PURIFICATION OF A FUNCTIONAL ENZYMATIC EDITING COMPLEX FROM TRYPANOSOMA-BRUCEI MITOCHONDRIA

Kinetoplastid mitochondrial RNA editing, the insertion and deletion of U residues, is catalyzed by sequential cleavage, U addition or remova l, and ligation reactions and is directed by complementary guide RNAs. We have purified a similar to 20S enzymatic complex from Trypanosoma brucei mitochondria that catalyzes a complete editing reaction in vitr o, This complex possesses all four activities predicted to catalyze RN A editing: gRNA-directed endonuclease, terminal uridylyl transferase, 3' U-specific exonuclease, and RNA ligase, However, it does not contai n other putative editing complex components: gRNA-independent endonucl ease, RNA helicase, endogenous gRNAs or pre-mRNAs, or a 25 kDa gRNA-bi nding protein. The complex is composed of eight major polypeptides, th ree of which represent RNA ligase, These findings identify polypeptide s representing catalytic editing factors, reveal the nature of this si milar to 20S editing complex, and suggest a new model of editosome ass embly.