RUI: Activity-based Mapping of Cellular Cathepsins B and L - An introduction to Chemical Biology in Undergraduate Curriculum

RUI：基于活性的细胞组织蛋白酶 B 和 L 作图 - 本科课程中的化学生物学简介

• 批准号: 1709711
• 负责人: Sanjai Pathak
• 金额: $ 28.25万
• 依托单位: CUNY Queens College
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-08-01 至 2021-07-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=1709711&HistoricalAwards=false
• 关键词: RUI; Activity; based; Mapping; Cellular

With this RUI award, the Chemistry of Life Processes Program in the Division of Chemistry is funding Professor Sanjai Kumar to investigate the function of two important enzymes found in human cells. A cell is a complex yet highly organized system where thousands of events are coordinated with great precision at any given moment. These processes are elegantly orchestrated by proteins, the main centers of activity of the cell. It is estimated that the total number of proteins in a human cell is between 250,000 to one million. Assigning individual functions to the various proteins is a challenging endeavor, especially since a protein may be active or inactive depending on the status of the cell. This status of a protein is often controlled by a group of enzymes known as proteases. The human genome has the capacity to produce about 550 proteases. How, why, and where these proteases function in the complex cellular environment remains poorly understood. This project involves examining two closely-related proteases of the cysteine cathepsin family of enzymes; cathepsin B and L. Selective probes are made to target these cathepsins so that their function can be understood. This research endeavor provides educational opportunities with hands-on research experience to graduate and undergraduate students. The laboratory environment is structured for an enhanced interdisciplinary experience for both undergraduate and graduate students. Additionally, there are outreach activities incorporating research training for students from Queensborough Community College and Bard High School Early College in New York. Special emphasis is given to include students from underrepresented groups in science disciplines, women, and veterans. Human cysteine cathepsins comprise a small group of eleven enzymes that are important in maintaining cell homeostasis, growth, differentiation, and survival. One of the main challenges of cathepsin biology is in assigning specific function to individual enzymes in a given cellular context. This is mainly because several functional redundancies and overlapping substrate specificities have been shown to exist in individual cell types. Since newly synthesized cellular cathepsins are subject to post-translational modifications, thereby leading to alteration of their activities (and hence function), specific investigative tools are needed for activity-based interrogation of cellular function in live cells. This project is developing selective tagless activity-based probes (TABPs) of two closely related cysteine cathepsins; cathepsin L and B. The developed TABPs are used in differential and quantitative mapping of cathepsin B and L activities in live pancreatic and neuronal cells. The knowledge gained from these studies significantly advances the functional understanding of cellular processes involving human cathepsin B and L.

有了这个RUI奖，化学系的生命过程化学计划正在资助Sanjai Kumar教授研究人类细胞中发现的两种重要酶的功能。细胞是一个复杂但高度组织化的系统，在任何给定的时刻，成千上万的事件都以非常精确的方式协调。这些过程是由蛋白质，细胞的主要活动中心精心策划的。据估计，人类细胞中的蛋白质总数在25万到100万之间。将个体功能与各种蛋白质结合是一项具有挑战性的奋进，特别是因为蛋白质可能根据细胞的状态而具有活性或无活性。蛋白质的这种状态通常由一组称为蛋白酶的酶控制。人类基因组能够产生大约550种蛋白酶。这些蛋白酶在复杂的细胞环境中如何，为什么以及在哪里发挥作用仍然知之甚少。 本计画包括研究半胱氨酸组织蛋白酶家族中两种密切相关的蛋白酶：组织蛋白酶B和L。选择性探针是针对这些组织蛋白酶，使他们的功能可以理解。 这项研究奋进为研究生和本科生提供了实践研究经验的教育机会。实验室环境的结构为增强本科生和研究生的跨学科经验。此外，还为来自皇后区社区学院和纽约巴德高中早期学院的学生开展了包括研究培训在内的外联活动。特别强调包括科学学科，妇女和退伍军人代表性不足的群体的学生。人半胱氨酸组织蛋白酶包括一小组的11种酶，这些酶在维持细胞稳态、生长、分化和存活中是重要的。组织蛋白酶生物学的主要挑战之一是在给定的细胞环境中将特定功能分配给单个酶。这主要是因为已经证明在单个细胞类型中存在几种功能冗余和重叠底物特异性。由于新合成的细胞组织蛋白酶受到翻译后修饰，从而导致其活动（因此功能）的改变，需要特定的研究工具，以活性为基础的讯问活细胞中的细胞功能。该项目正在开发两种密切相关的半胱氨酸组织蛋白酶（组织蛋白酶L和B）的选择性无标签活性探针（TABP）。开发的TABP用于活胰腺和神经元细胞中组织蛋白酶B和L活性的差异和定量映射。从这些研究中获得的知识显著地推进了对涉及人组织蛋白酶B和L的细胞过程的功能理解。

项目成果

Cell penetrable, clickable and tagless activity-based probe of human cathepsin L

• DOI: 10.1016/j.bioorg.2019.02.032
• 发表时间: 2019-04-01
• 期刊: BIOORGANIC CHEMISTRY
• 影响因子: 5.1
• 作者: Dana, Dibyendu;Garcia, Jeremy;Pathak, Sanjai K.
• 通讯作者: Pathak, Sanjai K.

Highly Efficient Cell-Penetrating Probes of Protein Arginine Deiminases for Functional Proteomics

用于功能蛋白质组学的蛋白质精氨酸脱亚胺酶的高效细胞穿透探针

• DOI: 10.1002/cbic.201800257
• 发表时间: 2018
• 期刊: ChemBioChem
• 影响因子: 3.2
• 作者: Kumar, Sanjai