CAREER: Quantification of Cellular Proteome Stress and Recovery Using Chemical Methods

凭借这一奖项，化学学部的生命过程化学项目资助了宾夕法尼亚州立大学的张欣博士开发新的工具，用于实时检测压力对活细胞中蛋白质的影响。细胞中的蛋白质必须在可溶于水的折叠态（可溶折叠态）和不溶于水的固态（不可溶聚集态）之间保持微妙的平衡。过度聚集对细胞有害，但这种现象发生在细胞暴露于老化、高温和某些化学物质等压力下时。为了了解蛋白质应激与细胞功能的关系，张博士开发了一类新的荧光传感器，当它们与细胞中未折叠的蛋白质接触时就会发光。该项目为实时测定细胞蛋白应力和恢复提供了多种新工具。该计划的跨学科性质允许研究生和本科生获得现代化学生物学研究方面的培训和专业知识。该项目还包括一个名为“寻找蛋白质聚集体探测器”的教育计划。该教育计划的中心主题是将K-12和本科生整合到研究和公众参与项目中，发现新的化学探测器，用于检测未开发的天然染料中的蛋白质聚集体及其与蛋白蛋白的相互作用。由于现有的蛋白质组应激传感器不足以对蛋白质组应激和恢复的动态进行定量研究，张博士正在开发一种新型荧光传感器，以AgHalo传感器为代表。该传感器使用荧光强度的变化作为直接读数来量化应力诱导的蛋白质聚集。在此基础上，张博士正在开发第二种荧光蛋白组应力传感器AgSNAP，它与AgHalo正交，从而实现一组双色传感器。为了检测作为应激后事件的蛋白质重折叠，利用抗溶剂致变色荧光团开发了一种荧光蛋白组恢复传感器。这些传感器用于定量细胞内蛋白质聚集和再折叠的细胞受到慢性应激。突变蛋白可以形成可溶性或不可溶性聚集体也是目标。这些实验的结果验证了突变蛋白的可溶性聚集体比不可溶性聚集体引起更严重的蛋白质组应激的假设。这些研究目标为蛋白质生物化学界提供了一套新的蛋白质组应激和恢复传感器，并提供了长期应激细胞中蛋白质组聚集和重折叠的定量知识。该奖项反映了美国国家科学基金会的法定使命，并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，被认为值得支持。

With this award, the Chemistry of Life Processes Program in the Chemistry Division is funding Dr. Xin Zhang from Pennsylvania State University to develop new tools examining the effects of stresses on proteins in live cells, in real time. The proteins in cells must be maintained in a delicate balance between a folded state that is dissolved in water (soluble folded) and an solid state that does not dissolve in water (insoluble aggregated state). Excessive aggregation is bad for the cells but this phenomenon occurs when cells are exposed to stresses such as aging, heat and certain chemicals. To understand how protein stress relates to cellular function, Dr. Zhang focuses develops a new class of fluorescent sensors that light up when they come into contact with unfolded proteins in cells. This project provides a variety of new tools for real-time determinations of cellular protein stresses and recovery. The interdisciplinary nature of the program allows graduate and undergraduate students to gain training and expertise in modern chemical biology research. The project also includes an education plan called "Seeking Protein Aggregates Detectors". The central theme of this education plan is to integrate K-12 and undergraduate students in research and public engagement programs that discover new chemical detectors for protein aggregates from unexplored natural dyes and their interactions with egg white proteins. As existing proteome stress sensors are insufficient to enable quantitative studies on the dynamics of proteome stress and recovery, Dr. Zhang is developing a novel class of fluorogenic sensors, represented by the AgHalo sensor. This sensor uses changes in fluorescence intensity as a direct readout to quantify stress-induced protein aggregation. Building on this foundation, Dr. Zhang is developing a second fluorogenic proteome stress sensor, AgSNAP, which is orthogonal to AgHalo, thus enabling a set of two-color sensors. To detect protein refolding as a post-stress event, a fluorogenic proteome recovery sensor is being developed using anti-solvatochromic fluorophores. These sensors are used to quantify intracellular protein aggregation and refolding in cells that are subjected to chronic stresses. Mutant proteins that can form either soluble or insoluble aggregates are also targeted. Results of these experiments test the hypothesis that the soluble aggregates of mutant proteins cause a more severe proteome stress than the insoluble aggregates. These research goals enable a set of novel proteome stress and recovery sensors for the protein biochemistry community and provide quantitative knowledge of proteome aggregation and refolding in chronically stressed cells.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.