CAREER: The stereochemical basis of target selectivity encoded by specificity-determining loops in peptide-binding domains

获得该奖项后，化学系的生命过程化学计划资助西华盛顿大学的Jeanine Amacher博士研究在不同蛋白质家族中结构保守的环如何影响它们与其他蛋白质的结合及其作为催化剂的功能。阿马赫博士的团队将确定组成这些环的氨基酸序列的差异如何帮助定义特定针对结合伙伴的蛋白质的相互作用，或影响其作为酶的活性。对这些结构环的研究结果将揭示分子原理，以了解控制细胞通信方式的蛋白质相互作用。该项目将培养蛋白质生物化学和结构生物学的本科生和硕士研究生。该项目的研究和教育部分通过对本科生和他们的导师进行计算生物学的专门培训来整合。此外，当地以本科为主的机构和社区学院的学生将通过在西华盛顿大学举办的定期生命科学研讨会，与来自地区研究密集型大学的博士生和博士后研究助理建立联系，进行近距离同行指导。科学交流在社会中的重要性将被整合到主办机构开设的高级荣誉选修课“生物医学研究中的新兴技术的生命伦理学”中。这个项目将检验一个假设，即关键信号或调控蛋白的催化和非催化结构域的选择性和/或活性是由保守的结构环决定的。这些研究将表征细菌分类酶中结构保守的环，其中两个已知影响特异性，以及先前发现的在人类Src癌基因蛋白的SH2和SH3区域中介导蛋白质-蛋白质相互作用的特异性决定环的位置特异性贡献。自然序列变异将被用于设计嵌合蛋白和其他环变异体。与编码靶标特异性有关的这些环的结构-功能关系将使用基于FRET(荧光共振能量转移)的活性分析、X射线结晶学和分子动力学模拟来确定。此外，生化和计算方法，包括结构预测程序Rosetta，将被用来追踪导致保守环序列的靶标特异性的进化根源。这个项目将确定共同的原则，扩大对几个蛋白质家族结构保守的环中序列和特异性之间关系的理解。最后，更好地了解分类酶的特异性将有可能扩大分类酶介导的连接作为蛋白质工程工具的应用。这一奖项反映了NSF的法定使命，并通过使用基金会的智力优势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. Jeanine Amacher from Western Washington University to investigate how loops that are structurally conserved in various families of proteins affect their binding to other proteins and their functions as catalysts. Dr. Amacher’s group will determine how differences in the amino acid sequences that make up these loops help define the interactions that specifically targets a protein to its binding partner or affect its activity as an enzyme. Results from studies on these structural loops will reveal molecular principles to understand protein interactions that control how cells communicate. This project will train undergraduate and masters level graduate students in protein biochemistry and structural biology. The research and educational components of this project are integrated through specialized training in computational biology for undergraduate students and for their mentor. In addition, students at local primarily undergraduate institutions and community colleges will be connected with PhD students and postdoctoral research associates from regional research-intensive universities for “near-peer” mentoring through a regular Life Sciences Symposium hosted at Western Washington University. The importance of scientific communication in society will be integrated into an upper-level Honors “Bioethics of Emergent Technology in Biomedical Research” elective course to be offered at the host institution. This project will test the hypothesis that the selectivity and/or activity of catalytic and noncatalytic domains of critical signaling or regulatory proteins are determined by conserved structural loops. The studies will characterize structurally-conserved loops in bacterial sortases, two of which are known to affect specificity, along with site-specific contributions of previously identified specificity-determining loops in SH2 and SH3 domains of the human Src oncogene protein that mediate protein-protein interactions. Natural sequence variation will be applied to design chimeric proteins and other loop variants. The structure-function relationship of these loops with respect to encoding target specificity will be determined using FRET(Fluorescence Resonance Energy Transfer)-based activity assays, X-ray crystallography, and molecular dynamics simulations. In addition, biochemical and computational methods, including the structure prediction program ROSETTA, will be used to trace the evolutionary roots that lead to target specificity of conserved loop sequences. This project will identify common principles that broaden the understanding of the relationship between sequence and specificity in structurally-conserved loops across several protein families. Finally, a better understanding for the specificity of sortases will potentially expand the application of sortase-mediated ligation as a tool for protein engineering.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.