Structural and Functional Characterization of Cyanoglobin

蛋白质的血红蛋白超家族在所有生物界都有代表性。在脊椎动物中，血红蛋白及其近亲肌红蛋白负责分子氧的运输和储存。这两种高度专业化的蛋白质的三维结构和功能之间的关系已经被广泛地探索和理解。在无脊椎动物中，血红蛋白表现出更广泛的生理作用，反映了珠蛋白折叠进化、氧气利用和血红素反应的基本方面。然而，对于这些蛋白质来说，结构和功能之间的联系还没有完全确定，而且还不可能根据氨基酸序列来预测其化学性质。这种知识的缺乏限制了基因组测序数据的有用性和对进化关系的解释。该项目的目标是研究非固氮蓝藻聚球藻的原始血红蛋白。PCC6803和聚球藻。PCC 7002。两者都是六配位的血红蛋白，利用两个组氨酸来连接铁离子，并且都经历了显然是它们所独有的血红素基团的翻译后连接。将应用生物物理方法(体外和体内核磁共振光谱、光学光谱)和生化方法(定点突变、遗传操作、表型鉴定)来研究聚球藻血红蛋白在体内的作用；确定聚球藻和聚球藻蛋白在经过血红素修饰前后的溶液中的三维结构；并分析决定血红素配位和反应活性的分子决定因素。该项目的研究部分将为研究生和本科生提供培训机会，并通过以蓝藻生理学和生物化学为重点的合作，建立化学系和生物化学及分子生物学系之间多学科互动的机制。该项目还包含三个具体的教育组成部分，将进一步加强其更广泛的影响。第一个是为高级本科生物化学课程开发三维可视化模块，并将其纳入本课程与研究相关的主题。这是为了让材料栩栩如生，并鼓励化学学生追求具有生命科学倾向的职业。第二个是针对研究生开发一门关于沟通技巧的短期课程。重点将放在如何向普通观众传达复杂的科学概念和问题上。三是继续组织和增加跨部门结构生物学研讨会的频率，讲者为高级学生。这些活动预计将为本科生、研究生和博士后在学术界、工业界和政府部门的专业生涯做好准备。

The hemoglobin superfamily of proteins has representatives in all kingdoms of organisms. In vertebrates, hemoglobin and its close relative myoglobin are responsible for the transport and storage of molecular oxygen. The relationship between three-dimensional structure and function in these two highly specialized proteins has been explored extensively and is well understood. In invertebrates, hemoglobins exhibit a broader range of physiological roles reflecting essential aspects of globin fold evolution, oxygen utilization, and heme reactivity. For these proteins, however, the connection between structure and function is incompletely defined and it is not yet possible to anticipate chemical properties on the basis of the amino acid sequences. This lack of knowledge limits the usefulness of genome sequencing data and the interpretation of evolutionary relationships. The goal of the project is to investigate primitive hemoglobins from the non-nitrogen fixing cyanobacteria Synechocystis sp. PCC 6803 and Synechococcus sp. PCC 7002. Both are hexacoordinate hemoglobins utilizing two histidines to ligate the iron ion, and both undergo a facile post-translational attachment of the heme group apparently unique to them. Biophysical methods (NMR spectroscopy in vitro and in vivo, optical spectroscopy) and biochemical methods (site-directed mutagenesis, genetic manipulations, phenotypic characterization) will be applied to investigate the role of Synechococcus hemoglobin in vivo; determine the three-dimensional structure of Synechocystis and Synechococcus proteins in solution before and after they have undergone the heme modification; and analyze the molecular determinants of heme coordination and reactivity. Comparison will be made to other invertebrate hemoglobins to develop an atomic level understanding of chemical and biochemical properties.The research component of the project provides training opportunities for graduate and undergraduate students and, through a collaboration focused on cyanobacterial physiology and biochemistry, establishes a mechanism for multidisciplinary interactions between the Chemistry department and the Biochemistry and Molecular Biology department. The project also contains three specific educational components that will enhance further its broader impact. The first is the development of three-dimensional visualization modules for an advanced undergraduate biological chemistry course and the inclusion in this course of research-related hemoglobin topics. This is designed to bring the material to life and encourage chemistry students to pursue careers with a life sciences slant. The second is the development of a short course in communication skills aimed at graduate students. The emphasis will be placed on how to convey sophisticated scientific concepts and issues to a general audience. The third is to continue the organization and increase the frequency of inter-departmental structural biology seminars, for which the speakers are advanced students. These activities are expected to prepare undergraduate, graduate, and post-doctoral students for professional careers in academia, industry, and government.