| Summary: | 碩士 === 國立臺灣大學 === 生物化學暨分子生物學研究所 === 104 === Due to inappropriate use of antibiotics, the emergence of antimicrobial-resistant bacteria, including methicillin-resistant Staphylococcus aureus (MRSA), Pseudomonas aeruginosa, Streptococcus pneumonia, has been a critical problem nowadays. In particular, infections of MRSA, resistant to most antibiotics, often cause a higher mortality in patient. In our previous study, two celecoxib derivatives, Cpd36 and Cpd46, were identified to exhibit inhibitory activity against MRSA. Subsequent mutation analysis of S. aureus resistant isolates by next-generation sequencing identified a membrane protein translocase, YidC2, as the potential target of these compounds. Here, we aim to investigate the interaction between the YidC2 and Cpd36 and Cpd46. The results showed that ectopic expression of YidC2 can decrease the susceptibility of S. aureus to Cpd46. The level of ATP synthase subunit c (ATPsc) on bacterial membrane were also reduced by Cpd46 treatment, leading to a reduction of ATP level in bacteria. Although YidC2 protein level in bacteria was increased with exposure to Cpd46, the raised YidC2 level didn’t reverse the reduction of ATPsc translocation. To further investigate whether Cpd46 bind to YidC2 directly, recombinant YidC2 was prepared by overexpressing protein in E. coli followed by extraction with detergent. Next, to explore the interaction between YidC2 and compounds, biophysical approaches were used to monitor the change of several important physiochemical parameters, including enthalpy, entropy, free energy and the thermodynamic profile. However, no significant biophysical evidences showed these compounds binding to YidC2 directly. Altogether, our results showed that YidC2 plays an important role in the antibacterial activity of Cpd46, and YidC2 is a potential target for new anti-MRSA drugs development.
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