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Disputation Luiz Schubert

25.10.2024 | 16:00
Thema der Dissertation:
Investigating Directional Proton Transfer in Rhodopsins by Novel Time-Resolved Infrared Spectroscopies
Abstract: Directional proton transport across the lipid bilayer of a biological membrane is a process of fundamental importance in nature. One of the few techniques, that allows one to observe protonation reactions is infrared (IR) difference spectroscopy. Its chemical sensitivity sheds insight into the changes in protonation state of single moieties within proteins but is at the same time sensitive to all other molecular alterations accompanying proton transfer. IR difference spectroscopy is therefore a powerful tool to study the functional mechanism of protontranslocating proteins.
A very intriguing question is how proteins that act as membrane-bound proton pumps achieve vectorial proton transport. The first part of this thesis aims to address this question by the spectroscopic investigation of a novel microbial rhodopsin acting as an inward proton pump. I characterized the sequence of protonation reactions with a focus on the cytoplasmic half channel of the protein. Here, deprotonation of an aspartic acid located on the cytoplasmic side is correlated to the deprotonation of the distantly located retinal Schiff base. The sequence of these protonation events has important implications on the functional mechanism of inward proton transport, which is compared to the mechanism of a prototypical outward proton pump. While time-resolved IR spectroscopy is an established tool to monitor light-induced absorption changes of reversible processes, it has found only limited application in the study of irreversible and light-insensitive reactions. In the second part of this thesis, I showcase two novel IR absorption techniques based on quantum cascade lasers (QCL) to study protein reactions. Both techniques were used to monitor protein conformational changes as well as protonation dynamics by single-shot experiments, i.e. without the need to average multiple acquisitions. After a thorough comparison of both techniques, I applied them to the study of the irreversible activation process of the G-protein-coupled receptor rhodopsin. This approach demonstrates that QCL-based IR spectroscopy is a powerful tool for the spectroscopic study of irreversible and, in the future, also light-insensitive (protein) reactions.

Zeit & Ort

25.10.2024 | 16:00

Hörsaal B (0.1.01)
Fachbereich Physik, Arnimallee 14, 14195 Berlin