Certain, but Not All, Tetraether Lipids from the Thermoacidophilic Archaeon Sulfolobus acidocaldarius Can Form Black Lipid Membranes with Remarkable Stability and Exhibiting Mthk Channel Activity with Unusually High Ca2+ Sensitivity

Abstract

Bipolar tetraether lipids (BTL) have been long thought to play a critical role in allowing thermoacidophiles to thrive under extreme conditions. In the present study, we demonstrated that not all BTLs from the thermoacidophilic archaeon Sulfolobus acidocaldarius exhibit the same membrane behaviors. We found that free-standing planar membranes (i.e., black lipid membranes, BLM) made of the polar lipid fraction E (PLFE) isolated from S. acidocaldarius formed over a pinhole on a cellulose acetate partition in a dual-chamber Teflon device exhibited remarkable stability showing a virtually constant capacitance (~28 pF) for at least 11 days. PLFE contains exclusively tetraethers. The dominating hydrophobic core of PLFE lipids is glycerol dialky calditol tetraether (GDNT, ~90%), whereas glycerol dialkyl glycerol tetraether (GDGT) is a minor component (~10%). In sharp contrast, BLM made of BTL extracted from microvesicles (Sa-MVs) released from the same cells exhibited a capacitance between 36 and 39 pF lasting for only 8 h before membrane dielectric breakdown. Lipids in Sa-MVs are also exclusively tetraethers; however, the dominating lipid species in Sa-MVs is GDGT (>99%), not GDNT. The remarkable stability of BLMPLFE can be attributed to strong PLFE–PLFE and PLFE–substrate interactions. In addition, we compare voltage-dependent channel activity of calcium-gated potassium channels (MthK) in BLMPLFE to values recorded in BLMSa-MV. MthK is an ion channel isolated from a methanogenic that has been extensively characterized in diester lipid membranes and has been used as a model for calcium-gated potassium channels. We found that MthK can insert into BLMPLFE and exhibit channel activity, but not in BLMSa-MV. Additionally, the opening/closing of the MthK in BLMPLFE is detectable at calcium concentrations as low as 0.1 mM; conversely, in diester lipid membranes at such a low calcium concentration, no MthK channel activity is detectable. The differential effect of membrane stability and MthK channel activity between BLMPLFE and BLMSa-MV may be attributed to their lipid structural differences and thus their abilities to interact with the substrate and membrane protein. Since Sa-MVs that bud off from the plasma membrane are exclusively tetraether lipids but do not contain the main tetraether lipid component GDNT of the plasma membrane, domain segregation must occur in S. acidocaldarius. The implication of this study is that lipid domain formation is existent and functionally essential in all kinds of cells, but domain formation may be even more prevalent and pronounced in hyperthermophiles, as strong domain formation with distinct membrane behaviors is necessary to counteract randomization due to high growth temperatures while BTL in general make archaea cell membranes stable in high temperature and low pH environments whereas different BTL domains play different functional roles.

Biomolecules under Pressure: Phase Diagrams, Volume Changes, and High Pressure Spectroscopic Techniques

“…Bipolar tetraether liposomes created from the polar lipid fraction of thermoacidophile archaeon Sulfolobus acidocaldarius have been studied by FTIR spectroscopy and SAXS [ 122 ]. Tetraether lipids were also shown to form stable black lipid membranes [ 128 ]. Although the whole phase diagram was not determined, a pressure-induced phase transition was detected from the shift of the symmetric CH 2 stretching mode.…”

Vesicular and Planar Membranes of Archaea Lipids: Unusual Physical Properties and Biomedical Applications

“…However, instability of the BLM made of conventional diester lipids has been a serious drawback [ 111 ]. Archaea tetraether lipids have been used to make BLMs (reviewed in [ 112 , 113 ]). Earlier studies [ 114 , 115 , 116 ] used archaea lipid extract to make free-standing planar membranes over micro-pores in a Teflon partition; however, those membranes lasted only 5–6 h or less, which was not superior to BLM made from diester lipids.…”

“…In contrast, in a recent study, PLFE has been shown to form stable BLM on the micro-pores of polydimethylsiloxane (PDMS) thin films embedded in a printed circuit board-based fluidics or in a glass/silicon microchip, exhibiting a constant electrical impedance for at least 55 h at 11–39 °C [ 117 , 118 ]. In a more recent study, it was demonstrated that BLM PLFE formed over a pinhole on a cellulose acetate partition in a dual-chamber Teflon device exhibited even more remarkable stability, showing a constant capacitance (~28 pF) for at least 11 days [ 113 ]. The extended stability may be related to the substrate material.…”

“…It appears that the chemical structure of the lipid used to make the BLM and the substrate used to hold the BLM both are critical for making stable BLM. In short, the remarkable stability of BLM PLFE on a cellulose acetate partition has been attributed to strong PLFE-PLFE and PLFE-substrate interactions [ 113 ].…”

“…For tetraether lipids isolated from the thermoacidophilic archaeon Sulfolobus solfataricus , the GDNT, not GDGT, component was able to form stable BLM [ 114 ]. While BLMs made of PLFE (with GDNT as the major component) can last for ≥11 days, BLMs made of tetraether lipids isolated from microvesicles (with GDGT as the major component) released from the same archaea cell ( S. acidocaldarius ) can last only for 8 h before membrane dielectric breakdown [ 113 ]. Additionally, pre-treatment of the torus with lipids and the use of a particular organic solvent to make lipid stock solutions also appear to be critical for the formation of stable BLM.…”