EXPLORING THE ROLE OF POST-TRANSLATIONAL MODIFICATIONS IN THE STRUCTURE AND MOLECULAR INTERACTIONS OF GRASP65 HOMOLOG PROTEIN 1

The Golgi complex, an essential organelle in eukaryotic cells, plays a key role in modifying, storing, and transporting proteins and lipids. The structure of the Golgi complex is determined mainly by the Golgi matrix proteins (PMGs), including Golgi Reassembly and Stacking Protein (GRASP) and Golgins. The unique structural arrangement of stacked flattened membranes can be influenced by factors such as mitosis, apoptosis and the progression of several diseases, such as cancers. Recently, significant advances have been made in understanding the biophysical and biochemical properties of PMGs. These discoveries have challenged the traditional understanding of Golgi organization and have important implications for understanding the processes of protein secretion. However, the molecular mechanisms by which these membrane proteins alter the Golgi membrane structure remain unknown. Membrane anchoring of GRASPs passes through an essential post-translational modification (PTM) at the N-terminus. However, studies involving members of this family involve heterologous expression in prokaryotic systems that cannot perform the necessary PTMs. This project aims to explore the molecular mechanisms underlying the architecture of the Golgi complex, particularly focusing on the impact of PTMs on the structure and stability of GRASPs. Our ongoing investigation successfully refined an effective approach for N-terminal acetylation of the ScGRASP, a GRASP homolog native to Saccharomyces cerevisiae. Concurrently, we have started to probe the influence of this critical PTM on the anchoring of ScGRASP to biological membrane models. Preliminary results using Circular Dichroism and Differential Scanning Calorimetry suggest that acetylation plays a crucial role in the ScGRASP anchoring, with the interaction primarily facilitated by an amphipathic helix positioned in the N-terminus. This interaction necessitates a negatively charged membrane surface to undergo a disorder-to-order transition. It is anticipated that the advancements in this project will enhance our comprehension of the intricate molecular mechanisms that define the structure of the Golgi complex.