Past Event

Thesis Defense In Chemistry, Presented by Zhiyu Sun

December 15, 2021
1:00 PM - 2:00 PM

Thesis Defense in Chemistry

Probing Allosteric Coupling and Dynamics with Solid-state NMR

Presented By Zhiyu Sun

Transmembrane allosteric coupling is a feature of many biological events, and we are intended to test if the allosteric coupling controls the c-type inactivation process with solid-state NMR. Since KcsA shares similar secondary structures with many human potassium channels, understanding the inactivation process on KcsA will shed light on more complex channel systems. Numerous studies have suggested that proton binding prompts a conformational switch, leading to a loss of affinity for potassium ions at the selectivity filter and therefore to channel inactivation. We tested this mechanism for inactivation using a KcsA mutant (H25R/E118A) that exhibits an open pH gate across a broad range of pH values. We are able to find a four order of magnitude potassium binding affinity change by doing solid state NMR. This result strongly supports our assertion that the open pH gate allosterically affects the potassium binding affinity of the selectivity filter. As the second part of the talk, I will go through our proton detection work on setting up the 1.3 mm fast magic angle spinning probe. We use microcrystalline ubiquitin as a model compound to set up 2D and 3D cross-polarization based proton-detected experiments. We also demonstrate non-uniform sampling for 3D experiments and T1, T2 and T1ρ relaxation measurement. At 60 kHz MAS, proton-detected ssNMR could serve as a powerful tool to study protein dynamics.

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