Abstract: Msh2-Msh6 is a eukaryotic ATP-dependent mismatch repair (MMR) protein that plays a key role in maintaining the fidelity of the genetic code. The protein recognizes and initiates repair for single base pair mismatches and short insertion/deletion loops (IDL). Fluorescence in solution assays have revealed that Msh2-Msh6 binds with the highest affinity to G:T mismatch DNA. Consequently, deficiencies in Msh2-Msh6 function are implicated in cancer etiology and specifically that of hereditary nonpolyposis colorectal cancer. Interestingly, fluorescence in-solution assays have revealed that Msh2-Msh6 binds to Holliday Junction DNA with 1:1 stoichiometry and a binding affinity of 12.9±1.3nM, similar to its affinity for G:T mismatch. However, the functional importance and details of this interaction are uncharacterized. We employ ATPase activity assays and equilibrium binding assays to understand the function of Msh2-Msh6’s interaction with Holliday Junction DNA; namely, we conduct experiments with G:T heteroduplex DNA and J3 Holliday Junction to establish control measurements. We follow with experiments conducted with J3 junction DNA that includes a G:T mismatch incorporated at various positions to probe the ATP hydrolysis and binding activity of Msh2-Msh6 in the presence of two favorable binding targets.
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Exploring-the-Binding-Activity-of-Msh2-Msh6-with-Mismatched-Four-Way-Junction-DNA-1-1Live Poster Session:
Thursday, July 29th 1:15-2:30pm EDT