Structural insight into the substrate specificity of DNA Polymerase mu

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Andrea F Moon1, Miguel Garcia-Diaz, Katarzyna Bebenek

  • 1Laboratory of Structural Biology, National Institute of Environmental Health Sciences (National Institutes of Health, US Department of Health and Human Services), 111 T.W. Alexander Drive, MD F3-09, Research Triangle Park, North Carolina 27709, USA.

Nature Structural & Molecular Biology
|December 13, 2006
PubMed

Insights

DNA polymerase mu (Pol mu), a DNA repair enzyme, has unique substrate specificity crucial for nonhomologous DNA end joining (NHEJ). Its crystal structure reveals distinct active site interactions, explaining its specialized function in DNA repair.

Area of Science:

  • Biochemistry
  • Molecular Biology
  • Structural Biology

Background:

  • DNA polymerase mu (Pol mu) is a Family X enzyme implicated in DNA repair pathways.
  • Its unique substrate specificity is essential for its role in nonhomologous DNA end joining (NHEJ).

Purpose of the Study:

  • To elucidate the structural basis of Pol mu's substrate specificity.
  • To understand how Pol mu functions differently from other Family X DNA polymerases.

Main Methods:

  • Determined the 2.4 A crystal structure of the polymerase domain of murine Pol mu bound to gapped DNA and a dNTP.
  • Utilized site-directed mutagenesis (H329A substitution) to assess enzyme activity.

Main Results:

  • The crystal structure revealed unique side chain interactions within the Pol mu active site compared to other Family X polymerases.
  • A specific mutation (H329A) differentially affected Pol mu's activity on various DNA substrates, particularly those lacking complementary template nucleotides at the 3' end.
  • This mutation impaired both template-independent and template-dependent synthesis during NHEJ.

Conclusions:

  • The structural and mutational data provide insights into the molecular mechanisms underlying Pol mu's specialized substrate specificity.
  • These findings help differentiate the functions of closely related mammalian Family X DNA polymerases in DNA repair processes.

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