Hindawi Publishing Corporation
Volume 2012, Article ID 138439, 11 pages
OnPhysical Propertiesof TetraetherLipidMembranes:
Parkson Lee-GauChong,Umme Ayesa,VarshaPrakashDaswani,andEllahChayHur
Department of Biochemistry, Temple University School of Medicine, 3420 North Broad Street, Philadelphia, PA 19140, USA
Correspondence should be addressed to Parkson Lee-Gau Chong, firstname.lastname@example.org
Received 6 July 2012; Accepted 8 August 2012
Academic Editor: Yosuke Koga
Copyright © 2012 Parkson Lee-Gau Chong et al. This is an open access article distributed under the Creative Commons
Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is
This paper reviews the recent findings related to the physical properties of tetraether lipid membranes, with special attention to
the effects of the number, position, and configuration of cyclopentane rings on membrane properties. We discuss the findings
obtained from liposomes and monolayers, composed of naturally occurring archaeal tetraether lipids and synthetic tetraethers as
well as the results from computer simulations. It appears that the number, position, and stereochemistry of cyclopentane rings
in the dibiphytanyl chains of tetraether lipids have significant influence on packing tightness, lipid conformation, membrane
thickness and organization, and headgroup hydration/orientation.
Archaea are subdivided into two kingdoms: euryarchaeota
and crenarchaeota . Euryarchaeota include methanogens
and halophiles, whereas crenarchaeota are traditionally com-
prised of thermophilic or hyperthermophilic archaea .
Halophiles and some methanogens are found mostly in high
salt water or hypersaline systems such as natural brines,
alkaline salt lakes, and salt rocks; while thermophilic and
hyperthermophilic archaea are found in very high temper-
ature environments . In recent years, crenarchaeota have
also been found in nonextreme environments such as soil
and pelagic areas [3, 4].
The plasma membranes of archaea are rich in tetraether
lipids (TLs) and diphytanylglycerol diethers, also known
as archaeols (reviewed in [11–13]). TLs are the domi-
nating lipid species in crenarchaeota, particularly in ther-
moacidophilic archaea (∼90–95%). They are also found in
methanogens (0–50%) but are virtually absent in halophiles.
Archaeal TLs contain either a caldarchaeol (GDGT) or
a calditoglycerocaldarchaeol (GDNT) hydrophobic core
(Figure 1) [13–17]. GDGT has two glycerols at both ends
of the hydrophobic core. GDNT has a glycerol backbone at
one end of the hydrophobic core and the calditol group at
the other end. Typically, TLs in methanogens contain only
GDGT, but TLs in thermoacidophiles, particularly in the
members of the order Sulfolobales, have both GDGT and
GDNT components. The Metallosphaera sedula TA-2 strain
from hot springs in Japan, which has only GDGT-based
lipids, is an exception . TLs have been thought to play
an important role in the thermoacidophile’s high stability
(e.g., 65-90◦C) and acidic environments (e.g., pH 2-3) .
However, more recent studies showed that GDGT-based
TLs are also abundant in nonextremophilic crenarchaeota
present in marine environments, lakes, soils, peat bogs,
and low temperature areas [20, 21]. The functional role of
tetraether lipids in crenarchaeota is not fully understood.
The hydrophobic core of archaeal TLs is made of
dibiphytanyl hydrocarbon chains, which may contain up
to 8 cyclopentane rings per molecule (reviewed in ).
The number of cyclopentane rings increases as growth
is a structural feature unique for archaeal tetraether lipids.
Therefore, it is of great interest to unravel its biological roles.
