# The Cell Membrane of _Sulfolobus_ spp.--Homeoviscous Adaption and Biotechnological Applications
## 3 Homeoviscous Adaption
### pH  
Homeoviscous adaption also occurs in response to pH changes. Acidophilic Archaea have adapted to life at low pH while maintaining a near neutral cytoplasmic pH. This is achieved by several mechanisms. The intracellular transmembrane potential is positive, thus repelling protons. Further, an overflow of protons is prevented by efflux systems. The membrane, in general, exhibits low proton permeability. Archaeal membranes comprised of monolayer forming TELs are an example of highly impermeable membranes [80; 88; 89; 90]. As already mentioned in the previous section, pH potentially has an effect on the average cyclization number. [PERSON] et al. showed that by decreasing the cultivation pH, the GDGT cyclization was increased in the crenarchaeon _A. sulfuriretucens_. [86]. This is contradicted by experiments in _Thermoplasma acidophilum_, which indicate a positive correlation between ring index and pH [91]. These results suggest additional factors to be involved in the pH adaption process, such as usages of different polar head groups. Several studies showed that the sugar moieties are faced outwards and their hydroxyl groups presumably aid the repulsion of protons on the cell surface [88; 92]. [PERSON] et al. applied experimental evolution to investigate the thermoacidophily of _S. acidocaldarius_. Over a three-year period employing adaptive evolution, a super-acid-resistant Crenarchaeota (SARC) strain with possible culture conditions of pH 0.8 and 80 \({}^{\circ}\)C was established. Transcriptomics showed upregulation in S-adenosyl methionine (SAM) proteins, which is required for calditol synthesis, and further increased the regulation of the membrane lipid biogenesis. They argue that, according to the thin layer chromatography (TLC) results, and after using lipid-specific dyes, no significant difference in the total lipids and glycolipids was observed. However, upon close inspection of the published images of the TLC plates, it appears that a difference in the total lipid compositions between SARC cultivated at pH 3 and pH 1 can be seen [93]. Sulfolobales contain GDNT in their membrane, in contrast to many other Archaea. Only _Acidianus_ and _Metallospira_, both Crenarchaeota, have been shown to incorporate GDNT, as well, in their cell membrane. SAM protein is required for calditol synthesis, and deletion of the gene locus encoding for this protein showed reduced acid tolerance of the mutant strain. The calditol synthase (_cds_) mutant was still able to produce glycosylated GDGTs, but was not able to survive at pH 1.6. Growth at pH 3.5 was not inhibited when compared to the parental strain, suggesting that the remaining glycolipids suffice as a protection. A hypothesis is that the hexose head group in the _cds_ mutant cells get removed due to the exposure to low pH. This process then results in an increased membrane permeability. Hence, the stronger calditol-linked tetraethers are needed for a higher acid tolerance [39]. Moreover, the presence of potassium ions in the supplied media are supposedly needed for acid tolerance in _S. acidocaldarius_. This leads to the proposal that other factors, such as media composition, also play a role in the coping mechanisms of Sulfolobales towards pH stress [94]. In addition, the presence of proteins, in particular heat shock protein 20 (Hsp20), can prevent stress-induced protein aggregation and membrane destabilization in archaeosomes [95]. Secretory vesicles and thus archaeosomes are unable to perform pH homeostasis, necessary for maintaining a neutral intrinsic pH. This results in a pH of 2 to 3 inside theses vesicles. It is unclear how long it takes for the pH to decline in these vesicles, but according to research conducted with archaeosomes generated from tetraether lipids derived from _T. acidophilum_, the presence of TEL increases the pH resistance measured by the leakage of fluorescent or radioactive entrapped compounds when compared to archaeosomes comprised of mainly DELs [67]. [PERSON] et al.