RNA 5'-triphosphatase, nucleoside triphosphatase, and guanylyltransferase activities of baculovirus LEF-4 protein.
Autographa californica nuclear polyhedrosis virus late and very late mRNAs are transcribed by an RNA polymerase consisting of four virus-encoded polypeptides: LEF-8, LEF-9, LEF-4, and p47. The 464-amino-acid LEF-4 subunit contains the signature motifs of GTP:RNA guanylyltransferases (capping enzymes). Here, we show that the purified recombinant LEF-4 protein catalyzes two reactions involved in RNA cap formation. LEF-4 is an RNA 5'-triphosphatase that hydrolyzes the gamma phosphate of triphosphate-terminated RNA and a guanylyltransferase that reacts with GTP to form a covalent protein-guanylate adduct. The RNA triphosphatase activity depends absolutely on a divalent cation; the cofactor requirement is satisfied by either magnesium or manganese. LEF-4 also hydrolyzes ATP to ADP and Pi (Km = 43 microM ATP; Vmax = 30 s-1) and GTP to GDP and Pi. The LEF-4 nucleoside triphosphatase (NTPase) is activated by manganese or cobalt but not by magnesium. The RNA triphosphatase and NTPase activities of baculovirus LEF-4 resemble those of the vaccinia virus and Saccharomyces cerevisiae mRNA capping enzymes. We suggest that these proteins comprise a novel family of metal-dependent triphosphatases.
Gross
C H
CH
Molecular Biology Program, Sloan-Kettering Institute, New York, New York 10021, USA.
Shuman
S
S
D017766:Acid Anhydride Hydrolases / Q000235:genetics / Q000378:metabolism*; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D017124:Conserved Sequence; D017931:DNA Primers / Q000235:genetics; D019143:Evolution, Molecular; D006157:Guanosine Monophosphate / Q000378:metabolism; D006868:Hydrolysis; D008780:Methyltransferases / Q000235:genetics / Q000378:metabolism; D008969:Molecular Sequence Data; D009097:Multienzyme Complexes / Q000235:genetics / Q000378:metabolism; D017924:Nucleopolyhedroviruses / Q000201:enzymology* / Q000235:genetics; D043583:Nucleoside-Triphosphatase; D009713:Nucleotidyltransferases / Q000235:genetics / Q000378:metabolism*; D010744:Phosphoric Monoester Hydrolases / Q000235:genetics / Q000378:metabolism; D012315:RNA Caps / Q000378:metabolism; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000201:enzymology / Q000235:genetics; D017386:Sequence Homology, Amino Acid; D014616:Vaccinia virus / Q000201:enzymology / Q000235:genetics; D014764:Viral Proteins / Q000235:genetics / Q000378:metabolism*; D000098556:mRNA Guanylyltransferases
D017931:DNA Primers; D009097:Multienzyme Complexes; D012315:RNA Caps; D011994:Recombinant Proteins; D014764:Viral Proteins; C084398:late expression factor 4, baculovirus; D006157:Guanosine Monophosphate; D008780:Methyltransferases; D009713:Nucleotidyltransferases; D000098556:mRNA Guanylyltransferases; D010744:Phosphoric Monoester Hydrolases; C027731:capping enzyme, vaccinia virus; D017766:Acid Anhydride Hydrolases; C023090:RNA triphosphatase; D043583:Nucleoside-Triphosphatase