Various polar headgroups can be attached to the glycerol
Figure 1: Illustrations of the molecular structures of the bipolar tetraether lipids in the polar lipid fraction E (PLFE) isolated from S.
acidocaldarius. PLFE contains (a) GDGT (or caldarchaeol) and (b) GDNT (or calditolglycerocaldarchaeol ). The number of cyclopentane
and GDG(N)T-4 contain 0 and 4 cyclopentane rings per molecule, respectively (taken from , reproduced with permission).
tetraether (BTL) lipids. Archaeal BTLs are glycolipids or
phosphoglycolipids (illustrated in Figure 1). Liposomes that
are made of BTLs containing two or more sugar moieties
exhibit lower proton permeability than those containing
only one sugar molecule . It has been proposed that
thermoacidophilic archaea cells adapt to low pH and high
temperature by increasing the number of sugar moieties
and cyclopentane rings [26, 27]. Increasing the number of
cyclopentane rings tightens membrane packing (discussed
later) . Sugar moieties and the phosphate group in the
BTL polar headgroup regions interact with each other to
form a strong hydrogen bond network at the membrane
BTLs are unique to archaea and cannot be biosyn-
thesized by eukaryotic or bacterial cells. The ether for-
mation from glycerol has been studied to a great extent
( and references cited therein). The calditol moiety
of GDNTs can be synthesized via an aldol condensation
between dihydroxyacetone and fructose . Calditol is
then reduced and alkylated to form GDNTs . An in
vitro study showed that with the aid of 1L-myo-inositol
1-phosphate synthase, archaetidylinositol phosphate (AIP)
synthase and AIP-phosphatase, archaeal inositol phospho-
lipid (see Figure 1 e.g.) can be formed from CDP-archaeol
and D-glucose-6-phosphate via myo-inositol-1-phosphate
and AIP . It has been proposed that the cyclopentane
rings in BTLs of Sulfolobus are synthesized from glucose by
a “cyclase” enzyme of the calditol carbocycle .
physical properties of tetraether lipid membranes, with spe-
figuration of cyclopentane rings on membrane properties.
We discuss the findings obtained from model membranes
composed of naturally occurring archaeal tetraether lipids
and synthetic tetraethers as well as the results from computer
Membranes Composed of
Thermoacidophilic Tetraether Lipids
2.1. Membranes Made of Total Polar Lipid Extracts. The
stability and physical properties of liposomes made from
the total polar lipids (TPLs) extracted from archaea have
been studied extensively (reviewed in [11, 12, 33, 34]). TPL
extracts contain both diether and tetraether lipids. The gen-
eral trend shows that membranes become more stable as the
mole fraction of tetraether lipids increases. As an example,
liposomes made of diether lipids such as Methanosarcina
mazei TPL (0wt% in caldarchaeols) were unstable against
simulated human bile while those made of TPL from
Methanobacterium espanolae (65% in caldarchaeols) and
Thermoplasma acidophilum (90% in caldarchaeols) were
relatively more stable . Solute and water permeability
made with archaeal TPLs increases .
TPL from the archaeon M. smithii AL1 can be highly fuso-
genic when exposed to low pH and α- and β-glucosidases.
It was suggested that, at low pH (4.8), the positively charged
glucosidases interact with the anionic phospholipids in M.
smithii TPL, which in turn causes archaeosomes to rapidly
aggregate . Aggregation is a prerequisite for membrane
fusion. This result is somewhat surprising because previous
studies showed that tetraether liposomes are resistant to
fusogenic compounds [38–40]. Since TPL of M. smithii
AL1 contains a significant amount of diethers, in addition
to caldarchaeols (∼40wt %), it is possible that the strong
fusogenic activity mentioned above comes from the diether
2.2. Membranes Made of Partially Purified Tetraether Lipid
Fractions. Since tetraethers are the dominating lipid species
in thermoacidophiles, and the presence of diethers in the
total polar lipid extracts makes the data interpretation more
difficult, it is of biophysical interest to study membranes
made only with tetraether lipids. The physical properties
of lipid membranes made of partially purified polar lipid
fractions from the archaeon Sulfolobus solfataricus have
been reviewed [11, 34]. In this section, we focus on the
recent studies of membranes made of partially purified
polar lipid fractions isolated from the archaeon Sulfolobus
2.2.1. PLFE. The polar lipid fraction E (PLFE) is one
of the major bipolar tetraether lipids (BTLs) found in
the thermoacidophilic archaeon S. acidocaldarius [41, 42].
