TRYPANOSOME CAPPING ENZYMES DISPLAY A NOVEL 2-DOMAIN STRUCTURE

The ubiquitous m(7)G 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 trypanosom es, the m7G cap is restricted to the spliced leader (SL) RNA and the p recursors of U2, U3, and U4 snRNAs. mRNA capping in these organisms oc curs posttranscriptionally by trans splicing, which transfers the capp ed SL sequence to the 5' ends of all mRNAs. The SL cap is the most ela borate cap structure known in nature and has been shown to consist of an m(7)G residue followed by four methylated nucleotides. Using Crithi dia fasciculata, we have characterized and purified the guanylyltransf erase (capping enzyme), which transfers GMP from GTP to the diphosphat e end of RNA. The corresponding gene codes for a protein of 697 amino acids, with the carboxy-terminal half of the C. fasciculata guanylyltr ansferase 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 e nzymes. Intriguingly, this region harbors a consensus sequence for a p hosphate-binding loop which is found in ATP- and GTP-binding proteins. This two-domain structure is also present in the Trypanosoma brucei c apping enzyme, which shows 44% overall identity with the C. fasciculat a capping enzyme. Thus, this structure appears to be common to all try panosomatid protozoa and defines a novel class of capping enzymes.