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

处于食物网底层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使用的形式。 该项目旨在研究一组广泛分布的浮游植物聚球藻如何通过将其色素成分与光环境相匹配来成功进行光合作用。这种现象被称为色彩适应，涉及到一些基因，这些基因允许这些微生物根据海洋过滤的光的可用颜色来整合不同的色素。 该团队将表征这些基因编码的蛋白质的结构和功能。这项研究的结果将有助于预测海洋聚球藻和食物链如何受到海洋变化的影响。这些研究有望扩大生物技术的荧光生物成像探针的库。 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培训。推广活动包括提高STEM专业的持久性和多样性的夏令营，高中教师培训模块，以及提高公众的科学意识。海洋聚球藻通过翻译后修饰构成光合天线的藻胆蛋白（称为藻胆体）实现广泛的色素多样性。藻胆蛋白的生物合成涉及通过胆色素裂解酶催化的酶促反应将化学上不同的胆色素附着到这些蛋白质上。胆色素裂解酶-异构酶将吸收绿色的胆色素连接并将其异构化为吸收蓝色的胆色素。 这些酶在IV型色适应（CA 4）中起关键作用，其中细胞在蓝光或绿色光下由于其藻胆体的大量重组而改变颜色。进行CA4的能力由4 - 5个基因组成的小基因组岛赋予，包括胆色素裂解酶。然而，聚球藻利用不同的胆色素裂解酶和裂解酶异构酶来实现其色素多样性的机制仍然是难以捉摸的。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相关的未鉴定的胆色素裂解酶进行了研究。该项目的目标是：（2）确定Unk10的功能作用，Unk10是一种在CA 4基因组岛中编码的新蛋白，被假设影响CA 4中涉及的那些胆色素位点的发色团化;（3）将联合收割机晶体学和诱变相结合，建立区分裂合酶和裂解酶的反应机理和结构基础-该奖项反映了NSF的法定使命，并被认为是值得通过使用基金会的知识价值和更广泛的影响审查标准进行评估的支持。

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.