Dissertation in Biophysics at the Center of Biochemistry and Molecular Biology & Department of Physics
(Prof. U.-P. Hansen), University of Kiel
Postdoc at the Center of Biochemistry and Molecular Biology & Department of Physics(Prof. U.-P. Hansen),
University of Kiel
Postdoc in the membrane biophysics group (Prof. G. Thiel), Department of Biology,
Technical University of Darmstadt
Postdoc in the Department of Biosciences (Prof. A. Moroni), University of Milan, Italy
Junior Group Leader in the Department of Biology, Technical University of Darmstadt
Heisenberg Fellow at the Institute of Physiology I, Jena University Hospital
Ion channels are involved in many physiological processes. Understanding the biophysical mechanisms of transport and gating in these proteins is therefore a prerequisite for the development of targeted therapies for many different diseases. The quantitative correlation of data from different sources, e.g. electrophysiological experiments, structural biology and computer simulations is needed for a full understanding of the molecular processes involved.
We employ electrophysiological, analytical and coarse-grained simulation techniques to improve this kind of quantitative correlation and to provide new information on channel function. The combination of these methods enables an interplay between computational and functional approaches leading to a stepwise improvement of the models on either side. To maximize the clearness of the experimental data, most experiments are done one the model system of viral potassium (K+) channels. More information about this can be found in Methods.
The selectivity filter gate
The selectivity filter (SF) gate in K+ channels is regulated by the occupation with K+ ions (orange circles) and a hydrogen bond network anchoring it to the pore helix (symbolized by the orange springs).
The selectivity filter (SF) is the central, most conserved core of K+ channels through all realms of life and has been shown to be a gate of physiological relevance. It is modulated by the occupation with K+ ions as they permeate the channel and by an intricate hydrogen bond network anchoring the selectivity filter to the pore helix. We could determine the occupation of K+ binding sites in the SF in a functioning channel under near-physiological conditions, and showed the correctness of these results by means of the electrostatic repulsion of TPrA by the ions in the SF. The quest to understand the signal chain between ion binding and the gate is continued within the framework of the DFG Research Unit 2518 ‘DynIon’.
Modularity of ion channel pores
Ion channels are modular proteins. Their central pore can function alone. However, in most channels, additional transmembrane or soluble domains provide regulatory functions. Prominent examples are the voltage-sensing domain (VSD) and various ligand-binding domains.
However, also the pore module itself is modular. The pores of all K+, Na+ and even some Ca2+ channels share a remarkably similar structure, but evolution has resulted in different strategies, for e.g. cytosolic gates. In this project, we will transfer “functional modules”, e.g. an isolated gate or the selectivity filter, from Kcv channels to eukaryotic channels and vice versa and investigate if and how these modules can transfer their function to the host channel. Being able to understand and create channel pores as modular systems from distinct building blocks will be a huge step in terms of both understanding disease-relevant mutations in a certain domain in the context of the whole protein and also towards rational protein engineering.
Cell-free expression and characterization of viral ion channels
To characterize the function of our model channels, we employ a fully cell-free system. This avoids e.g. the changes in lipid composition of the cell membrane or endogenous proteins that could influence the results in uncontrollable ways. The channels are expressed with a commercial in vitro expression kit into lipid nanodiscs and reconstituted into planar lipid bilayers for electrophysiological recordings.
Increasing the temporal resolution in electrophysiological experiments
Pore gating is the process of an ion channel switching between conductive (open) and nonconductive (closed) states. Often, this process is faster than the temporal resolution of the experimental setup and cannot be directly resolved, resulting in loss of information. Another problem is that molecular dynamics simulations typically span the temporal range of nanoseconds to microseconds.
We approach this problem in two ways. On the technical side, the use of next-generation amplifiers allows the recording of currents at higher bandwidths, while keeping the noise manageable. On the analytical side, by analyzing the so-called “excess noise” by “Extended Beta Distributions” we are able to retrieve this information and increase the effective temporal resolution. In house, we reach a resolution of microseconds, in cooperation with the Shepard lab at the Columbia University, even a resolution of 30 nanoseconds was reached well within the range of molecular dynamics simulations.
Lolicato, M., Bucchi, A., Arrigoni, C., Zucca, , Nardini, M., Schroeder, I., Simmons, K., Aquila, M., Difrancesco, D., Bolognesi, M., Schwede, F., Kashin, D., Fishwick, C.W.G., Johnson, A.P., Thiel, G., and Moroni, A. (2014)
Project P10 " Novel approaches to study the signal chain of the voltage-dependent gating in the selectivity filter of viral K+ channels " of the Research Unit 2518 DynIon funded by the DFG (2020)
Heisenberg fellowship by the DFG “Understanding fast gating of ion channels with new strategies for the interplay between computational and functional approaches” (2019)
Project “Functional impact of transmembrane domain and turret on potassium channel gating " funded by the DFG (2014)
Many years of teaching experience in biology, biomolecular engineerinf and (bio)physics.
Here in Jena: Lectures on ion channels in the research-oriented line of medical studies (FOM). Biophysics teaching in physics.
The Covid-19 pandemic requires increased flexibility and creativity in teaching from all of us. Many theoretical subjects can be covered by online offerings. We have to take care that the interaction between students on the one hand and between students and teachers on the other hand is not neglected and that active learning also finds its way into digital teaching. This can already be achieved by very simple means such as a quiz, which is solved by teams competing for the highest score in the fastest time. Seminars in which students not only reproduce a scientific publication, but also critically examine it in small groups, can also be realized well via video conferencing.