Characterize FliL and Stator Interaction in Bacterial Flagellar Motor System by Fluorescent Microscopy
Tsai-Shun Lin1*, Shi-Wei Zhu2, Chien-Jung Lo1
1Department of Physics, National Central University, Taoyuan, Taiwan
2Division of Biological Science, Nagoya University, Nagoya, Japan
* presenting author:Tsai-Shun Lin, email:103282003@cc.ncu.edu.tw
Flagellar motor is essential for bacteria motility to search various environments striving for survival. In a flagellar motor, stator units are the torque-generation units involving in energy transferring process to drive motor rotation. Recent studies pointed out membrane protein FliL might support the torque generation, especially in the high load state. However, how FliL proteins interact with stator units in the flagellar motor is still unknown. We aim to observe the stator protein behavior directly under control of FliL proteins by fluorescent microscopy. Through genetic approach, GFP was fused to stator protein or FliL protein. Under control of FliL protein, Fluorescence recovery after photobleaching (FRAP) experiment showed the turnover rate of stator units were similar suggesting the dynamics of stator and FliL might be independent. In addition, we applied total internal reflection fluorescence (TIRF) microscopy in stepwise photobleaching method to measure single GFP intensity and calculate the number of stator units or FliL assemble in the flagella motor. We compared the number of assembled stator units in absence of FliL. Also from the number of FliL around a motor, we estimated the stoichiometry of FliL and stator units assembled in the flagellar motor system may close to 2:1. We propose a new model of FliL and stator unit interaction.


Keywords: FliL, Bacterial flagellar motor, FRAP, TIRF