Chemical Toolbox for Deciphering the Molecular Origins of Selectivity in Human Extracellular Sulfatases

用于破译人细胞外硫酸酯酶选择性分子起源的化学工具箱

• 批准号: 2004243
• 负责人: Kamil Godula
• 金额: $ 45万
• 依托单位: University of California-San Diego
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-07-01 至 2023-06-30
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=2004243&HistoricalAwards=false
• 关键词: Chemical; Toolbox; Deciphering; Molecular; Origins

With this award, the Chemistry of Life Processes Program in the Chemistry Division is funding the research of Dr. Kamil Godula at the University of California – San Diego, to study the effects of two human enzymes known as Sulf1 and Sulf2. These enzymes act on the exterior of cell surfaces by recognizing and modifying the patterns of negatively charged groups on sugar polymers that are part of the outside coating of cells. In doing so they control the activity of proteins involved in critical cell signaling and development events. Despite being a key element in regulating important cellular functions, the structures of these enzymes and the sugars they act on are still poorly understood. Additionally, their precise actions in a cellular context need better understanding. This research project develops a set of new tools and techniques at the interface of chemistry and biology to fill the gaps in the basic understanding of the biological functions of these enzymes. These activities allow graduate students to obtain training in the interdisciplinary area of chemical biology focusing on carbohydrate chemistry and biology. The students also gain a deeper understanding of the varied roles of carbohydrates in biology. The educational and technical resources enabled by this award allow for the dissemination of the knowledge, tools, and techniques stemming from this research project to the broader research community and the general public. This research develops novel methods to produce human sulfatase enzymes enabling detailed investigations of structure and mechanism. New chemical glycomics tools are envisioned to capture and structurally characterize the polysaccharide substrates for these enzymes. Further, biochemical and cell-based assays are under development to investigate their function during embryonic stem cell differentiation. The sulfatase enzymes are thought to have specificity for unique negatively charged regions of their sulfated polysaccharide substrates complementing the organization of positively charged residues in their substrate-binding domains. However, little information exists regarding the structure of the enzymes and the composition of their preferred substrates. Arrays comprising structurally defined sulfated polysaccharides from synthetic and biological sources are in use to identify preferred substrates for the sulfatase enzyme isoforms and to delineate how the enzymatic removal of sulfates from the polysaccharides impacts their interactions with growth factors. The chemical toolbox includes substrate-based fluorogenic probes for imaging sulfatase activity in biological environments and reactive molecules for chemical fingerprinting of substrates for structural characterization by mass spectrometry analysis. In combination, these tools are deployed to map how sulfatase enzymes control growth factor signaling during neural and mesenchymal differentiation of murine embryonic stem cells. The tools are general and suitable for the investigation of other biological systems where extracellular sulfatases exert important regulatory 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.

凭借该奖项，化学部的生命过程化学项目正在资助加州大学圣地亚哥分校的 Kamil Godula 博士的研究，以研究两种人类酶 Sulf1 和 Sulf2 的作用。这些酶通过识别和修改糖聚合物上带负电基团的模式来作用于细胞表面的外部，糖聚合物是细胞外涂层的一部分。在此过程中，它们控制参与关键细胞信号传导和发育事件的蛋白质的活性。尽管这些酶是调节重要细胞功能的关键元素，但人们对这些酶及其作用的糖的结构仍然知之甚少。 此外，它们在细胞环境中的精确作用需要更好的理解。 该研究项目开发了一套化学和生物学交叉领域的新工具和技术，以填补对这些酶的生物学功能的基本理解的空白。这些活动使研究生能够获得化学生物学跨学科领域的培训，重点是碳水化合物化学和生物学。 学生们还对碳水化合物在生物学中的不同作用有了更深入的了解。 该奖项提供的教育和技术资源可以将本研究项目产生的知识、工具和技术传播给更广泛的研究界和公众。这项研究开发了生产人类硫酸酯酶的新方法，能够详细研究结构和机制。预计新的化学糖组学工具可以捕获这些酶的多糖底物并对其进行结构表征。此外，生物化学和基于细胞的测定正在开发中，以研究它们在胚胎干细胞分化过程中的功能。硫酸酯酶被认为对其硫酸化多糖底物的独特带负电区域具有特异性，以补充其底物结合域中带正电残基的组织。然而，关于酶的结构及其优选底物的组成的信息很少。包含来自合成和生物来源的结构确定的硫酸化多糖的阵列用于鉴定硫酸酯酶异构体的优选底物，并描述从多糖中酶促去除硫酸盐如何影响它们与生长因子的相互作用。化学工具箱包括用于生物环境中硫酸酯酶活性成像的基于底物的荧光探针和用于底物化学指纹识别的反应分子，以便通过质谱分析进行结构表征。结合起来，这些工具被用来绘制硫酸酯酶如何在小鼠胚胎干细胞的神经和间充质分化过程中控制生长因子信号传导。这些工具是通用的，适用于细胞外硫酸酯酶发挥重要调节功能的其他生物系统的研究。该奖项反映了 NSF 的法定使命，并通过使用基金会的智力价值和更广泛的影响审查标准进行评估，被认为值得支持。

项目成果

Biologically Derived Neoproteoglycans for Profiling Protein–Glycosaminoglycan Interactions

用于分析蛋白质与糖胺聚糖相互作用的生物衍生新蛋白聚糖

• DOI: 10.1021/acschembio.2c00205
• 发表时间: 2022
• 期刊: ACS Chemical Biology
• 影响因子: 4
• 作者: Porell, Ryan N.;Follmar, Julianna L.;Purcell, Sean C.;Timm, Bryce;Laubach, Logan K.;Kozirovskiy, David;Thacker, Bryan E.;Glass, Charles A.;Gordts, Philip L.;Godula, Kamil
• 通讯作者: Godula, Kamil