Palmitoylation is the reversible attachment of long lipid chains to proteins. It controls essential biological functions including cell signaling, immune regulation, and ion transport. Detachment of these lipid chains and recycling of membrane bound proteins is performed by acyl protein thioesterases (APTs), a conserved class of enzymes linked with cancer progression, neural degeneration, and bacterial pathogenicity. This research project will investigate the biological regulation of APTs, focusing on the role of protein flexibility and dynamics in controlling APT activity. The complex regulation of APT activity presents an intricate system for understanding how protein dynamics can regulate protein function. The dynamic regulation of APTs also has applications in understanding essential signaling pathways and disease states across human and bacterial systems. The research will also provide training opportunities for multiple undergraduate students within the investigator's laboratory and for broader investigations within linked classroom undergraduate laboratories. For classroom applications, an inquiry-based molecular biophysics laboratory will be added to the biochemistry major at Butler University and will be assessed for its effect on student learning and STEM career outcomes. This course and its outcomes will also be integrated with a previous NSF funded inquiry-based laboratory in biochemistry studying protein structure and function. To reach the broader community, a molecular biophysics symposium will be held at local undergraduate research conference and hands-on demonstrations of basic protein structure will be presented to young scientists at a public event aimed at exciting K-12 students about scientific discovery.
APTs catalyze the depalmitoylation of S-acylated proteins attached to the plasma membrane. An APT homologue from a gram-negative pathogen exists in conformational equilibrium between a closed and open state that is hypothesized to regulate its biological activity. Interconversion between these closed and open states is dependent on the structural dynamics of a flexible loop overlapping the active site. The structural properties of this flexible loop provide a straightforward model for how the enzymatic activity of APTs could be regulated and localized at the plasma membrane. The goal of this project is to understand the interplay between the protein loop dynamics and regulation of the catalytic and membrane binding activity of human and bacterial APTs. The central hypothesis is that structural rearrangement of this flexible, hydrophobic loop dually controls the catalytic and membrane binding activity of APTs. To address this hypothesis, a series of studies will be conducted to understand the loop dynamics, structural transition, and biological importance of hypothesized structural rearrangements within human and bacterial APTs. Confirmation of the dynamic nature of this flexible loop and its dual roles will provide a novel model for controlling the biological function of APTs.
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.
