Crystallographic Evidence for an Induced Fit Mechanism to Enhance Fidelity in DNA Polymerase Beta. Michael R. Sawaya, Huguette Pelletier, Rajendra Prasad, Samuel H. Wilson, Joseph Kraut University of California, San Diego, 9500 Gilman Drive, La Jolla, California 92093-0506; National Institute of Environmental Health Sciences, P.O. Box 12233, Research Triangle Park, NC 27709-2233; Baylor Medical College, One Baylor Plaza, Houston, TX 77030.

Structural evidence suggests that DNA polymerase beta (pol beta) may employ an induced fit mechanism to enhance fidelity in filling single nucleotide gaps formed in the base excision repair pathway of mammalian cells. Human pol beta has been crystallized in complexes representing three intermediates in the gap-filling reaction: (1) the binary complex with a gapped DNA substrate, (2) the ternary complex including ddCTP and (3) the binary complex with nicked DNA product (2.2-2.6 Angstrom resolution). In the ternary complex, the presence of a Watson-Crick base pair between the templating residue and a dNTP analogue (ddCTP) appears to induce rotation of the entire thumb subdomain into a closed position relative to the two binary complexes. In the closed position, the thumb contacts the face and edge of the forming base pair which have been exposed by a unique 90 degree kink in the DNA located at the 5'-phosphate of the templating residue. Thumb closure also triggers further conformational changes which position catalytic residue Asp192, ddCTP, and the template for nucleotidyl transfer, effectively assembling the active site. We speculate that unfavorable contacts between the thumb and a non-Watson-Crick base pair would hinder thumb closure and assembly of the active site, hence hindering incorporation of an incorrect nucleotide.