PLFE is a mixture of GDNT and GDGT (Figure 1). The
GDNT component (∼90% of total PLFE) contains phospho-
myo-inositol on the glycerol end and β-glucose on the
calditol end, whereas the GDGT component (∼10% of total
PLFE) has phospho-myo-inositol attached to one glycerol
and β-D-galactosyl-D-glucose to the other glycerol skeleton
(Figure 1). The nonpolar regions of these lipids consist of
a pair of 40-carbon biphytanyl chains, each of which may
contain up to four cyclopentane rings .
2.2.2. PLFE Liposomes. PLFE lipids can form stable unil-
amellar (∼60–800nm in diameter), multilamellar, and giant
unilamellar (∼10–150μm) vesicles [40, 41, 43]. The lipids in
these vesicles span the entire lamellar structure, forming a
bilayer structure formed by monopolar diester (or diether)
phospholipids. Compared to liposomes made of diester
or diether lipids, PLFE liposomes exhibit extraordinary
membrane properties (reviewed in [11, 12, 34]). PLFE
liposomes exhibit low proton permeability and dye leakage
[45, 46], high stability against autoclaving and Ca2+-induced
vesicle fusion [40, 47], tight and rigid membrane packing
, and low enthalpy and volume changes associated with
the phase transitions [48, 49].
It is known that a decrease in archaeal cell growth
in archaeal TLs . In the case of S. acidocaldarius, the
average number of cyclopentane rings per tetraether lipid
molecule decreases from 4.8 to 3.4 when Tg drops from
82◦C to 65◦C . Recent experimental work (see below)
has addressed the effect of Tg, inferentially the number of
cyclopentane rings, on the physical properties of tetraether
2.2.3. Effect of Cyclopentane Rings on Phase Behavior of
PLFE Liposomes. The phase behavior of PLFE liposomes has
been characterized by small angle X-ray scattering, infrared
and fluorescence spectroscopy, and differential scanning
calorimetry (DSC). PLFE liposomes exhibit two thermally-
induced lamellar-to-lamellar phase transitions at ∼47–50◦C
and ∼60◦C [34, 43, 48, 49] and a lamellar-to-cubic phase
transition at ∼74–78◦C [48, 49] all of which involve small
or no volume changes as revealed by pressure perturbation
calorimetry (PPC) . The calorimetry experiments also
suggested that the number of cyclopentane rings in the dibi-
phytanyl chains affect membrane packing in PLFE liposomes
because the liposomes derived from different cell growth
temperatures showed different thermodynamic properties
. DSC allows us to determine the enthalpy change (ΔH)
of the phase transition. PPC, on the other hand, allows us to
determine the relative volume change (ΔV/V) at the phase
transition and the thermal expansivity coefficient (α) at each
For PLFE liposomes derived from cells grown at 78◦C,
the DSC heating scan exhibited an endothermic transi-
tion at 46.7◦C, which can be attributed to a lamellar-to-
lamellar phase transition and has an unusually low ΔH
(3.5kJ/mol), when compared to that for the main phase
transitions of saturated diacyl monopolar diester lipids (e.g.,
1,2-dimyristoyl-sn-glycero-3-phosphocholine, DMPC). The
PPC scan revealed that, at this same phase transition, the
relative volume change (ΔV/V) in the membrane is very
small (∼0.1%) and much lower than the ΔV/V value 2.8%
for the main phase transition of DMPC. The low ΔH
and ΔV/V values may arise from the restricted transauche
conformational changes in the dibiphytanyl chain due to
the presence of cyclopentane rings, branched methyl groups,
and to the spanning of the lipid molecules over the whole
For PLFE liposomes derived from cells with growth
temperature of 65◦C, similar DSC and PPC profiles were
obtained. However, the lower cell growth temperature
30 405060 7080 90
Figure 2: Adiabatic compressibilities (ko
squares), 76◦C (open circles), and 81◦C (open triangles). Solid line:
DPPC liposomes for comparison (taken from , reproduced with
S) of PLFE liposomes
yielded a higher ΔV/V (∼0.25%) and ΔH (14kJ/mol) value
for the lamellar-to-lamellar phase transition measured at
pH 2.1. The lower growth temperature also generated less
ΔH, and the temperature dependence of α can be attributed
to the decrease in the number of cyclopentane rings in PLFE
due to the lower growth temperature . A decrease in the
and less rigid; thus, a higher ΔV/V value is shown through
the phase transition.
