Computational analyses on the structure-function relation in
ion channels
Creators: name: Furini, Simone <1979>
Publisher: name: Alma Mater Studiorum - Università di Bologna
Year: 2008
Subjects: ING-INF/06 Bioingegneria elettronica e informatica

description: Ion channels are protein molecules, embedded in the lipid bilayer of the cell
membranes. They act as powerful sensing elements switching chemicalphysical
stimuli into ion-fluxes. At a glance, ion channels are water-filled
pores, which can open and close in response to different stimuli (gating), and
one once open select the permeating ion species (selectivity). They play a
crucial role in several physiological functions, like nerve transmission,
muscular contraction, and secretion. Besides, ion channels can be used in
technological applications for different purpose (sensing of organic
molecules, DNA sequencing). As a result, there is remarkable interest in
understanding the molecular determinants of the channel functioning.
Nowadays, both the functional and the structural characteristics of ion
channels can be experimentally solved. The purpose of this thesis was to
investigate the structure-function relation in ion channels, by computational
techniques. Most of the analyses focused on the mechanisms of ion
conduction, and the numerical methodologies to compute the channel
conductance. The standard techniques for atomistic simulation of complex
molecular systems (Molecular Dynamics) cannot be routinely used to
calculate ion fluxes in membrane channels, because of the high
computational resources needed. The main step forward of the PhD research
activity was the development of a computational algorithm for the
calculation of ion fluxes in protein channels. The algorithm - based on the
electrodiffusion theory - is computational inexpensive, and was used for an
extensive analysis on the molecular determinants of the channel
conductance.
The first record of ion-fluxes through a single protein channel dates back to
1976, and since then measuring the single channel conductance has become
a standard experimental procedure. Chapter 1 introduces ion channels, and
the experimental techniques used to measure the channel currents. The
abundance of functional data (channel currents) does not match with an
equal abundance of structural data. The bacterial potassium channel KcsA
was the first selective ion channels to be experimentally solved (1998), and
after KcsA the structures of four different potassium channels were revealed.
These experimental data inspired a new era in ion channel modeling. Once
the atomic structures of channels are known, it is possible to define
mathematical models based on physical descriptions of the molecular
systems. These physically based models can provide an atomic description
of ion channel functioning, and predict the effect of structural changes.
Chapter 2 introduces the computation methods used throughout the thesis to
model ion channels functioning at the atomic level.
In Chapter 3 and Chapter 4 the ion conduction through potassium channels is
analyzed, by an approach based on the Poisson-Nernst-Planck
electrodiffusion theory. In the electrodiffusion theory ion conduction is
modeled by the drift-diffusion equations, thus describing the ion
distributions by continuum functions. The numerical solver of the Poisson-
Nernst-Planck equations was tested in the KcsA potassium channel (Chapter
3), and then used to analyze how the atomic structure of the intracellular
vestibule of potassium channels affects the conductance (Chapter 4). As a
major result, a correlation between the channel conductance and the
potassium concentration in the intracellular vestibule emerged. The atomic
structure of the channel modulates the potassium concentration in the
vestibule, thus its conductance. This mechanism explains the phenotype of
the BK potassium channels, a sub-family of potassium channels with high
single channel conductance.
The functional role of the intracellular vestibule is also the subject of
Chapter 5, where the affinity of the potassium channels hEag1 (involved in
tumour-cell proliferation) and hErg (important in the cardiac cycle) for
several pharmaceutical drugs was compared. Both experimental
measurements and molecular modeling were used in order to identify
differences in the blocking mechanism of the two channels, which could be
exploited in the synthesis of selective blockers. The experimental data
pointed out the different role of residue mutations in the blockage of hEag1
and hErg, and the molecular modeling provided a possible explanation based
on different binding sites in the intracellular vestibule.
Modeling ion channels at the molecular levels relates the functioning of a
channel to its atomic structure (Chapters 3-5), and can also be useful to
predict the structure of ion channels (Chapter 6-7). In Chapter 6 the structure
of the KcsA potassium channel depleted from potassium ions is analyzed by
molecular dynamics simulations. Recently, a surprisingly high osmotic
permeability of the KcsA channel was experimentally measured. All the
available crystallographic structure of KcsA refers to a channel occupied by
potassium ions. To conduct water molecules potassium ions must be
expelled from KcsA. The structure of the potassium-depleted KcsA channel
and the mechanism of water permeation are still unknown, and have been
investigated by numerical simulations. Molecular dynamics of KcsA
identified a possible atomic structure of the potassium-depleted KcsA
channel, and a mechanism for water permeation. The depletion from
potassium ions is an extreme situation for potassium channels, unlikely in
physiological conditions. However, the simulation of such an extreme
condition could help to identify the structural conformations, so the
functional states, accessible to potassium ion channels.
The last chapter of the thesis deals with the atomic structure of the !-
Hemolysin channel. !-Hemolysin is the major determinant of the
Staphylococcus Aureus toxicity, and is also the prototype channel for a
possible usage in technological applications. The atomic structure of !-
Hemolysin was revealed by X-Ray crystallography, but several experimental
evidences suggest the presence of an alternative atomic structure. This
alternative structure was predicted, combining experimental measurements
of single channel currents and numerical simulations.
This thesis is organized in two parts, in the first part an overview on ion
channels and on the numerical methods adopted throughout the thesis is
provided, while the second part describes the research projects tackled in the
course of the PhD programme. The aim of the research activity was to relate
the functional characteristics of ion channels to their atomic structure. In
presenting the different research projects, the role of numerical simulations
to analyze the structure-function relation in ion channels is highlighted.