Laser Trapping-Induced Fibrous Structure Formation of Cytoskeletal Proteins in Deuterium Oxide

• Main Authors: Shih, Yang-Hsin, 施仰欣
• Other Authors: Teruki Sugiyama
• Format: Others
• Language: en_US
• Published: 2019
• Online Access: http://ndltd.ncl.edu.tw/handle/mvvt4h

碩士 === 國立交通大學 === 應用化學系分子科學碩博士班 === 107 === We here present laser trapping-formed microtubules consisting of tubulin protein by focusing a continuous wave laser of 1064 nm at air/solution/glass interface in D2O. It has been already preformed the laser trapping experiments in H2O and the spatiotemporal control of microtubule formation was successfully demonstrated. Laser trapping increases not only local concentration of tubulin but also local temperature elevation at laser focus. Therefore, it has not been cleared that either concentration or temperature caused by laser irradiation mainly contribute to microtubule formation because, for example, 20 µmol/l tubulin forms microtubules at high temperature (30 ºC). In D2O, local temperature increased by laser trapping is suppressed twenty times compared to that in H2O, by which laser trapping effects on microtubule formation becomes clear. A systematic study on microtubule formation was performed under various concentration of tubulin, ranging from 20 to 80 μM. When a focused laser beam was introduced to sample solutions, the condensed area of tubulin was generated at the laser focus and it was extended to the outside, eventually to be approximately 10 μm in diameter. During the irradiation, an anisotropic area, which was confirmed under a cross Nicol condition, was observed around the laser focus. In order to confirm whether or not microtubules are involved in the formed anisotropic area, the temporal change in anisotropic area was observed under a low temperature of 4 ºC, because the microtubules are dissolved under such a low temperature. As a result, at all concentrations the anisotropic area disappeared within 1 hour, and it took more time at high concentrations. We also found that orientation of microtubules might be aligned by laser polarization direction. Based on all on the results obtained here, we propose dynamics and mechanism of laser trapping-induced fibrous structure formation in view of effective laser trapping at air/solution/glass interface. Microtubule consisting of cytoskeleton plays a crucial role in the cell from tubulin dimer, and contribute a great part to maintain the cell shape. Although further investigation should be necessary to discuss the details of the formation mechanism, we believe that the findings in this study will give a new insight into the mechanism of microtubule formation in future.