2.2.4. Effect of Cyclopentane Rings on Compressibility and
Membrane Volume Fluctuations of PLFE Liposomes. The
isothermal and adiabatic compressibility and relative volume
fluctuations of PLFE liposomes have been determined by
using calorimetry (DSC and PPC) and molecular acoustics
(ultrasound velocimetry and densimetry) . The com-
pressibility values of PLFE liposomes were low, compared
to those found in a gel state of 1,2-dipalmitoyl-sn-glycero-3-
phosphocholine (DPPC) . Relative volume fluctuations
of PLFE liposomes at any given temperature examined were
1.6–2.2 times more damped than those found in DPPC
liposomes . Volume fluctuations are closely related to
solute permeation across lipid membranes  and lateral
motion of membrane components . Thus, the low values
of relative volume fluctuations explain why PLFE liposomes
exhibit unusually low proton permeation and dye leakage
[45, 46] as well as limited lateral mobility, especially at low
temperatures (e.g., <26◦C) [43, 53].
Zhai et al.  have used the growth temperature Tg
to alter the structure of PLFE lipids. They determined
the compressibilities and volume fluctuations of PLFE
liposomes derived from different cell growth temperatures
(Tg = 68, 76, and 81◦C). The compressibility and volume
fluctuation values of PLFE liposomes exhibit small but
significant differences with Tg. Figure 2 shows that adiabatic
isothermal compressibility (ko
coefficient (βT) and relative volume fluctuations, a similar,
but somewhat different, trend is seen: (Tg = 68◦C) >
(Tg = 81◦C) ≥ or ≈ (Tg = 76◦C). These data indicate
that, among the three employed growth temperatures, the
inferentially the most tightly packed PLFE lipid membranes.
Note that 76◦C is in the temperature range for optimal
growth of S. acidocaldarius (75–80◦C, [54, 55]). This finding
suggests that membrane packing in PLFE liposomes may
actually vary with the number of cyclopentane rings in a
nonlinear manner, reaching maximal tightness when the
tetraether lipids are derived from cells grown at the optimal
growth temperatures .
S) of PLFE liposomes changes significantly
T), isothermal compressibility
2.2.5. Future Studies of Physical Properties of Tetraether Lipid
Membranes. PLFE is a mixture of GDNT- and GDGT-
derived BTLs with varying numbers of cyclopentane rings.
Furthermore, at any given growth temperature, there is
always a broad distribution of the number of cyclopen-
tane rings. In order to gain more insight into the effect
of cyclopentane rings on compressibility and membrane
volume fluctuations, it will be necessary to use purified
archaeal BTLs with a well-determined number and location
of cyclopentane rings. It has been reported that intact
of the archaeon Thermoplasma acidophilum can be sep-
arated with single cyclopentane ring resolution by high-
performance liquid chromatography (HPLC) as detected by
evaporative light-scattering detection [26, 56]. However, the
study by Shimada et al. on T. acidophilum was limited to
GDGT-based BTLs. To separate intact archaea BTLs at single
cyclopentane ring resolution when both GDNT- and GDGT-
derived BTLs are present remains a major challenge.
