Limited terminal transferase in human DNA polymerase μ defines the required balance between accuracy and efficiency in NHEJ

DNA polymerase mu (Pol mu) is a family X member implicated in DNA repair, with template-directed and terminal transferase (template-independent) activities. It has been proposed that the terminal transferase activity of Pol mu can be specifically required during non-homologous end joining (NHEJ) to create or increase complementarity of DNA ends. By site-directed mutagenesis in human Pol mu, we have identified a specific DNA ligand residue (Arg(387)) that is responsible for its limited terminal transferase activity compared to that of human TdT, its closest homologue (42% amino acid identity). Pol mu mutant R387K ( mimicking TdT) displayed a large increase in terminal transferase activity, but a weakened interaction with ssDNA. That paradox can be explained by the regulatory role of Arg387 in the translocation of the primer from a nonproductive E: DNA complex to a productive E: DNA: dNTP complex in the absence of a templating base, which is probably the rate limiting step during template-independent synthesis. Further, we show that the Pol mu switch from terminal transferase to templated insertions in NHEJ reactions is triggered by recognition of a 5'-P at a second DNA end, whose 3'-protrusion could provide a templating base to facilitate such a special "pre-catalytic translocation step.'' These studies shed light on the mechanism by which a rate-limited terminal transferase activity in Pol mu could regulate the balance between accuracy and necessary efficiency, providing some variability during NHEJ.