Investigate How Mismatch Repair Protein MutS Inhibits RecA-Catalyzed Homeologous Strand-Exchange Reaction Using a Single-Molecule Method

• Main Authors: Hung-Chi Liu, 劉泓其
• Other Authors: Hung-Wen Li
• Format: Others
• Language: zh-TW
• Published: 2015
• Online Access: http://ndltd.ncl.edu.tw/handle/09716667923834719471

碩士 === 國立臺灣大學 === 化學研究所 === 104 === Homologous recombination (HR) plays an important role in meiosis and DNA damage repair. In prokaryotes, RecA nucleoprotein filaments search complementary DNA sequence, and carry out the strand-exchange reaction of essential steps in HR. Moreover, RecA can catalyze strand-exchange reaction even if DNA sequence not totally homologous, i.e. homeologous. Previous studies indicate that mismatch repair protein, MutS, inhibits RecA-mediated homeologous recombination. However, the inhibition mechanism remains unknown. We found the DNA mismatch density plays an important role in inhibiting the homeologous strand-exchange reaction. Using gel-shift electrophoresis assay, an intermediate state of strand exchange reaction with higher molecular weight appeared in the presence of MutS. This suggests that MutS proteins promote the accumulation of the reaction intermediate, thus reduce final reaction products. Using single-molecule tethered particle motion experiments, we measured interaction between RecA filaments and homeologous DNA substrates in the presence of MutS. These experiments offer information about how MutS inhibition might take place.