Hydrolyzed BTLs can also be separated with single
cyclopentane ring resolution using normal phase HPLC
and positive ion atmospheric pressure chemical ionization
mass spectrometry . Figure 3 shows the structures of the
cyclopentane-containing GDGT hydrophobic cores previ-
ously identified from the archaeon Sulfolobus solfataricus.
These structures were determined by mass spectrometry.
Compounds F?and G?(Figure 3) were reported as minor
components in S. solfataricus . The relative distribution
of these GDGT structures varies from species to species.
The GDGT fraction of S. solfataricus is dominated by those
structures with one (Structures E and G, Figure 3) or two
(F) biphytanyl chains with two cyclopentane rings. The
distribution of GDGTs in the extract of the archaeon M.
sedula is somewhat different. In this case, the distribution is
dominated by structures containing one or two biphytanyls
with one cyclopentane ring. Physical properties of liposomes
made of hydrolyzed BTLs (without sugar and phosphate
[M + H]+
Figure 3: Structures of cyclopentane ring containing GDGTs previously reported to exist in archaea . The number of cyclopentane rings
in the first and second hydrocarbon chains is indicated in the parentheses. The mass-to-charge ratio (m/z) of the protonated form [M+H]+
for each structure is also listed.
moieties) are not expected to be the same as those obtained
from the liposomes made of intact BTLs .
2.2.6. Disruption of PLFE Liposome Stability. While BTL
liposomes (such as PLFE liposomes) exhibit remarkable
stability against a number of chemical and physical stressors
ished under certain conditions. The most striking finding
in this regard is that PLFE liposomes become excessively
disrupted by the presence of two archaeal proteins, namely,
CdvA and ESCRT-III (ESCRT: endosomal sorting complex
protein that forms structures at mid-cell prior to nucleoid
to aid in the final steps of cell division in some species
of archaea. Negative stain electron microscopy revealed
extensive deformation of PLFE liposomes in the presence
of both CdvA and ESCRT-III together, but not individually
. The molecular mechanism underlying this disruption
is not clear.
PLFE liposomes are “autoclavable.” However, low pH
(<4) and low salt concentrations (<50mM) are unfavorable
for autoclaving PLFE-based liposomes . PLFE liposomes
and PLFE-based stealth liposomes (e.g., 95mol% PLFE,
polyethylene glycols (2000) (DSPE-PEG(2000)) and 2mol%
DSPE-PEG(2000)-maleimide) are extraordinarily stable
against autoclaving between pH 4–10 . These liposomes
retained their particle size and morphology against multiple
autoclaving cycles. One autoclaving cycle refers to the
incubation of a sample for 20min at 121◦C under a steam
pressure of ∼18psi. However, at pH 2-3, one or two
autoclaving cycles appeared to disrupt these liposomal
membranes, causing a significant increase in particle size
and autoclaving conditions. As the salt concentration was
decreased from 160 to 40mM, the percent of dye molecules
that leaked out from PLFE-based stealth liposomes after one
autoclaving cycle increased from 10.8% to 56.3% .
by surfactants. The effect of the surfactant n-tetradecyl-
β-D-maltoside (TDM) on unilamellar vesicles composed
of PLFE and POPC (1-palmitoyl-2-oleoyl-sn-glycero-3-
phosphocholine, a monopolar diester lipid) has been exam-
ined . TDM disrupts the POPC/cholesterol vesicles
effectively; however, higher concentrations (∼10 times) of
TDM were required to disrupt PLFE/POPC vesicles.
2.2.7. Structural and Packing Properties of PLFE Monolayer
Films Spread at the Interface between Air and Water. Effects
of cell growth temperature, subphase temperature and pH,
and lateral film pressure on PLFE lipid monolayers at the air-
water interface have been examined using X-ray reflectivity
(XRR) and grazing incidence X-ray diffraction (GIXD)
. XRR and GIXD determine the vertical and horizontal
structure of the monolayers, respectively.
