Specific binding of human MSH2·MSH6 mismatch-repair protein heterodimers to DNA incorporating thymine- or uracil-containing UV light photoproducts opposite mismatched bases

Previous studies have demonstrated recognition of DNA-containing UV light photoproducts by bacterial (Feng, W.-Y., Lee, E., and Hays, J. B. (1991) Genetics 129, 1007-1020) and human (Mu, D., Tursun, M., Duckett, D. R., Drummond, J. T., Modrich, P., and Sancar, A. (1997) Mol. Cell Biol. 17, 760-769) long-patch mismatch-repair systems. Mismatch repair directed specifically against incorrect bases inserted during semi-conservative DNA replication might efficiently antagonize UV mutagenesis. To test this hypothesis, DNA 51-mers containing site-specific T-T cis-syn-cyclobutane pyrimidine-dimers or T-T pyrimidine-(6-4')pyrimidinone photoproducts, with all four possible bases opposite the respective 3'-thymines in the photoproducts, were analyzed for the ability to compete with radiolabeled (T/G)mismatched DNA for binding by highly purified human MSH2·MSH6 heterodimer protein (hMutSα). Both (cyclobutane-dimer)/AG and ((6- 4)photoproduct)/AG mismatches competed about as well as non-photoproduct T/T mismatches. The two respective pairs of photoproduct/(A(T or C)) mismatches also showed higher hMutSα affinity than photoproduct/AA 'matches'; the apparent affinity of hMutSα for the ((6-4)photoproduct)/AA-'matched' substrate was actually less than that for TT/AA homoduplexes. Surprisingly, although hMutSα affinities for both non-photoproduct UU/GG double mismatches and for (uracil-cyclobutane-dimer)/AG single mismatches were high, affinity for the (uracil-cyclobutane-dimer)/GG mismatch was quite low. Equilibrium binding of hMutSα to DNA containing (photoproduct/base) mismatches and to (T/G)-mismatched DNA was reduced similarly by ATP (in the absence of magnesium).