NMR Analysis of Designed Mini-Protein Molecular Motion

<p>Computationally designed mini-proteins are useful as these proteins can be designed to be thermally and chemically stable. Due to their stability, mini-proteins can be used as small-molecule drugs; drug transportation can be done easily as the miniproteins do not have to be carefully stored and will still have high activity. Furthermore, binding assays have shown that mini-proteins can be designed to have high affinity binding to their targets. The flexibility or rigidity of the mini-protein structure influences the stability of the protein as well, and the more rigid the mini-protein is, the more effective it is as a small-molecule drug. This thesis will focus on the molecular motion of these mini-proteins, which will be studied through Nuclear Magnetic Resonance (NMR) Spectroscopy.</p> <p>T₁, T_1œÅ, and NOE experiments are conducted in order to obtain R₁, R_1œÅ, and Œ∑ values. With the obtained values, R2 values can be calculated, and all of these values will be utilized in a program called ModelFree to obtain Lipari-Szabo order parameters. For the protein EHEE_rd2_0005, it is found that this protein is heterogeneous and is not a static structure, indicating that this protein is fairly rigid but has certain regions in the protein, such as the C-terminus, loops, and regions in the Œ≤-strands within the protein that are more flexible, or experience more motion. Circular Dichroism (CD) Spectroscopy will also be performed to try to obtain a melting temperature of the miniproteins of interest.</p>