Structural insight into the substrate specificity of DNA Polymerase μ

DNA polymerase μ (Pol μ) is a family X enzyme with unique substrate specificity that contributes to its specialized role in nonhomologous DNA end joining (NHEJ). To investigate Pol μ's unusual substrate specificity, we describe the 2.4 Å crystal structure of the polymerase domain of murine Pol μ bound to gapped DNA with a correct dNTP at the active site. This structure reveals substrate interactions with side chains in Pol μ that differ from other family X members. For example, a single amino acid substitution, H329A, has little effect on template-dependent synthesis by Pol μ from a paired primer terminus, but it reduces both template-independent and template-dependent synthesis during NHEJ of intermediates whose 3′ ends lack complementary template strand nucleotides. These results provide insight into the substrate specificity and differing functions of four closely related mammalian family X DNA polymerases.

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