In vitro mapping studies of the MD145 norovirus (Caliciviridae) ORF1
polyprotein identified two stable cleavage products containing the
viral RNA-dependent
RNA polymerase (RdRp) domains:
ProPol (a precursor comprised of both the
proteinase and polymerase) and Pol (the mature polymerase). The goal of this study was to identify the active form (or forms) of the norovirus polymerase. The recombinant
ProPol (expressed as Pro(-)Pol with an inactivated
proteinase domain to prevent autocleavage) and recombinant Pol were purified after synthesis in bacteria and shown to be active RdRp
enzymes. In addition, the mutant His-E1189A-ProPol
protein (with active
proteinase but with the natural
ProPol cleavage site blocked) was active as an RdRp, confirming that the norovirus
ProPol precursor could possess two enzymatic activities simultaneously. The effects of several
UTP analogs on the RdRp activity of the norovirus and feline calicivirus Pro(-)Pol
enzymes were compared and found to be similar. Our data suggest that the norovirus
ProPol is a bifunctional
enzyme during virus replication. The availability of this recombinant
ProPol enzyme might prove useful in the development of
antiviral drugs for control of the noroviruses associated with acute
gastroenteritis.