The lipid-bilayer membrane that encloses all cells
and their internal organelles has due to its important role in biology and
medicine been subject to intense investigations. Many functions of the cell
membrane are controlled by proteins that transduce signals across the lipid
bilayer, either via molecular-transport reactions or structural rearrangements.
An evidence of their important role is the fact that they are the targets of
more than half of today’s drugs, and significant effort has therefore been
devoted to the development of bioanalytical instruments capable of measuring
membrane-protein controlled reactions.
However, the enormous complexity of cell membranes,
being built up by numerous different lipids and proteins, combined with the fact
that membrane proteins must reside in this environment to function, makes
functional characterization of this class of protein extremely demanding. In
particular, isolation procedures needed for in-depth studies of an individual type
of membrane protein is complicated by the high tendency of membrane proteins to
lose function upon detergent-based solubilization procedures.
This demanding challenge has spurred intense activities
directed towards membrane-protein separation and enrichment protocols
applicable directly on natural cell-membranes, with the aim to bypass the
destructive solubilization step. Supported lipid bilayers, which are continuous
two dimensional cell-membrane mimics (originally developed for biosensor
applications) have due to preserved lateral mobility emerged as a promising system
for this type of membrane-protein separation. Up to now, however, the creation
of planar supported cell-membrane mimics from compositionally complex cell-membrane
derived vesicles has remained relatively elusive.
Our group has contributed to resolve this challenge
by first learning how to control the motion of a supported lipid bilayer in a
microfluidic channel , and then use this principle to induce fusion of
adsorbed cell-membrane derived vesicles with the front edge of a
hydrodynamically driven lipid bilayer (Fig. 1a) . Moreover, hydrodynamic forces generated in the microfluidic
channels were successfully utilized for both separation and accumulation of membrane-associated
proteins (Fig. 1b) [2-4].
Figure 1. (a) Schematic illustration
of a continuous lipid bilayer derived from native cell membrane, (b) micrograph
of the separation of two types of fluorescently labeled membrane-associated
proteins, (c) microfluidic setup, (d) polymer-cushioned planar cell membrane,
and (e) electrophoretic separation of transmembrane proteins.
In running projects, the group is further developing these
microfluidic-based (Fig. 1c) concepts with the aim to improve the spatial
mobility also of membrane proteins that span across the entire lipid bilayer, which
due to their close contact with the underlying support typically become
immobile. One approach is to utilize polymer cushions (Fig. 1d) in order
to separate the lipid membrane from the underlying support and thereby maintain
the mobility of transmembrane proteins. Another strategy is to introduce a
conductive polymer to the underlying substrate (Fig. 1e). The aim is in
this case to tune the interaction between protein and support and thereby also
influence the spatial mobility of the lipid membrane and its components.
Once local enrichment of transmembrane proteins is successfully
accomplished, it is our aim to employ surface analytical tools, such as SPR,
TIRF microscopy, electrical impedance spectroscopy and QCM-D, for functional
studies of membrane proteins in a near-native cell-membrane environment.
1. Jonsson, P.,
et al., Shear-Driven Motion of Supported
Lipid Bilayers in Microfluidic Channels. Journal of the American Chemical
Society, 2009. 131(14): p.
L., et al., Continuous Lipid Bilayers
Derived from Cell Membranes for Spatial Molecular Manipulation. Journal of
the American Chemical Society, 2011. 133(35):
P., A. Gunnarsson, and F. Hook, Accumulation
and Separation of Membrane-Bound Proteins Using Hydrodynamic Forces. Analytical
Chemistry, 2011. 83(2): p. 604-611.
B., et al., Hydrodynamic separation of
proteins in supported lipid bilayers confined by gold barriers. Soft
Matter, 2013. 9(39): p. 9414-9419.
Fredrik Höök (project
Swedish Research Council
Swedish Foundation for
Göran Gustafsson Foundation