Collaborative Research: Biochemical, genetic and structural studies of bilin lyases

位于食物网底部的海洋浮游植物在光合作用过程中从大气中吸收二氧化碳，并将其转化为海洋中所有生命都可以利用的形式。该项目旨在研究一组广泛存在的浮游植物聚球藻是如何通过将它们的色素成分与光环境相匹配来成功进行光合作用的。这种现象被称为显色适应，涉及到一些基因，这些基因允许这些微生物根据透过海洋的可用光线的颜色来吸收不同的色素。该团队将描述由这些基因编码的蛋白质的结构和功能。这项研究的发现将有助于预测海洋聚球菌和食物链如何受到海洋变化的影响。这些研究有望扩大用于生物技术的荧光生物成像探针库。本研究将对4名研究生、6-8名本科生、6名高中教师和3-6名高中生进行遗传学、生物化学和结构生物学的培训。外展活动包括将提高STEM专业学生的持久性和多样性的夏令营，高中教师培训单元，以及促进公众的科学意识。海洋聚球藻通过对组成称为藻胆体的光合作用触角的藻胆蛋白进行翻译后修饰来实现广泛的色素多样性。藻胆蛋白的生物发生涉及通过胆碱裂解酶催化的酶反应，将化学上不同的胆碱色素附着到这些蛋白上。胆碱裂解酶异构酶将绿色吸收的胆碱与蓝色吸收的胆碱进行异构化。这些酶在IV型染色驯化(CA4)中起着关键作用，在CA4中，细胞由于在蓝光或绿光下藻胆体的大规模重组而改变颜色。执行CA4的能力是由一个由4-5个基因组成的小基因组岛所赋予的，其中包括一个胆碱裂解酶。然而，聚球藻利用不同的胆碱裂解酶和裂解酶异构酶来实现其色素多样性的机制仍然不清楚。本研究结合生物化学、分子遗传学和结构生物学对一组与CA4相关的未知胆碱裂解酶进行了研究。该项目的目标是：(1)识别和表征新的裂解酶和裂解酶异构酶，这些裂解酶和裂解酶异构酶对CA4期间发生的胆碱含量的关键变化负责；(2)确定Unk10的功能作用，Unk10是一种编码在CA4基因组岛上的新蛋白质，假设它会影响CA4中涉及的胆林部位的着色素化；(3)结合结晶学和突变建立区分裂解酶和裂解酶异构酶功能的反应机制和结构基础。该奖项反映了NSF的法定使命，并通过使用基金会的智力优势和更广泛的审查标准进行评估，被认为值得支持。

Marine phytoplankton at the base of the food web take carbon dioxide from the atmosphere during photosynthesis and turn it into a form that all life in the ocean can use. This project aims to study how a group of widespread phytoplankton called Synechococcus succeed in photosynthesis by matching their pigment composition to the light environment. This phenomenon, known as chromatic acclimation, involves a few genes that allow these microbes to incorporate different pigments depending on the available color of light filtering through the ocean. The team will characterize the structures and functions of the proteins encoded by these genes. Findings of this research will help predict how marine Synechococcus and the food chain are affected by changes in the ocean. These studies promise to expand the repertoire of fluorescent bio-imaging probes for biotechnology. This research will train 4 graduate students, 6-8 undergraduate students, 6 high school teachers, and 3-6 high school students in genetics, biochemistry and structural biology. Outreach activities include camps which will increase the persistence and diversity of STEM majors, high school teacher training modules, and promotion of the public’s awareness of science.Marine Synechococcus achieve extensive pigment diversity via post-translational modifications of phycobiliproteins that constitute the photosynthetic antennae called phycobilisomes. Phycobiliprotein biogenesis involves attachment of chemically distinct bilin pigments to these proteins via enzymatic reactions catalyzed by bilin lyases. Bilin lyase-isomerases attach a green-absorbing bilin and isomerize it to a blue-absorbing bilin. These enzymes play a key role in Type IV Chromatic Acclimation (CA4), in which the cells change color as a result of massive restructuring of their phycobilisomes under blue or green light. The ability to perform CA4 is conferred by a small genomic island composed of 4-5 genes, including a bilin lyase. However, the mechanism by which Synechococcus utilizes different bilin lyases and lyase-isomerases to achieve their pigment diversity remains elusive. This research integrates biochemistry, molecular genetics and structural biology to study a group of uncharacterized bilin lyases involved in CA4. The project aims are: (1) to identify and characterize novel lyases and lyase-isomerases responsible for key changes in bilin content that occur during CA4; (2) to determine the functional role of Unk10, a novel protein encoded in the CA4 genomic island, which is hypothesized to influence chromophorylation at those bilin sites involved in CA4; (3) to combine crystallography and mutagenesis to establish the reaction mechanism and structural basis that distinguishes between lyase and lyase-isomerase functions.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.