Collaborative Research: Site-Directed Mutagenesis of the Manganese Stabilizing Protein of Photosynthesis

导致光合作用水氧化的分子事件在很大程度上仍不清楚，特别是所涉及的蛋白质和形成锰的配体的特定残基，锰是氧气释放的重要辅助因子。目前的模型表明，参与反应中心的D1D2复合体和一个外在的33kD蛋白，称为锰稳定蛋白。这项研究将结合定点突变技术，使用易转化的蓝藻聚球藻6803，对含有所谓的锰稳定多肽的选定氨基酸残基进行修饰，结合EPR和Endor光谱方法，这些方法能够直接评估连接到反应中心的锰位和其他内源电子给体的结构和动力学性质的这些变化，如Cytb559和Tyr-160.光合作用是地球表面最普遍的生物反应。它产生空气中几乎所有的氧气，而空气中的氧气对所有有氧生物的氧化新陈代谢和化石燃料驱动的发动机的燃烧都是必不可少的。因此，光合作用的氧气释放为大多数生物和商业机器提供动力。目前，还没有一种人造催化剂能够实现从水中高效地放出氧气，也没有一种具有商业用途的能源。另一方面，大自然已经开发出一种独特的酶，这种酶存在于所有放氧植物和藻类中，最适合催化这一过程。这项研究将有助于理解绿色植物能够利用自然界最强大的氧化剂的分子基础，即含锰酶分解水分。通过了解这一生物过程，我们希望获得洞察力，从而为合成水氧化催化剂的设计绘制新的蓝图。

The molecular events leading to photosynthetic water oxidation are still largely unknown, particularly the proteins involved and the specific residues that form ligands to manganese, an essential cofactor for oxygen evolution. Current models suggest the participation of the reaction center D1D2 complex and an extrinsic 33kD protein, called the manganese stabilizing protein. This research will combine the techniques of site-directed mutagenesis, using the readily transformable cyanobacterium Synechocystis 6803, to modify selected amino acid residues comprising the so-called manganese stabilizing polypeptide, together with EPR and ENDOR spectroscopic methods which enable a direct assessment of these changes on the structure and kinetic properties of the manganese site and other endogenous electron donors coupled to the reaction center, such as Cytb559 and Tyr- 160.%%% Photosynthesis is the most pervasive biological reaction on the face of the earth. It produces nearly all of the oxygen in the air which is essential for oxidative metabolism in all aerobic organisms and for combustion in fossil fuel powered engines. Thus, photosynthetic O2 evolution powers most biological and commercial machines. Presently there are no man-made catalysts capable of carrying out the efficient evolution of O2 from water and an energy source with a commercially useful yield. Nature, on the other hand, has developed a unique enzyme, found in all O2-evolving plants and algae, optimized for catalyzing this process. This research will contribute to the understanding of the molecular basis by which green plants are capable of harnessing nature's most power oxidant, the splitting of the water by a manganese-containing enzyme. By understanding this biological process we hope to gain insight that will lead to a new blueprint for the design of synthetic water oxidizing catalysts.*** //