research topics

Ion channels are ubiquitous proteins, vital to cellular processes such as cell membrane excitability and propagation of nervous stimuli. Defects within ion channel proteins alter their function and regulation and in turn, cause human diseases.

The main focus of the lab is to elucidate the molecular mechanisms of gating, selectivity, and modulation of potassium channels.  We investigate these phenomena using a combination of molecular biology, biochemistry, electrophysiology, x-ray crystallography, and cryo-electron microscopy.

calcium activated channels

MthK is a bacterial Ca2+ gated potassium channel from Methanobacterium thermoautotrophicum and has historically functioned as a model system for eukaryotic ligand-gated channels. Two distinct gates have been identified in MthK. A voltage-dependent gate at the selectivity filter and a C-type inactivation gate regulate the passage of ions through the channel.  By measuring the kinetic response of MthK combined with structural data, we were able to describe molecular details of gating and selectivity of potassium channels.  Recent structural data of MthK reconstituted into lipid nanodiscs have provided evidence for the ball-and-chain inactivation mechanism.

Image: Structural and functional studies of MthK reveal mechanistic details of gating

Cyclic nucleotide-gated channels

Cyclic nucleotide-gated (CNG) channels are central players in visual and olfactory signal transduction cascades. They are closely related to hyperpolarization-activated (HCN) channels, which are critical for pacemaker activity in the heart and are neuropathic pain targets. Cyclic nucleotides, such as cAMP and cGMP, are used to regulate the activity of these channels. Their architecture is modular and features a voltage-sensing domain, a pore domain, a cytosolic cyclic nucleotide-binding domain, and a C-linker domain that connects the CNBD to the pore.

We are interested in elucidating how different domains and structural elements work together to control CNG channel function. The eukaryotic channels from this family are difficult to express and purify, which impedes the analysis of these proteins under defined conditions. To that end, we recently introduced a bacterial CNG channel, SthK from Spirocheta thermophila, as model system to study this family of channels. SthK can be expressed and purified from bacteria and is suitable for many different in vitro assays including single channel bilayer recordings, stopped-flow based flux assays, equilibrium titrations, and ITC measurements. The single particle cryo-EM structure of SthK revealed the same architecture as its eukaryotic homologues. We now use SthK to address long standing questions about the regulation of CNG and HCN channels in molecular detail.

lipid modulation of ion channels

Integral membrane proteins, naturally, are in close contact with lipids and specific protein-lipid interactions have been identified in high-resolution structures. Given this intimate relationship, it is not surprising that proteins incorporated in membranes affect lipid bilayer properties and vice versa. We use both our model proteins, MthK and SthK, to analyze the effect of the bilayer composition on protein activity. Analysis of single channel characteristics as well as changes in macroscopic activity are used to gain insights into lipid modulation of these channels. The results are correlated with structural information on the channels reconstituted into nanodiscs of different lipid compositions.