<b>Biophysics Seminar Series</b><br><i>Solid State NMR Studies of Protein Structure and Dynamics in Heterogeneous Environments</i>
Speaker: Dr. Joanna R. Long (University of Florida)
Obtaining high resolution structural information on biomolecules in heterogeneous environments, such as lipid membranes, mineralized tissue, or extracellular matrices, is not possible using standard high resolution NMR or x-ray techniques. We are developing and applying solid state NMR (ssNMR) techniques to the study of peptide structure and dynamics in unoriented membranes, immobilized on surfaces, and at varying levels of hydration. Many of these experiments rely on double-quantum filtering to simplify complex spectra and allow observation of specific interactions in peptides which comprise a small percentage of the sample under study. The advantages, challenges, and accuracy of these methodologies will be discussed along with experiments validating structural data obtained using model systems.
The second half of my talk will focus on the application of ssNMR to the study to peptide mimetics being developed for lung surfactant replacement therapies. Lipid-associated proteins play key roles in determining the unique physical properties of pulmonary surfactant, yet little is known about their atomic-level structure and dynamics in the membrane environment. Using ssNMR we have characterized the structure of a peptide mimetic, KL4, of lung surfactant protein B, SP-B, which is critical in forming a stable air-water interface in alveoli. With dipolar recoupling experiments, structures of KL4 when interacting with POPC/POPG and DPPC/POPG vesicles have been determined. The backbone torsion angles measured in KL4 are consistent with a distorted a-helix in POPC/POPG vesicles; in DPPC/POPG vesicles, a second conformation is observed with a lower helical pitch. These structures, coupled with 31P and 2H NMR studies of the effects of KL4 on lipid dynamics, leads to a molecular model for how KL4 differentially partitions into the two lipid environments. Given a nearly parallel alignment of the helix axis in a bilayer leaflet, with an extended conformation of the lysine sidechains, KL4 can partition more deeply into DPPC/POPG lipid bilayers while still allowing for the lysines to "snorkel" at a level where water can penetrate into the bilayers. This depth of peptide insertion would lead to negative membrane curvature strain. In contrast, the POPC/POPG conformer, with the lysine residues spread out more over the surface of the helix, is consistent with partitioning at the lipid/ water interface which would cause a positive curvature strain. The differential partitioning of the peptide into the lipids provides a mechanism for KL4 to enrich DPPC lipids at the lung air-water interface. These studies point to the importance of high-resolution, atomic level characterization to understanding lipid-associated peptides and proteins in their native environments.