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BP: Fachverband Biologische Physik
BP 17: Protein Structure and Dynamics
BP 17.8: Talk
Wednesday, March 29, 2023, 17:00–17:15, BAR 0106
Multi-state Unfolding Processes: Discrimination of protein domains by urea-induced thermal shift — •Ji Young Yang1,2, Oliver Burkert2, Boris Mizaikoff1, and Jens Smiatek3,4 — 1Institute for Analytical and Bioanalytical Chemistry, University of Ulm, Ulm, Germany — 2Boehringer Ingelheim Pharma GmbH & Co. KG, Analytical Development Biologicals, Biberach(Riss), Germany — 3Boehringer Ingelheim Pharma GmbH & Co. KG, Development NCE, Biberach (Riss), Germany — 4Institute for Computational Physics, University of Stuttgart, Stuttgart, Germany
Co-solute induced molecular denaturation and aggregation mechanisms related to stability changes for multi-domain proteins like mAbs are often hard to monitor experimentally. In addition, a thorough theoretical explanation is often missing. We performed intrinsic fluorescence (IF) measurements of monoclonal antibody (mAb) samples for different aqueous urea concentrations under thermal denaturation. Our results show that the denaturating effect of urea on individual mAb domains can be explained by linear mapping of the thermal shifting curve to the actual urea concentration. Notably, the achieved thermal shifting curves can be assigned to certain protein domains, which enables discrimination of overlapping denaturation processes. Our approach highlights the benefits of direct monitoring of co-solute effects on the conformational stability of mAb domains and its colloidal stability. We will discuss the experimental approach and present the corresponding outcomes in terms of the underlying molecular mechanisms.