ROTATIONAL DIFFUSION WITH 2 PHOTON FCS

我们正在使用单光子共聚焦和双光子荧光 光谱学检测单分子水平上的蛋白质反应。 通过测量单个蛋白质的过程，我们不会受到 系综求平均值。因此，蛋白质的异质性程度， 并且可以监控可用的各种反应路径 直接去吧。初步的实验是关于结合的 8-苯胺基-1-萘磺酸(ANS)合成脱辅酶原球蛋白。这个 ANS的量子产额增加了两个数量级以上 配体与蛋白质结合，使区分蛋白质成为可能 在蛋白质的连接构象和非连接构象之间。这个 蛋白质用俄勒冈州绿色单独标记，使它们更容易 定位并固定在硅胶中。鼻息肉通过 玻璃，以及随时间监测的结合事件。利率 然后可以从时间中提取反应系数 绑定事件的序列。

We are using one-photon confocal and two-photon fluorescence spectroscopy to detect protein reactions on the single molecule level. By measuring processes on single proteins, we are not subject to ensemble averaging. Thus, the degree of heterogeneity in proteins, and the variety of reaction pathways available can be monitored directly. Initial experiments are performed on the binding of 8-anilino-1 -naphthalene sulfonic acid (ANS) to apornyoglobin. The quantum yield of ANS increases by over 2 orders of magnitude when the ligand is bound to the protein, making it possible to distinguish between the ligated and unligated conformations of the protein. The proteins are singly labeled with oregon green to make them easy to locate and are immobilized in a silica gel. The ANS diffuses through the glass, and the binding events monitored with time. The rate coefficients of the reaction can then be extracted from the time sequence of binding events.