For PLFE derived from cells grown at 76◦C, a total
monolayer thickness of ∼30˚ A was found in the XRR mea-
surements for all monolayers studied. This finding suggests
that both head groups of a U-shaped conformation of the
molecules are in contact with the subphase and that a
single hydrocarbon chain region is protruded into the air.
Similar U-shaped monolayer structures have been reported
in other tetraether lipid membranes . However, some
other studies [60, 61] suggest that the U-shaped and the
upright conformations may coexist in the monolayer at the
same time or occur sequentially after spreading the TL lipids
at the water-air interface.
At the subphase temperatures 10◦C and 20◦C, large,
highly crystalline domains were observed by GIXD; and the
thickness of the crystalline part of the monolayer is slightly
larger than 30˚ A, which indicates a tight packing of the whole
lipid monolayer, including both the hydrocarbon chain and
the head group regions. The area per hydrocarbon chain
of PLFE (∼19.3˚ A2) found by GIXD is significantly smaller
than that of DPPC (∼23.2˚ A2) or 1,2-dipalmitoyl-sn-glycero-
3-phosphoglycerol (DPPG) (∼22.6˚ A2). In fact, both the two
hydrocarbon chains of a single PLFE lipid and the chains
of neighbouring lipid molecules adopted an extremely tight
For PLFE lipids derived from cells grown at higher
temperatures, a slightly more rigid structure in the lipid
dibiphytanyl chains was observed. However, the growth
temperature, inferentially the number of cyclopentane rings,
does not affect the parameters of the unit cell in GIXD
measurements. This suggests that there exists a nearly
identical crystalline packing of all the PLFE lipids examined
and that, at high film pressures, membrane packing is
primarily governed by the lipid headgroup region . It is
interesting to mention that the lack of cyclopentane rings
in the bipolar tetraether lipids from M. hungatei has been
suggested to be the cause of the U-shaped configuration
adopted by these lipids in the monolayer film at the air-water
would hinder the dibiphytanyl hydrocarbon chains from
bending to form the U-shaped configuration.
3.PhysicalPropertiesof Membranes Made of
Synthetic Tetraether Lipids
The process of isolating well-defined archaeal tetraether
lipids can be difficult and time consuming. In addition,
archaeal tetraether lipids have several structural features
distinctly different from conventional diester lipids. There-
fore, it is rather difficult to elucidate the structure-activity
relationship for each of the individual structural features
when using native archaeal lipids. To resolve these problems
3.1. Importance of the Stereochemistry of the Cyclopentane
Ring. Jacquemet et al. were able to study the effect of the
stereochemistry of the cyclopentane ring on BTL membrane
properties by using two synthetic tetraether lipids [8, 9]
(Compounds 1 and 2 in Figure 4). Both lipids have a
bridging hydrocarbon chain with a single 1,3-disubstituted
cyclopentane ring at the center. The substitutes on the
ring are ether-linked to C3 of the two opposite glycerol
moieties, while C2 of the glycerols is ether-linked to a
phytanyl chain and C1 is linked to a lactosyl polar headgroup
(Figure 4). The only difference between these two isomers
is the configuration (cis or trans) of the 1,3-disubstituted
cyclopentane ring [8, 9].
The trans-isoform showed multilamellar vesicles whereas
the cis-counterpart led to nonspherical nanoparticles, as
revealed by cryo-transmission electron microscopy .
