Research Initiation Award: Redox Regulated Phosphorylation of BCAT by PKC

研究启动奖为历史悠久的黑人学院和大学的初级和职业生涯中期教师提供支持，这些教师正在建立新的研究计划或重新定向和重建现有的研究计划。预计该奖项有助于进一步提高教师的研究能力和有效性，改善所在机构的研究和教学，并让本科生参与研究经验。授予北卡罗来纳农业与技术州立大学的奖项有可能增加对细胞膜的了解。这项拟议的研究旨在培养本科生和研究生学习一种影响细胞膜功能的酶。国际学生联合会将积极参与并招募少数族裔学生参与该项目，使这些学生能够从事前沿研究。这项活动将提高学生的技术和批判性思维能力。本计划的目标是通过生化和生物物理方法研究PKC、α和BCATC的相互作用。以往的等电点显示，胞浆人支链氨基转移酶(BCATC)的膜定位调节自噬小体的合成，从而进一步调节阿尔茨海默病(AD)，而BCATC的膜定位受蛋白激酶Cα(PKCα)介导的磷酸化的氧化还原调节。在这项拟议的工作中，PI将识别PKCα介导的BCATC中的磷酸化位点，表达和纯化PKCα磷酸化的BCATC，并解析磷酸化的BCATC的晶体结构，以分析由PKCα磷酸化引发的BCATC的结构变化。这项工作将通过共结晶和其他生物物理方法(如SPRI)来分析PKCα-BCATc复合体的结构，以揭示蛋白质-蛋白质相互作用的界面，并进一步了解磷酸化过程的机制。根据项目成果构建的模型将使PI能够理解PKCα氧化还原调节BCATC的磷酸化如何调节其构象变化，从而进一步调节其膜定位的机制。这是第一次在分子水平上描述这些相互作用，这将是对该领域的重大贡献，并有助于促进对这两种相互关联的BCATC监管模式的理解。该奖项反映了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。

Research Initiation Awards provide support for junior and mid-career faculty at Historically Black Colleges and Universities who are building new research programs or redirecting and rebuilding existing research programs. It is expected that the award helps to further the faculty member's research capability and effectiveness, improve research and teaching at the home institution, and involves undergraduate students in research experiences. The award to North Carolina Agricultural & Technical State University has potential to increase the understanding of the cell membrane. The proposed study intends to train undergraduate and graduate students in the study of an enzyme that affects cell membrane function. The PI will actively engage and recruit underrepresented minority students into the project so that the students may engage in cutting edge research. This activity will enhance the technical and critical thinking skills of students.The goal of this proposal is to study the PKCα-BCATc interaction via biochemical and biophysical methods. Previously the PI showed that the membrane localization of cytosolic human Branched-Chain Aminotransferase (BCATc) regulates the autophagosome synthesis which further regulates Alzheimer’s Disease (AD), and the membrane localization of BCATc is redox regulated by the Protein Kinase C α (PKCα) mediated phosphorylation. In this proposed work, the PI will identify the PKCα-mediated phosphorylation site in BCATc, express and purify the PKCα-phosphorylated BCATc, and solve the crystal structure of the phosphorylated BCATc to analyze the structural changes of BCATc triggered by the phosphorylation by PKCα. The work will analyze the structure of PKCα-BCATc complex via co-crystallization and other biophysical methods, e.g. SPRi to uncover the protein-protein interaction interface and further understand the mechanism of the phosphorylation process. The constructed model built from the project outcomes will enable the PI to understand the mechanism of how the redox-regulated phosphorylation of BCATc by PKCα regulates its conformational changes that further regulate its membrane localization. This is the first time that these interactions will be described at the molecular level, and this would be a significant contribution to the field and help advance the understanding of these two inter-related modes of BCATc regulation.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.