ATP hydrolysis induces expansion of MutS contacts on heteroduplex: a case for MutS treadmilling?
An unsolved problem in E. coli mismatch repair is how the MutS-MutL complex communicates positional information of a mismatch to MutH. MutS is bound to a mismatch in the absence of ATP, exhibiting a short DNase I footprint that is dramatically expanded in ATP hydrolysis. The same is corroborated by restriction enzyme site protection far away from the mismatch. High-resolution gel-shift analyses revealed that super-shifted specific complexes, presumably containing multiple MutS homodimers on the same heteroduplex, were generated during ATP hydrolysis. Such complexes are largely nonspecific in "minus ATP" or in ATP gamma S conditions. Specific ternary complexes of MutS-MutL-heteroduplexes were formed only during ATP hydrolysis. These results suggest that MutS loading onto a mismatch induces the formation of a higher-order complex containing multiple MutS homodimers, presumably through a putative "treadmilling action" that is ATP-hydrolysis dependent. Such a higher-order MutS complex productively interacts with MutL in ATP-hydrolyzing conditions and generates a specific ternary complex, which might communicate with MutH. This model should neither depend on nor give rise to the spooling of DNA. This was corroborated when we observed footprint extension in ATP-hydrolyzing conditions, despite the heteroduplex ends being tethered to agarose beads that block helical rotations.
Joshi
Amita
A
Department of Biological Sciences, Tata Institute of Fundamental Research, Homi Bhabha Road, Bombay 400 005, India.
Rao
Basuthkar J
BJ
D000251:Adenosine Triphosphatases; D000255:Adenosine Triphosphate / Q000378:metabolism*; D001426:Bacterial Proteins / Q000378:metabolism*; D001483:Base Sequence; D004247:DNA; D018983:DNA Footprinting; D004262:DNA Restriction Enzymes / Q000378:metabolism; D004268:DNA-Binding Proteins; D029968:Escherichia coli Proteins; D006868:Hydrolysis; D000070956:MutL Proteins; D051719:MutS DNA Mismatch-Binding Protein; D009692:Nucleic Acid Heteroduplexes; D011485:Protein Binding
D001426:Bacterial Proteins; D004268:DNA-Binding Proteins; D029968:Escherichia coli Proteins; C074770:MutL protein, E coli; D009692:Nucleic Acid Heteroduplexes; D000255:Adenosine Triphosphate; D004247:DNA; D004262:DNA Restriction Enzymes; D000251:Adenosine Triphosphatases; D000070956:MutL Proteins; D051719:MutS DNA Mismatch-Binding Protein; C497175:MutS protein, E coli