Small angle X-ray scattering (SAXS) studies further showed
that the cis-isomer exhibited Lc-Lα-QII(cystal, lamellar, and
bicontinuous cubic phase (Pn3m), resp.) phase transitions
whereas the trans-isomer remained in Lα phase from 20
to 100◦C. The electron density profiles calculated from the
SAXS data were consistent with a stretched conformation
of these synthetic BTLs within the Lα phase . The
difference in the phase behaviors was attributed to the
conformation equilibrium of 1,3-disubstituted cyclopenatne
is pseudorotational . Pseudorotation is more restricted
for the trans-isomer whereas several more orientations of
the two substituents on the ring can be created for the cis-
1,3-dialkyl cyclopentane ring [9, 68, 69]. Even though this
study shows that the stereochemistry at the cyclopentane
work is still required in order to explain why liposomes
made of PLFE, which naturally occurs and contains trans-
1,3-disubstituted cyclopentyl rings, can undergo the Lα-
to-QII phase transition [48, 49], while the synthetic trans
Figure 4: Synthetic tetraether lipids that have been used to study the effect of configuration (Compounds 1 and 2 [8, 9]) and position
(Compounds 3 and 4 ) of the cyclopentane rings on membrane properties.
BTL (Figure 4) cannot . Note that the placement and
the number of cyclopentane rings are different between
PLFE lipids (Figure 1) and the synthetic BTLs mentioned
above (Figure 4). Apparently, BTLs with subtle differences
in chemical structures can display distinctly different phase
The difference in the polar headgroups between PLFE
and the above-mentioned synthetic BTLs also leads to
other subtle structural differences. The d-spacing of PLFE
liposomes increases with increasing temperature , which
is contrary to that obtained from the synthetic trans-
isomer mentioned above (Compound 2 in Figure 4) . The
increased d-spacing with temperature is probably due to an
increase in hydration at the polar headgroup of PLFE .
For unknown reasons, there is no change in hydration at
3.2. Influence of the Position of the Cyclopentane Ring. Brard
et al. studied the effect of the position of the cyclopentane
ring on physical properties of tetraether lipid membranes
. They synthesized two tetraether glycolipids, each of
which contains a single cis-1,3-disubstituted cyclopentane
ring in the bridging chain. One glycolipid contained a
cyclopentane ring in the middle of the bridging chain while
the other had one at three methylene units from the glycerol
backbone (Compound 3 and 4 in Figure 4). This helped
them determine the influences of the different positions of
the cyclopentane ring.
The cyclopentane ring position appears to have a
profound impact on hydration properties, lyotropic liq-
uid crystalline behavior, and membrane organization .
Moreover, the synthetic BTL with the cyclopentane ring
positioned at the center (Compound 3 in Figure 4) can be
completely dispersed in water, and it can form sponge-like
compound with the cyclopentane ring away from the center
(Compound 4 in Figure 4) can only be partitially dispersed
in water and it forms multilamellar vesicles. It has been
suggested that the position of the cyclopentane ring in the
bridging chain influences the orientation of the glycosidic
polar headgroups attached to the glycerol backbone, which
leads to different membrane organizations .
4.Membrane PropertiesRevealed by
4.1. Effect of Cyclopentane Rings on Membrane Packing and
Headgroup Orientation. An increase in growth temperature
is known to increase the number of cyclopentane rings in the
dibiphytanyl chains of archaeal lipids . The number of
cyclopentane rings may vary from 0 to 4 in each biphytanyl
chain (i.e., 0 to 8 per dibiphytanyl unit). To evaluate how
the number of cyclopentane rings might affect membrane
packing, Gabriel and Chong have conducted molecular
modeling studies on a membrane containing 4 × 4 GDNT
molecules (with sugar moieties, Figure 1) . It was found
that when 8 cyclopentane rings are contained, the headgroup
of GDNT runs almost parallel to the membrane surface.
