LB-233

Metal ions are necessary for DNA synthesis by translesion Y family DNA polymerase iota (polι). Previous characterization at 5 - 8 mM Mg2+ revealed a very distributive behavior of the enzyme in vitro, especially error-prone incorporation opposite template T. Here we report our analysis of polι primer extension on an undamaged template in the presence of eight divalent metal cations. We compared polι with two other closely related human Y-family enzymes, DNA polymerases eta (polη) and pol kappa (polκ), in their requirements for metal ions. Both polη and polκ, have performed the most processive DNA synthesis in the presence of Mg2+ ions. In contrast, polι was most processive in Mn2+ supplemented reactions. We found that polι pauses less opposite template T in Mn2+ reactions, in agreement with the kinetic parameters which had revealed less misincorporation at template T in the presence of Mn2+, than in presence of Mg2+. polι was active in a broad range of Mn2+ concentrations and demonstrated ability to bypass thymine dimer with an efficiency up to 39% of an undamaged template. Taken together with the higher affinity of polι to Mn2+ vs Mg2+ ions, our results suggest that Mn2+ is the biologically relevant metal ion cofactor for polι DNA synthesis.

98th AACR Annual Meeting-- Apr 14-18, 2007; Los Angeles, CA