Trypanosome capping enzymes display a novel two-domain structure.
The ubiquitous m7G cap of eukaryotic mRNAs and of precursors to the spliceosomal small nuclear RNAs (snRNAs) is the result of an essential RNA modification acquired during transcript elongation. In trypanosomes, the m7G cap is restricted to the spliced leader (SL) RNA and the precursors of U2, U3, and U4 snRNAs. mRNA capping in these organisms occurs posttranscriptionally by trans splicing, which transfers the capped SL sequence to the 5' ends of all mRNAs. The SL cap is the most elaborate cap structure known in nature and has been shown to consist of an m7G residue followed by four methylated nucleotides. Using Crithidia fasciculata, we have characterized and purified the guanylyltransferase (capping enzyme), which transfers GMP from GTP to the diphosphate end of RNA. The corresponding gene codes for a protein of 697 amino acids, with the carboxy-terminal half of the C. fasciculata guanylyltransferase containing the six signature motifs previously identified in yeast capping enzymes. The amino-terminal half contains a domain that displays no resemblance to any other domain associated with capping enzymes. Intriguingly, this region harbors a consensus sequence for a phosphate-binding loop which is found in ATP- and GTP-binding proteins. This two-domain structure is also present in the Trypanosoma brucei capping enzyme, which shows 44% overall identity with the C. fasciculata capping enzyme. Thus, this structure appears to be common to all trypanosomatid protozoa and defines a novel class of capping enzymes.
Silva
E
E
Departments of Internal Medicine, Yale University School of Medicine, New Haven, Connecticut 06520-8022, USA.
Ullu
E
E
Kobayashi
R
R
Tschudi
C
C
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D003001:Cloning, Molecular; D017124:Conserved Sequence; D016831:Crithidia fasciculata / Q000201:enzymology* / Q000235:genetics; D016054:DNA, Protozoan; D006157:Guanosine Monophosphate / Q000378:metabolism; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D012316:RNA Nucleotidyltransferases / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D014346:Trypanosoma brucei brucei / Q000201:enzymology* / Q000235:genetics
D016054:DNA, Protozoan; D011993:Recombinant Fusion Proteins; D006157:Guanosine Monophosphate; C035410:GTP-RNA guanylyltransferase; D012316:RNA Nucleotidyltransferases