Disruption of the helix-u-turn-helix motif of MutS protein: Loss of subunit dimerization, mismatch binding and ATP hydrolysis

The DNA mismatch repair protein, MutS, is a dimeric protein that recognizes mismatched bases and has an intrinsic ATPase activity. Here, a series of T ag MutS proteins having C-terminal truncations in the vicinity of a highly conserved helix-u-turn-helix (HuH) motif are assessed for subunit oligomeri zation, ATPase activity and DNA mismatch binding. Those proteins containing an intact HuH region are dimers; those without the HuH region are predomin antly monomers in solution. Steady-state kinetics of truncated but dimeric MutS proteins reveals only modest decreases in their ATPase activity compar ed to full-length protein. Ln contrast, disruption of the HuH region result s in a greatly attenuated ATPase activity. In addition, only dimeric MutS p roteins are proficient for mismatch binding. Finally, an analysis of the mi smatch repair competency of truncated Escherichia coli MutS proteins in a r ifampicin mutator assay confirms that the HuH region is critical for in viv o function. These findings indicate that dimerization is critical for both the ATPase and DNA mismatch binding activities of MutS, and corroborate sev eral key features of the MutS structure recently deduced from X-ray crystal lographic studies.