However, without containing any rings, the headgroup
is oriented perpendicular to the membrane surface. The
molecular modeling further showed that an increase in the
number of cyclopentane rings in the dibiphytanyl chains
of GDNT from 0 to 8 made GDNT membrane packing
tighter, more rigid, and more negative in interaction energy
(−156.5kcal/mol for 0 cyclopentane ring to −191.6kcal/mol
with 8 rings ). The resulting energy lowering effect is
neither due to the decrease in polar headgroup separation,
nor the change in the van der Waals interactions. Instead, it
is due to the more favorable hydrogen bonding, and bonded
interactions including harmonic bending, theta expansion
bond angle bending, dihedral angle torsion, and inversion
4.2. Effect of Macrocyclic Linkage on Membrane Properties.
Most archaeal BTLs are macrocyclic molecules with two
biphytanyl hydrocarbon chains linked to two opposite
glycerol or calditol backbones (illustrated in Figure 1 for
the case of PLFE). The effect of the macrocyclic linkage
on membrane properties has been studied by molecular
dynamics simulations [10, 70, 71]. For simplicity, coarse
graining approaches were employed and BTL molecules
were modeled as di-monopolar lipids such as di-DPPC
 and diphytanyl phosphatidylcholine (DPhPC) . In
essence, two monopolar molecules were tethered together
either at one pair of the hydrocarbon chains (acyclic di-
DPPC or di-DPhPC) or at both pairs (cyclic di-DPPC or
di-DPhPC). The simulations showed that in the membranes
composed of macrocyclic BTL-like molecules, the upright
configuration gains favor over the U-shaped configuration
. The macrocyclic linkage also leads to a condensing
effect on the membrane surface, increases the order of
the lipid hydrocarbon chains, slows lateral mobility in the
membrane, and increases membrane thickness [10, 70, 71].
Furthermore, the molecular dynamics simulations made by
the dissipative particle dynamics method  revealed the
formation of two types of membrane pores. Hydrophobic
pores are unstable and transient and exist at the low
temperature. Hydrophilic pores are more stable with much
longer lifetimes and are observed at high temperatures. The
simulation data  suggested that hydrophilic pores can
lead to the rupture of membrane vesicles. More intriguingly,
it was proposed that hydrophobic pores, which occur at low
temperatures, may result in the permeation of encapsulated
small molecules . This implies that although BTL
membranes are extremely stable and tightly packed, some
small leakage of entrapped molecules can still occur due to
volume fluctuations [6, 50] (discussed earlier).
5.Applications of Tetraether LipidMembranes
The extraordinary stability of tetraether lipid membranes
against a variety of physical and biochemical stressors
has provided the basis for using these lipids to develop
technological applications. BTLs can be used as a stable lipid
nanoparticles for targeted imaging and therapy (reviewed in
It has been proposed that liposomes made of archaeal
lipids (also called archaeosomes) are taken up via a phago-
cytosis receptor in the plasma membrane of the target
cells . This uptake occurs in a liposomal composition-
dependent manner . Total polar lipids from the
archaeon, Halobacterium salinarum CECT 396, have been
used to make archaeosomes and archaeosomal hydrogels
as a possible topical delivery system for antioxidants .
Compared to conventional liposomes, those archaeosomes
and archaeosomal hydrogels showed better stability and
more sustained drug release . It is of interest to
salinarum CECT 396) to tetraether lipids (e.g., PLFE lipids
isolated from thermoacidophiles). BTL-based liposomes are
suitable for oral delivery of therapeutic agents because BTL
liposomes are stable against the harsh conditions (such
as bile salts, pancreatic enzymes, and low pH) in the
gastrointestinal tract . Tetraether lipid membranes have
also been tailored and evaluated as an intranasal peptide
delivery vehicle . PEGylated tetraether lipids have been
synthesized and tested for their stability in test tubes and for
liposomal encapsulation potential . Knowledge gained
from the physical studies of cyclopentane rings, sugar
moieties, and macrocyclic structures should help to optimize
the numerous potential applications.
bipolar tetraether lipids
differential scanning calorimetry
ESCRT:endosomal sorting complex required for
GIXD: grazing incidence X-ray diffraction
HPLC: high-performance liquid chromatography
PLFE: polar lipid fraction E
PPC:pressure perturbation calorimetry
TPL: total polar lipids
XRR: X-ray reflectivity.
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