Biomedical Engineering Reference
In-Depth Information
receptor. As distinct from valinomycin and gramicidin, the M2
peptide does not insert spontaneously into a lipid bilayer from the
bulk solution. Therefore, its incorporation was achieved by pre-
loading the vesicles used for the formation of the distal leaflet of
the tBLM with it. The M2 pore allows the passage of small mono-
valent cations such as Na
+
and K
+
, but not larger cations such as
tetramethylammonium (TMA). Upon incorporating M2, the re-
sistance of the lipid bilayer is reduced to one fifth of its value in
KCl solution, but increases again by exchanging the KCl solution
with a TMACl solution.
In spite of the lack of lateral mobility and low hydration of
these tBLMs, attempts have been made to incorporate the exotoxin
D-hemolysin from
Staphylococcus aureus
, a water soluble, mono-
meric, 293-residue polypeptide that forms heptameric pores in
lipid bilayers. Three different polyethyleneoxy-based tBLMs have
been employed: HS(CH
2
CH
2
O)
6
(CH
2
)
17
CH
3
(THEO-C
18
),
172
DPTL
and DPHDL,
173
which differs from DPTL by the presence of six
ethyleneoxy units, instead of four, and of two lipoic acid residues,
instead of one. With all thiolipids, incorporation of D-hemolysin
causes a decrease in resistance and an increase in capacitance of
the tBLM. The effect of D-hemolysin on the tBLM obtained by
self-assembling an egg-PC monolayer on the THEO-C
18
thiolipid
is bimodal and poorly reproducible. In some experiments, the fit-
ting of two RC meshes in series to the impedance spectra indicates
that the parameters ascribed to the hexaethyleneoxy moiety are
practically unaffected. This suggests that the protein penetrates
only the distal monolayer of the tBLM. In a second group of ex-
periments, the resistance ascribed to the hexaethyleneoxy moiety
decreases by two orders of magnitude. As a whole, the changes in
the hexaethyleneoxy and hydrocarbon moieties induced by D-
hemolysin suggest that this toxin penetrates the tBLM partially or
totally. The resistance of the DPHDL monolayer was very low
(0.33 k: cm
2
) when compared with that, 9.9 M: cm
2
, of the
DPTL monolayer. This denotes a loosely packed DPHDL mono-
layer. However, the formation of a DPhyPC monolayer on top of it
by vesicle fusion seems to heal some of its defects. Upon incorpo-
rating D-hemolysin, the resistance of the lipid bilayer moiety of the
DPHDL/DPhyPC tBLM decreases by three orders of magnitude,
and that of the DPTL/DPhyPC tBLM by one order of magnitude.
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