讲座主题： “Seeing” the movements of Protein Molecules with Single-molecule Spectroscopy
讲座时间： 2013年10月8日（周一） 下午3点
讲座地点： 教三301
Abstract: The advent of single-molecule fluorescence techniques has provided a new perspective for the study of biomolecules. Utilizing single-molecule techniques, we are now able to study protein conformational dynamics and protein-protein interaction networks directly. For complex systems like proteins, energy landscape is an effective way of correlating function with structure and free energies. We use single-molecule optical microscopy and enzymatic assays to measure the energy landscape of a model enzyme system, adenylate kinase (AK) from Escherichia coli. AK’s lid domain undergoes a large conformational change at the catalytic, millisecond timescale. Using the high-resolution time-dependent single-molecule FRET (Förster Resonance Energy Transfer), we have measured AK||s lid movements on the millisecond scale and map out its entire conformational distribution along the FRET coordinate without a presumed model. Using this information, we have quantitatively recovered AK||s energetic landscape and related its stochastic lid dynamics to its catalytic function. To study protein-protein interactions during signal transduction events, we have chosen brassinosteroids signaling pathway in plants as our model system. Brassinosteroids (BRs) are the sixth class of plant hormones that involved in numerous plant development processes such as leaf expansion, shoot elongation and pollen tube formation. Once the signal transduction is initiated by the membrane receptor kinase BRI1 (brassinosteroid insensitive 1), the signal transmits from the cytoplasm to the nucleus. In the cytoplasm, the signaling pathway is realized by three proteins: BIN2 (brassinosteroid insensitive 2), BES1 (BRI1 ems suppressor1) and a kind of 14-3-3s protein . BRs signaling pathway have been extensively studied via genetics, proteomics, genomics and cell biology techniques. However, these bulk methods can’t follow the transduction process in situ or resolve molecular details at a rate matching the true signaling time-scale. Here we use a single molecular assay based on Total-Internally Reflected Fluorescence (TIRF) microscopy to observe the interaction of these three proteins. The result shows that BIN2 can phosphorylate BES1 on the order of seconds, and the dimeric 14-3-3s can only bind with BES1 in its phosphorylated form. In addition, we have, for the first time, found that the interaction between BIN2 and BES1 is oxygen dependent. This result may have implications on BRs signaling pathway’s involvement of stress acclimation in plants.
谭砚文教授，台湾台北人。1998年自台湾大学物理系毕业后，于中央研究院原子分子研究所范文祥实验室从事发光共轭高分子研究并于2000年取得台湾大学物理硕士学位。同年至美国哥伦比亚大学Horst Stormer实验室攻读博士学位。2006年底博士毕业后，加入杨皓实验室先后在加州伯克力大学与普林斯顿化学系，以单光子荧光光谱学的方法，研究蛋白质分子构像动力相关的生物物理问题。2010年加入复旦大学物理系。
研究方向：①蛋白质与生物的生理时钟；②利用生物信息学的方法来解决蛋白质工程的难题；③发展能够快速控温的单分子荧光共焦显微系统。利用此种新技术来达成分子层面的随机共振；④发展与改良现有的单分子显微技术。
