Functional characterisation of different fucoxanthin chlorophyll proteins (FCP) in diatoms

本项目旨在了解硅藻天线复合物、岩藻黄素-叶绿素蛋白（FCP）的激发能传输、FCP可能的保护功能以及FCP蛋白在光系统周围的大分子组织。尽管在基因水平上可以区分出几组fcp，但通常只有一两个主要的天线复合物被纯化，而且不同fcp的生化和光谱数据很少。因此，对这些不同多肽可能的功能异质性知之甚少。为了表征单天线蛋白，我们希望通过添加标签修饰不同FCP基因的硅藻，以便能够在所有相关蛋白的背景下分离特定的FCP。此外，假定的色素结合位点将发生突变，以获得具有不同色素沉着的fcp。然后，基因工程的fcp应该允许进行详细的生化和光谱表征。根据其在不同fcp中的定位，将特别强调硅黄素在荧光猝灭中的作用。对突变体FCP的研究将伴随着对不同光系统下不同FCP蛋白排列的研究。此外，使用his标签在电子显微镜下标记fcp将允许在光系统的超配合物中明确定位fcp。

The aim of this project is to understand the excitation energy transport in the antenna complexes of diatoms, fucoxanthin-chlorophyll proteins (FCPs), the possible protective function of FCPs and the macromolecular organisation of the FCP proteins around the photosystems. Despite the fact that several groups of FCPs can be distinguished on the level of genes, usually only one or two main antenna complexes are purified and biochemical and spectroscopic data on distinct FCPs are scarce. Thus, little is known about possible functional heterogeneities of these different polypeptides. In order to characterise single antenna proteins we want to transform diatoms with different FCP genes modified by addition of tags in order to be able to separate specific FCPs against the background of all related proteins. Additionally, putative pigment binding sites will be mutated to obtain FCPs with different pigmentation. The genetically engineered FCPs should then allow for a detailed biochemical and spectroscopic characterisation. Special emphasis will be on the role of diatoxanthin in fluorescence quenching depending on its localisation in the different FCPs. The studies on mutant FCPs will be accompanied by studies on the arrangement of the different FCP proteins with respect to the different photosystems. Additionally, using the His-tags to label FCPs for electron microscopy will allow to specifically locate the FCPs inside supercomplexes of the photosystems.