CAREER: Dynamic Regulation of Protein Quality Control

Miami University, Oxford OH

Investigators

Abstract

Title: CAREER: Dynamic Regulation of Protein Quality Control The three-dimensional structures of proteins determine the roles and functions proteins play within cells. Upon exposure to chemical or mechanical stresses, proteins can misfold, resulting in large changes in three-dimensional structure and loss of protein function. To survive, cells have developed quality control systems that guide misfolded proteins towards pathways that lead to them either being repaired or discarded. This project will determine the biological principles that allow cells to respond to protein misfolding by directing misfolded proteins for destruction. Another goal of this project is to develop techniques that will enable further investigations into how cellular responses to protein misfolding can be controlled. The integrated education objective of this CAREER project is to increase retention of underrepresented minorities in STEM through direct outreach at the high school and undergraduate levels via hands-on research experience and through different outreach programs. The project will integrate research directly into graduate and undergraduate courses with an emphasis on hands-on interpretation of real data and attention to development of visual, verbal and written communication skills. The requirement for cells to respond to mechanical, thermal or chemical stresses is ubiquitous in biology. The E3 ubiquitin ligase CHIP and the 70 kilodalton heat shock protein, Hsp70, play pivotal roles in the response to cellular stress through the convergence of refolding and degradative pathways. The research will forge connections between dynamics on the picosecond to millisecond timescales and the biological function of chaperoned ubiquitination complexes. The incorporation of gain-of-function mutants developed using insights from high-resolution X-ray crystallographic information will identify the effects of enhancing the CHIP/Hsp70 interaction. The project will redefine how the protein quality control field views the role of interactions between Hsp70 and CHIP in regulating how cells respond to protein misfolding by employing either refolding or degradative approaches. The combination of NMR, SAXS, and EPR with biolayer interferometry will generate advances in the use of hybrid methods for structural biology. A novel real-time ubiquitination assay will demonstrate the utility of biolayer interferometry for rapid and sensitive monitoring of a processive post-translational modification.