Collaboration between CC- and A-adding enzymes to build and repair the 3′-terminal CCA of tRNA in Aquifex aeolicus

The universal 3'-terminal CCA sequence of all transfer RNAs (tRNAs) is repaired, and sometimes constructed de novo, by the CCA-adding enzyme [ATP(CTP):tRNA nucleotidyltransferase]. This RNA polymerase has no nucleic acid template, yet faithfully builds the CCA sequence one nucleotide at a time using cytidine triphosphate (CTP) and adenosine triphosphate (ATP) as substrates. AU previously characterized CCA-adding enzymes from all three kingdoms are single polypeptides with CCA-adding activity. Here, we demonstrate through biochemical and genetic approaches that CCA addition in Aquifex aeolicus requires collaboration between two related polypeptides, one that adds CC and another that adds A.