Biophysical Society Conference | Tahoe 2022

Molecular Biophysics of Membranes

Thursday Speaker Abstracts

INTEGRATIVE STRUCTURAL MODELING OF MEMBRANE PROTEINS USING SPARSE PARAMAGNETIC NMR AND NEUTRON SCATTERING DATA Kaitlyn V Ledwitch 1 ; Georg Kuenze 3 ; Elleansar Okwei 1 ; Katherine Larochelle 1 ; Shuo Qian 2 ; Hugh O'Neill 2 ; Jens Meiler 1,3 ; 1 Vanderbilt University, Center for Structural Biology, Nashville, TN, USA 2 Oak Ridge National Lab, Oak Ridge, TN, USA 3 University of Leipzig, Institute of Drug Discovery, Leipzig, Germany Combining different types of experimental information with computation is a powerful approach to investigate complex biological systems. Here, we combine experimentally-derived nuclear magnetic resonance (NMR) restraints and small-angle neutron scattering (SANS) data to guide membrane protein structural modeling in Rosetta. We demonstrate this integrative modeling strategy using the disulfide bond formation protein B (DsbB) in a nanodisc as a model membrane protein system. In this work, we developed two new technology platforms: 1) an unnatural amino acid-based paramagnetic NMR tagging strategy for membrane proteins and 2) a SANS deuteration strategy for producing ‘stealth’-like circularized nanodiscs (cNW9). NMR restraints from paramagnetic labeling gives structural information in the form of long-range distance and angular restraints. We complement these atomic details with SANS-derived envelopes to probe the overall geometric shape of DsbB in the nanodisc. These datasets are used as experimental input to evaluate how the membrane mimetic perturbs the overall structure and architecture of DsbB. We carried out these experiments for DsbB in dodecylphosphocholine (DPC) detergent micelles in parallel to compare differences between lipid-loaded nanodiscs and micelles. We demonstrate this approach for the model membrane protein system DsbB but our integrated biophysical framework is equally applicable to other complex membrane protein systems.

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