Dr. Aaron Smith
University of Maryland, Baltimore County
Department of Chemistry/Biochemistry
Friday, October 8, 2021
“Structural and Regulatory Elements of Post-Translational Arginylation”
Eukaryotic post-translational arginylation, mediated by the family of enzymes known as the arginine transferases (ATE1s), is an important post-translational modification that can alter protein function and even dictate cellular protein half-life. Multiple major biological pathways are linked to the fidelity of this process, including neural development, cardiovascular development, cell division, and even the stress response. Despite this significance, the structural, mechanistic, and regulatory mechanisms that govern ATE1 function remain enigmatic. Research in my lab seeks to close this gap in understanding in order to target arginylation as a future therapeutic. While exploring arginine transferase function, we have discovered that ATE1s bind a previously undiscovered [Fe-S] cluster. We have used biochemical, spectroscopic, and analytical methods to decipher the composition and reactivity of this [Fe-S] cluster. Fascinatingly, we find that ATE1 cluster-binding preserves oligomeric homogeneity while increasing arginylation efficacy, demonstrating that this evolutionarily-conserved [Fe-S] cluster regulates arginylation rates. Furthermore, using a combination of X-ray crystallography, cryo-EM, and size-exclusion chromatography-coupled small angle X-ray scattering (SEC-SAXS), our lab has successfully solved the structure of Saccharomyces cerevisiae ATE1 (ScATE1). The three-dimensional structure of ScATE1 reveals a bilobed protein containing a canonical GCN5-related N-acetyltransferase (GNAT) fold. Structural superpositions and electrostatic analyses indicate this domain as the location of catalytic activity and tRNA binding. Additionally, our structure reveals the spatial connectivity of the N-terminal domain that binds the [Fe-S] cluster, hinting at the atomic-level details of the cluster’s regulatory influence. Coupled with a new regulatory framework, the first atomic-level structure of any ATE1 brings us closer to answering pressing questions regarding the molecular-level mechanism of eukaryotic post-translational arginylation.