CAREER:Yeast-based disulfide trapping for engineering selective inhibitors of a protein kinase

这项由生物技术、生化和生物质工程计划颁发的NSF奖支持Park教授的实验室设计和表征Erk-2激酶的新型多肽抑制剂的工作。ERK-2在细胞发育、增殖和恶性肿瘤中发挥重要作用，是细胞内磷酸化依赖信号转导的重要模型系统。考虑到哺乳动物基因组编码了大量参与磷酸化调节的酶，能够特异性地靶向选择的激酶活性应该会导致更好的细胞内沟通模型，以及更有效地治疗由过度激活的激酶引起的遗传性疾病。该提案试图通过结合酵母展示和二硫化物捕获来设计高亲和力、高特异性的Erk-2多肽/蛋白抑制剂。设计的抑制剂将提供一个超越小分子药物的激酶调节的正交化解决方案，并显著增加用于调节激酶通路的候选分子的潜在池。所设计的多肽抑制剂有望具有比小分子抑制剂更有利的生物物理性质，包括：1)相互作用具有更高的特异性，以减少不必要的副作用；2)对靶酶的偶然突变具有健壮性，以最大限度地减少耐药性的发生。建议的二硫键捕获策略已被证明可以使在同一酵母细胞中共表达的相互作用的蛋白质发生交联，与传统的筛选方法相比，该策略提供了一个独特的优势，因为相互作用的特异性本质上是建立在筛选中的，因此所识别的克隆更有可能是功能相关的。因此，该协议的成功实施将与其他特定蛋白质相互作用的工程研究高度相关。朴教授积极参与了密歇根大学旨在鼓励和留住理工科少数族裔学生的各种外展项目。这笔资金将支持他继续参与大学、学校和院系层面发起的教育活动，并创造场所发起其他包容性计划，使社区和大学更紧密地联系在一起。此外，将根据所描述的研究开发新的教学材料，用于本科生的化学和生物工程课程。该提案的教育部分将共同提高多个层次的教育质量，同时为公众提供科学管理。

This NSF award by the Biotechnology, Biochemical and Biomass Engineering program supports work in Prof. Park's lab to engineer and characterize novel peptide inhibitors of the Erk-2 kinase. Erk-2 plays a role in cell development, proliferation, and malignancies, and is an important model system for phosphorylation-dependent signaling in the cell. Considering that the mammalian genome encodes a large number of enzymes involved in phosphorylation regulation, being able to specifically target the activity of select kinases should lead to a better model of intracellular communication as well as to more effective therapeutics against genetic diseases caused by overactive kinases. The proposal seeks to engineer high affinity, high specificity peptide/protein inhibitors of Erk-2 by combining yeast display with disulfide trapping. The designed inhibitors will provide an orthogonal solution to kinase regulation beyond small molecule drugs, and significantly increase the potential pool of candidate molecules for regulating the kinase pathways. The designed peptide inhibitors are expected to have more favorable biophysical properties than small molecule inhibitors, including i) higher specificity of interaction to reduce unwanted side effects, and ii) robustness against fortuitous mutations in the target enzyme to minimize the occurrence of drug resistance. The proposed disulfide trapping strategy, which has been shown to crosslink interacting proteins that are co-expressed in the same yeast cell, provides a unique advantage over conventional screening methods in that the specificity of interaction is intrinsically built in the screen, so that the identified clones are far more likely to be functionally relevant. The successful implementation of the protocol will therefore be highly relevant to other engineering studies of specific protein interactions. Prof. Park has been an active participant of various outreach programs at UB designed to encourage and retain minority students in science and engineering. The funding will support his continued engagement in educational activities initiated at the university, school and department levels, as well as create venues to initiate other inclusive programs to bring the community and university closer together. Additionally, new pedagogical materials will be developed based on the described research for use in undergraduate chemical and biological engineering classes. The educational components of the proposal will collectively improve the quality of education at multiple levels, while providing scientific stewardship to the public.