251:Adenosine Triphosphatases / Q000302:isolation & purification / Q000378:metabolism*; D000595:Amino Acid Sequence; D000818:Animals; D016367:Baculoviridae / Q000235:genetics; D001483:Base Sequence; D015153:Blotting, Western; D002352:Carrier Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D002460:Cell Line; D003001:Cloning, Molecular; D017931:DNA Primers; D006367:HeLa Cells; D006801:Humans; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D017361:Viral Nonstructural Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*
D002352:Carrier Proteins; D017931:DNA Primers; D017361:Viral Nonstructural Proteins; C076023:2C protein, viral; D000251:Adenosine Triphosphatases