Specific binding of human MSH2 center dot 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 p hotoproducts by bacterial (Feng, W,-Y., Lee, E., and Hays, J.B. (1991) Gene tics 129, 1007-1020) and human (Mu, D., Tursun, M., Duckett, D, R., Drummon d, J.T., Modrich, P., and Sancar, A. (1997) Mol. Cell. Biol. 17, 760-769) l ong-patch mismatch-repair systems. Mismatch repair directed specifically ag ainst incorrect bases inserted during semi-conservative DNA replication mig ht efficiently antagonize UV mutagenesis. To test this hypothesis, DNA 51-m ers containing site-specific T-T cis-syn-cyclobutane pyrimidine-dimers or T -T pyrimidine-(6-4')pyrimidinone photoproducts, with all four possible base s opposite the respective 3'-thymines in the photoproducts, were analyzed f or the ability to compete with radiolabeled (T/G)-mismatched DNA for bindin g by highly purified human MSH2 . MSHG heterodimer protein (hMutS alpha). B oth (cyclobutane-dimer)/AG and ((6-4)photoproduct)/AG mismatches competed a bout as well as non-photoproduct T/T mismatches. The two respective pairs o f photoproduct/(A(T or C)) mismatches also showed higher hMutS alpha affini ty than photoproduct/AA "matches"; the apparent affinity of hMutS alpha for the ((6 - 4)photoproduct)/AA"matched" substrate was actually less than tha t for TT/AA homoduplexes. Surprisingly, although hMutS alpha 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 hMutS alpha to DN A containing (photoproduct/base) mismatches and to (T/G)-mismatched DNA was reduced similarly by ATP (in the absence of magnesium).