Purification and Characterization of polκ, a DNA Polymerase Encoded by the Human DINB1 Gene*

By V. Gerlach, W. J. Feaver, P. Fischhaber, E. Friedberg. Journal of Biological Chemistry, 2001

112 citations recorded by the source corpus.

The Escherichia coli dinB gene encodes DNA polymerase (pol) IV, a protein involved in increasing spontaneous mutations in vivo. The protein-coding region ofDINB1, the human ortholog of DNA pol IV, was fused to glutathione S-transferase and expressed in insect cells. The purified fusion protein was shown to be a template-directed DNA polymerase that we propose to designate polκ. Human polκ lacks detectable 3′ → 5′ proofreading exonuclease activity and is not stimulated by recombinant human proliferating cell nuclear antigen in vitro. Between pH 6.5 and 8.5, human polκ possesses optimal activity at 37 °C over the pH range 6.5–7.5, and is insensitive to inhibition by aphidicolin, dideoxynucleotides, or NaCl up to 50 mm. Either Mg2+ or Mn2+ can satisfy a metal cofactor requirement for polκ activity, with Mg2+ being preferred. Human polκ is unable to bypass a cisplatin adduct in the template. However, polκ shows limited bypass of an 2-acetylaminofluorene lesion and can incorporate dCTP or dTTP across from this lesion, suggesting that the bypass is potentially mutagenic. These results are consistent with a model in which polκ acts as a specialized DNA polymerase whose possible role is to facilitate the replication of templates containing abnormal bases, or possessing structurally aberrant replication forks that inhibit normal DNA synthesis.

Subfields: Cancer Research, Molecular Biology

View Paper: https://www.semanticscholar.org/paper/dae84efb3bd99b88062072d74efe544fbe1a2b35

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Citations in the corpus

Mechanisms of human DNA repair: an update. (2003, background): “The human polymerase kappa (Pol␬; DINB; Gerlach et al., 1999, 2001; Johnson et al., 2000a; Ogi et al., 1999) is the homologue of the E. coli dinB-encoded DNA polymerase IV.”
Regulation of the Rev1–pol ζ complex during bypass of a DNA interstrand cross‐link (2015, unknown): “Because TLS polymerases b , g , j , ι , and k are insensitive to aphidicolin (Nealon et al , 1996; Masutani et al , 1999; Tissier et al , 2000; Gerlach et al , 2001; Shimazaki et al , 2002) and pol f is not required for approach (Ra¨schle et al , 2008), these data imply that a replicative DNA…”