3-D STRUCTURE OF GRANUM-STROMA THYLAKOID ASSEMBLIES IN HIGHER

这个子项目是许多研究子项目中利用 资源由NIH/NCRR资助的中心拨款提供。子项目和 调查员(PI)可能从NIH的另一个来源获得了主要资金， 并因此可以在其他清晰的条目中表示。列出的机构是 该中心不一定是调查人员的机构。 摘要： 颗粒类囊体系统的螺旋排列主要是从单个颗粒或一对颗粒上化学固定的标本的连续切片EM研究中明显看出的。这些并没有揭示与基质类囊体相互连接的基粒的3D排列(网络)。基粒的个体发育、大小的变化、在不同环境条件(高光、弱光、短期适应、长期驯化)下的灵活性、膜的光磷化的结构后果以及物理化学环境(渗透压、离子强度)的作用也很难理解。此外，由于化学固定可能存在伪影，因此存在一些不确定因素。颗粒叶绿体还没有被用电子断层扫描技术研究，可能是因为完整的成熟叶绿体的大小和致密结构。因此，我们还计划尝试对完整的叶绿体(和类囊体膜，即没有包膜和可溶性成分的质体)进行成像，方法是使用带有小颗粒的质体和通过超声波分离和/或用洗涤剂切割的基粒基质组件。 通过大量的其他技术，我们对颗粒-基质组合体中的复合体和片层的组织有了详细的了解，例如揭示了宏域组织(直径300-400 nm)的圆二色谱，以及提供了周期性超结构变化及其光合作用功能的小角X射线和中子散射。人们还证实，这些结构能够通过一种以前未知的生物学物理机制--热光效应--进行光诱导的可逆结构组织。 在上一个报告所述期间，进行了几次断层扫描重建。米尔迪博士和加拉布博士访问了RVBC一周。他们观察并参与了数据收集和断层重建。一张断层照片显示了基粒是如何在类囊体内联系在一起的，他们一直希望澄清这一点。他们发表了一场演讲，展示了理解叶绿体结构/功能所用的各种技术。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. ABSTRACT: The helical arrangement of the granal thylakoidal system is evident mostly from serial section EM studies on chemically fixed specimens on single granum or on a pair of grana. These do not reveal the 3D arrangement (network) of grana that interconnects with stroma thylakoids. The ontogeny of grana, the variation of its size, its flexibility under various environmental conditions (high light  low light, short term adaptation, long-term acclimation), structural consequences of the phosophorylation of membranes, and the role of the physico-chemical environment (osmolality, ionic strength) are also poorly undrestood. Futher, there are some uncertainties because of possible artifacts due to the chemical fixation. Granal chloroplasts have not been studied by electron tomographic techniques  perhaps because of the large size and dense structure of whole, mature chloroplasts. Thus we plan, in addition, to attempt the imaging of intact chloroplasts (and thylakoid membranes, i.e. plastids without their envelope membranes and soluble components), by the use of plastids with small-sized grana, and granum stroma assemblies isolated by sonication and/or cleaved by detergents. We have detailed knowledge concerning the organization of complexes and lamellae in the granum-stroma assemblies by a wealth of other techniques, such as circular dichroism spectroscopy that reveals the macrodomain organization (300-400 nm in diameter) and small angle X-ray and neutron scattering which provide information on the variations in the periodic ultrastructure, as well as their photosynthetic functions. It has also been established that these structures are capable of undergoing light-induced reversible structural organization by a previously unknown physical mechanism in biology, the thermo-optic effect. In the previous reporting period, several tomographic reconstructions were made. Drs. Must¿rdy and Garab visited the RVBC for one week. They observed and participated in data collection and tomographic reconstruction. A tomogram was made showing how the grana are linked within the thylakoid, a point they had been hoping to clarify. They presented a lecture showing a variety of techniques brought to bear on understanding structure/function of the chloroplast.