Human DNA Polymerase μ Can Use a Noncanonical Mechanism for Multiple Mn2+-Mediated Functions
Journal
Journal of the American Chemical Society
Journal Volume
141
Journal Issue
21
Pages
8489-8502
Date Issued
2019
Author(s)
Abstract
Recent research on the structure and mechanism of DNA polymerases has continued to generate fundamentally important features, including a noncanonical pathway involving "prebinding" of metal-bound dNTP (MdNTP) in the absence of DNA. While this noncanonical mechanism was shown to be a possible subset for African swine fever DNA polymerase X (Pol X) and human Pol λ, it remains unknown whether it could be the primary pathway for a DNA polymerase. Pol μ is a unique member of the X-family with multiple functions and with unusual Mn2+ preference. Here we report that Pol μ not only prebinds MdNTP in a catalytically active conformation but also exerts a Mn2+ over Mg2+ preference at this early stage of catalysis, for various functions: incorporation of dNTP into a single nucleotide gapped DNA, incorporation of rNTP in the nonhomologous end joining (NHEJ) repair, incorporation of dNTP to an ssDNA, and incorporation of an 8-oxo-dGTP opposite template dA (mismatched) or dC (matched). The structural basis of this noncanonical mechanism and Mn2+ over Mg2+ preference in these functions was analyzed by solving 19 structures of prebinding binary complexes, precatalytic ternary complexes, and product complexes. The results suggest that the noncanonical pathway is functionally relevant for the multiple functions of Pol μ. Overall, this work provides the structural and mechanistic basis for the long-standing puzzle in the Mn2+ preference of Pol μ and expands the landscape of the possible mechanisms of DNA polymerases to include both mechanistic pathways. © 2019 American Chemical Society.
Other Subjects
Polymers; Human DNA polymerase; Important features; Mechanistic pathways; Multiple function; Nonhomologous end joining; Possible mechanisms; Recent researches; Single nucleotides; DNA; 8 oxo deoxyguanosine triphosphate; deoxynucleotide triphosphate; DNA polymerase; DNA polymerase mu; magnesium ion; manganese; nucleotide derivative; single stranded DNA; unclassified drug; DNA directed DNA polymerase; DNA polymerase mu; manganese; Article; binding affinity; catalysis; complex formation; controlled study; DNA end joining repair; enzyme binding; enzyme conformation; enzyme mechanism; enzyme specificity; protein nucleic acid interaction; chemistry; human; isolation and purification; metabolism; molecular model; DNA-Directed DNA Polymerase; Humans; Manganese; Models, Molecular
Type
journal article