Identification And Characterization Of A Ribose 2'-O-methyltransferase Encoded By The Ronivirus Branch Of Nidovirales

The order Nidovirales currently comprises four virus families:Arteriviridae,Coronaviridae(divided into the subfamilies Coronavirinae and Torovirinae),Roniviridae and the recently recognized Mesoniviridae.RNA cap formation and methylation have been best studied for coronaviruses,with emphasis on the identification and characterization of two virus-encoded methyltransferases(MTases)involved in RNA capping,a guanine-N7-MTase and a ribose-2'-O-MTase.Although bioinformatic analyses suggest that these MTases may also be encoded by other nidoviruses with large genomes,such as toroviruses and roniviruses,no experimental evidence has been reported thus far.In this study,we show that a ronivirus,gill-associated nidovirus,encodes the 2'-O-MTase activity,although we could not detect 2'-O-MTase activity for the homologous protein of a torovirus,equine torovirus,which is more closely related to coronaviruses.Like the coronavirus 2'-O-MTase,the roniviral 2'-O-MTase harbors a catalytic K-D-K-E tetrad and possibly a Rossman-like fold that is conserved among 2'-O-MTases and can only target the N7-methylated cap structure of adenylate-primed RNA substrates.However,in contrast with coronavirus,roniviral 2'-O-MTase does not require a protein cofactor for stimulation of its activity,and differs in its preference for several biochemical parameters such as reaction temperature and pH.Furthermore,the ronivirus 2'-O-MTase can be targeted by MTase inhibitors.These results extend our current understanding of nidovirus RNA cap formation and methylation beyond the coronavirus family.