The 5' termini of many viral and cellular mRNAs contain sequences of the type 
m7G(5")pppNm. An RNA (guanine-7-)-methyltransferase that specifically methylates 
the 5'-terminal guanosine residue of RNAs ending in the dinucleoside 
triphosphate G(5')pppN- has been purified from the cytoplasm of HeLa cells. 
Approximately two-thirds of the methyltransferase activity detected in an assay 
employing umnethylated vaccinia virus mRNA as acceptor was located in the 
cytoplasm when cells were disrupted by Dounce homogenization; 30% of the 
cytoplasmic activity was associated with ribosomes but was removed by washing 
with 0.5 M KCl. The enzyme was purified 165-fold from the cytoplasm by removing 
nucleic acid by phase partition followed by ammonium sulfate precipitation and 
column chromatography on DEAE-cellulose, denatured DNA-agarose, and CM-Sephadex. 
The partially purified enzyme preparation methylated heterologous tRNAs as well 
as vaccinia mRNA, but the tRNA methyltransferases could be separated from the 
mRNA activity by sucrose gradient sedimentation and gel filtration on Sephadex 
G-200. The product of the partially purified enzyme using vaccinia mRNA as 
substrate was exclusively 7-methylguanosine located in the terminal dinucleoside 
triphosphate. In addition to RNAs and synthetic polyribonucleotides terminating 
in a dinucleoside triphosphate, free G(5')pppG could be methylated but GTP, GDP, 
and G(5')pppG could not. The enzyme also methylated the dinucleoside diphosphate 
G(5')pppG but much less efficiently than G(5')pppG. An S20, W of 3.8, a Stokes 
radius of 3.6 nm, and a molecular weight of 56,000 were obtained from sucrose 
gradient sedimentation and Sephadex G-200 column chromatography.