PROTEIN METHYLATION IN PHOTOTAXIS AND CHEMOTAXIS

我们将研究蛋白质甲基化与受体的关系 光感和化学感信号转导中的兴奋 Halobacterium halobium盐生菌的代谢途径。 一项主要的研究发现， 细菌的趋化性是细胞膜的可逆甲基化， “信号转导”蛋白在适应中起着关键作用， 趋化性刺激 最近，一种趋光性系统使用视紫红质样 光感受器（慢循环细菌视紫红质，“SR”）已经被证明是 控制H.盐生植物 类似于细菌的趋化性。 sR蛋白经历两个 不同的光化学反应周期，取决于激发 波长 一个周期产生吸引信号，另一个周期产生吸引信号。 产生排斥信号 这些信号的产生可以是 通过控制激发的光谱组成来控制 光 几个实验室的初步研究表明， 甲基化反应参与sR介导的趋光性， 在这些细胞中的趋化性行为。 我们的目标是：（1） 描述了H. 盐生菌趋光性;（2）探讨它们与光激发的关系。 两个sR光化学反应循环激活甲基化 系统 蛋白质甲基化将通过用（3 H）甲基进行体内标记来监测， 通过聚丙烯酰胺凝胶电泳和放射性标记进行蛋白质分离 通过自动荧光照相术和闪烁计数进行检测。 在同一个在 体内制剂，其中光刺激诱导的甲基化是 监测，sR的光化学反应将通过 时间分辨闪光光解 我们的理由是， 详细的受体信息和蛋白质甲基化数据将阐明 受体兴奋与甲基化/去甲基化的关系 反应. 利用光来控制sR信号的能力将使我们能够 探索趋光性甲基化系统对 具有时间分辨率的刺激和难以获得的控制， 通过化学梯度刺激趋化系统。

We will study the relationship of protein methylation to receptor excitation in the photosensory and chemosensory signal transduction pathways of Halobacterium halobium. A major finding from studies of bacterial chemotaxis is that reversible methylation of integral membrane "signal transducer" proteins plays a critical role in adapatation to chemotaxis stimuli. Recently a phototaxis system using a rhodopsin-like photoreceptor (the slow-cycling bacterial rhodopsin, "SR") has been shown to govern excitation and adaptation motility responses in H. halobium analogous to those of bacterial chemotaxis. The sR protein undergoes two different photochemical reaction cycles, depending on excitation wavelength. One cycle generates an attractant signal, and the other generates a repellent signal. The generation of these signals can be controlled by controlling the spectral composition of the excitation light. Initial work from several laboratories indicates protein methylation reactions participate in the sR-mediated phototaxis as well as in chemotaxis behavior in these cells. Our objectives are: (1) to characterize the components of the protein methylation system in H. halobium phototaxis; (2) to assess their relationship of photoexcitation of the two sR photochemical reaction cycles to activation of the methylation system. Protein methylation will be monitored by in vivo labeling with (3H)methyl, protein separation by polyacrylamide gel electrophoresis, and radiolabel detection by autofluorgraphy and scintillation counting. In the same in vivo preparations in which the photostimulus-induced methylation is monitored, the photochemical reactions of sR will be analyzed by time-resolved flash photolysis. Our rationale is tht this combination of detailed receptor information and protein methylation data will clarify the relationship of receptor excitation to the methylation/demethylation reactions. The ability to use light to control sR signaling will permit us to probe some properties of the phototaxis methylation system's response to stimuli with a time resolution and control difficult to obtain with stimulation of chemotaxis systems by chemical gradients.