Stereoselective synthesis of perdeuterated phytanic acid, its phospholipid derivatives and their formation into lipid model membranes for neutron reflectivity studies.

We describe a straightforward method, for synthesis of large scale (gram quantities) of highly deuterated phytanic acid from commercially available phytol while preserving the stereochemistry around the chiral centres. The subsequent synthesis of tail-deuterated analogues of the archeabacterial membrane lipids 1,2-di(3RS,7R,11R-phytanyl)-sn-glycero-3-phosphocholine (DPEPC) and 1,2-di(3RS,7R,11R-phytanoyl)-sn-glycero-3-phosphocholine (DPhyPC) from perdeuterated phytanic acid is also described. Both lipids were employed in construction of two different model membranes, namely Langmuir monolayers and a tethered bilayer membrane (TBM) on a solid substrate, characterised by pressure area isotherm and neutron reflectometry techniques. At 10 mN/m pressure the head-group thickness of both monolayers was similar while the thickness of the tail region was significantly larger for tail-deuterated DPhyPC, which was evident from a smaller area per molecule. At 20 mN/m the thickness of the head and tail regions in both lipids was comparable, yet the area per molecule of tail-deuterated DPhyPC was 10% smaller than tail-deuterated DPEPC. In the TBM bilayer model membrane, the thickness of the lipid tails in both inner and outer leaflets was 8.2 Å, giving a total of 16.4 Å. Deuteration enabled unambiguous determination of the relative proportion of the hydrogenous tether, phospholipid and subphase.

Synthesis of deuterated [D32 ]oleic acid and its phospholipid derivative [D64 ]dioleoyl-sn-glycero-3-phosphocholine.

Oleic acid and its phospholipid derivatives are fundamental to the structure and function of cellular membranes. As a result, there has been increasing interest in the availability of their deuterated forms for many nuclear magnetic resonance, infrared, mass spectroscopy and neutron scattering studies. Here, we present for the first time a straightforward, large-scale (gram quantities) synthesis of highly deuterated [D32 ]oleic acid by using multiple, yet simple and high yielding reactions. The precursors for the synthesis of [D32 ]oleic acid are [D14 ]azelaic acid and [D17 ]nonanoic acid, which were obtained by complete deuteration (>98% D) of their (1) H forms by using metal catalysed hydrothermal H/D exchange reactions. The oleic acid was produced with ca. 94% D isotopic purity and with no contamination by the trans-isomer (elaidic acid). The subsequent synthesis of [D64 ]dioleoyl-sn-glycero-3-phosphocholine from [D32 ]oleic acid is also described.

Molecular basis for fungicidal action of neothyonidioside, a triterpene glycoside from the sea cucumber, Australostichopus mollis.

Neothyonidioside is a triterpene glycoside (TG) isolated from the sea cucumber, Australostichopus mollis, that is potently cytotoxic to S. cerevisiae, but does not permeabilize cellular membranes. We mutagenized S. cerevisiae and isolated a neothionidioside-resistant (neo(R)) strain. Using synthetic genetic array mapping and sequencing, we identified NCP1 as the resistance locus. Quantitative HPLC revealed that neo(R)/ncp1 mutants have reduced ergosterol content. Ergosterol added to growth media reversed toxicity, demonstrating that neothionidioside binds directly to ergosterol, similar to the polyene natamycin. Ergosterol synthesis inhibitors ketoconazole and atorvastatin conferred resistance to neothionidioside in a dose-dependent manner showing that a threshold ergosterol concentration is required for toxicity. A genome-wide screen of deletion mutants against neothionidioside revealed hypersensitivity of many of the component genes in the ESCRT complexes relating to multivesicular body formation. Confocal microscopy of cells stained with a vital dye showed blockage at this step. Thus, we propose neothionidioside may affect membrane curvature and fusion capability in the endosome-vacuole pathway.

Mollisosides A, B1, and B2: minor triterpene glycosides from the New Zealand and South Australian sea cucumber Australostichopus mollis.

Three new monosulfated triterpene glycosides, mollisosides A (2), B(1) (3), and B(2) (4), have been isolated from the sea cucumber Australostichopus mollis. Their structures were determined by NMR and mass spectra. The presence of sulfated glycosides in sea cucumbers belonging to the family Stichopodidae is uncommon.