The long term goal is to understand the how physiologically relevant environments influence protein-protein interactions both in vitro and in living cells. Quinary structure occurs solely in the crowded cellular environment and involves nonspecific interactions between a protein and the macromolecules that surround it. These interactions by definition are absent in dilute buffered solutions. Protein-protein interactions are generally understood as protein complexes comprise >50% of the RCSB Protein Data Bank structures. Nearly all of these structures were studied in dilute buffered solutions where the macromolecule concentration is negligible. In a proteins native environment, the cell, the macromolecule concentration is >300 g/L, giving rise to quinary interactions. Our knowledge of how quinary interactions influence protein-protein interactions is lacking. Thus is it imperative to understand how the cellular environment effects protein-protein interactions. To do so we use the simplest model, a homodimer to investigate the influence of quinary interactions on protein-protein interactions using three specific aims.
Aim 1 : Determine how hard-core repulsions influence protein-protein interactions using 19F NMR and scaled particle theory Aim 2: Determine how chemical interactions influence protein-protein interactions by using protein cosolutes, lysates, and by modulating the electrostatic environment.
Aim 3 : Determine how the cellular environment effects protein-protein interactions using in-cell NMR and the domain swapped dimer. At the conclusion of these studies we will understand how the complex cellular environment effects protein- protein interactions. This knowledge will provide fundamental knowledge for understanding the forces that influence protein complexes in cells and how their disruption can lead to disease.
The environment inside of cells is drastically different from the dilute solutions predominately used to study proteins. Numerous in vitro and in cell studies have shown how this cellular milieu influences the equilibrium thermodynamics of protein folding for monomeric globular proteins but avoided studying protein-protein interactions due to their complexity. This project takes the next step to determine how the cellular environment affects protein-protein interactions and will provide foundational knowledge that will further our understanding of how disease relevant protein-protein interactions are influenced by the cellular environment.
| Guseman, Alex J; Perez Goncalves, Gerardo M; Speer, Shannon L et al. (2018) Protein shape modulates crowding effects. Proc Natl Acad Sci U S A 115:10965-10970 |
| Guseman, Alex J; Speer, Shannon L; Perez Goncalves, Gerardo M et al. (2018) Surface Charge Modulates Protein-Protein Interactions in Physiologically Relevant Environments. Biochemistry 57:1681-1684 |
