Physics And Chemistry Basis Of Biotechnology - De Cuyper - tiera.ru

Supported lipid membranes for reconstitution of membrane proteins
stability of the artificial bilayer since it would impose self-healing properties to the
bilayer. A fluid membrane is also of greatest significance for the successful
incorporation of an active membrane protein.
It has been observed that the composition of the lipid layers is not of crucial
importance for a successful incorporation of an active membrane protein. This is rather
surprising considering the great effort Nature has taken to vary the lipid composition in
the biological membrane. In practice, artificial cell membranes rarely consist of more
than a few different phospholipids and in many cases only one has been used.
Therefore they are much more homogeneous than the real plasma membrane and in
some respects they differ significantly from the forerunner. The net surface charge of
the artificial bilayer membrane is for instance often rather large and can vary between -
0.4 and + 0.4 C*m -2 and the effective interfacial width is only around 0.6 nm [8].
Furthermore, in many cases only one side of the biomimetic membrane faces an
aqueous phase whereas the other one faces a solid support.
2.2. 1, Unsupported artificial bilayer membranes
Two types of unsupported synthetic cell membranes have been used successfully for
decades in experimental studies: Firstly, it has long been recognised that a solution of
phospholipids under special conditions spontaneously forms uni- or multi-lamellar
vesicles consisting of one or several hundred concentric lipid bilayer membranes. The
size of the vesicles varies within large limits and they are sometimes classified
according to their size. Thus D.D. Lasic distinguishes between large multilamellar
vesicles (MLV’s), and large and small unilamellar vesicles (LUV's and SUV's) [9] and
this terminology is also adopted in this paper. Vesicle studies have found many
applications both in theoretical and experimental sciences, i.e.; topology investigations
of two-dimensional surfaces in three-dimensional space, phase transition studies in two
dimensions, artificial photosynthesis, drug delivery and medical diagnostics, etc. In
biochemistry and biology the focus is on reconstitution of membrane proteins into
artificial membranes and the study of model biological membranes. The long term
stability of the vesicles is good but since it is an unsupported membrane the bioactivity
of incorporated proteins cannot always be investigated in a straightforward way.
Secondly, so called black lipid membranes can easily be formed by painting a lipid
solution across an aperture in a hydrophobic septum that separates two aqueous phases.
In the pioneering work by Mueller et al variations in the dielectric properties of such an
artificial membrane caused by spontaneous adsorption of various water-soluble
macromolecules was reported [10]. Although these types of BLM provide excellent
models for biological membranes they suffer from some severe drawbacks. The total
surface area is small, typically less than 1 mm 2 , most often only low protein densities
can be reconstituted into them. Furthermore, they rarely last more than a few hours.
Less fragile painted lipid membranes can be formed on polycarbonate ultrafiltration
membranes for use in FIA-systems [ 1 1 -13].
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