Project Details
Projekt
Novel approaches to study the signal chain of the voltage-dependent gating in the selectivity filter of viral K+ channels
Applicant
Professorin Dr. Indra Schröder
Subject Area
Biophysics
Term
from 2020 to 2024
Project identifier
Deutsche Forschungsgemeinschaft (DFG) - Project number 291198853
The selectivity filter currently emerges as one of the main gates of potassium (K+) channels. It is known that both the occupation of a series of K+ binding sites in the selectivity filter (SF) and membrane voltage regulate this gate. The molecular processes underlying the gate function are only poorly understood. To allow experimental testing of theoretical models, we have recently determined from current recordings of a K+ channel the site-specific ion distribution in the SF and the correlation with the gating. We have also identified a specific K+ binding site as the voltage sensor and further concluded that changes in protein flexibility rather than conformational changes mediate gating. The subject of this proposal is a voltage-dependent gating process in the SF of a viral prototype K+ channel (Kcv) with sub-millisecond kinetics. By a combination of cell-free single-channel recordings and computational studies, we will identify a) how ion occupation shapes the flexibility of the SF, b) how the signal is transmitted from the sensor binding site to the gate and c) the location and mechanism of the gate. Single channels will be characterized by high-resolution recordings in planar lipid bilayers. Modified ion-filter interactions will be induced by different permeant ions and point mutations. The role of the hydrogen bond network between the pore helix and the SF will also be addressed by point mutations. Anisotropic network model (ANM) calculations will be used to screen mutations and different ion occupations. This will provide estimates for the related changes in flexibility and the principal components of structural fluctuations. This will allow to identify molecular mechanisms, which will then be quantitatively tested by global fits of Hidden Markov Models. The researchers of P3 will provide structural information from solid-state NMR experiments. Atomistic MD simulations will be performed by P5 to test the proposed molecular mechanisms. The expertise regarding fits of Hidden Markov Models in P2 will be used to improve the analyses. By this combination of experimental data with coarse-grained and atomistic computations, detailed biophysical insights into the mechanisms of SF gating in Kcv channels is expected.
DFG Programme
Research Units
