Phosphatidylglycerophosphate Methyl Ester Research Articles

Average Conformation of Branched Chain Lipid PGP-Me That Accounts for the Thermal Stability and High-Salinity Resistance of Archaeal Membranes.

The average conformation of the methyl-branched chains of archaeal lipid phosphatidyl glycerophosphate methyl ester (PGP-Me) was examined in a hydrated bilayer membrane based on the 2H nuclear magnetic resonance (NMR) of enantioselectively 2H-labeled compounds that were totally synthesized for the first time in this study. The NMR results in combination with molecular dynamics simulations revealed that the PGP-Me chain appeared to exhibit behavior different from that of typical membrane lipids such as dimyristoylphosphatidylcholine (DMPC). The C-C bonds of the PGP-Me chain adopt alternative parallel and tilted orientations to the membrane normal as opposed to a DMPC chain where all of the C-C bonds tilt in the same way on average. This characteristic orientation causes the intertwining of PGP-Me chains, which plays an important role in the excellent thermal and high-salinity stabilities of archaeal lipid bilayers and membrane proteins.

Stereoselective synthesis of the head group of archaeal phospholipid PGP-Me to investigate bacteriorhodopsin-lipid interactions.

Phosphatidylglycerophosphate methyl ester (PGP-Me), a major constituent of the archaeal purple membrane, is essential for the proper proton-pump activity of bacteriorhodopsin (bR). We carried out the first synthesis of the bisphosphate head group of PGP-Me using H-phosphonate chemistry that led to the production of a simplified PGP-Me analogue with straight alkyl chains. To investigate the role of this head group in the structural and functional integrity of bR, the analogue was used to reconstitute bR into liposomes, in which bR retained the original trimeric structure and light-induced photocycle activity. Enhanced ordering of an alkyl chain of the (2)H-labelled analogue was observed in (2)H NMR spectra upon interaction with bR. These results together suggest that the bisphosphate moiety plays a role in the proper functioning of bR through the lipid-protein interaction.

Deciphering archaeal glycolipids of an extremely halophilic archaeon of the genus Halobellus by MALDI-TOF/MS

Polar membrane lipids of an archaeal microorganism recently isolated from the natural salt lake Fuente de Piedra (Málaga, Spain) have been studied by means of TLC in combination with MALDI-TOF mass spectrometry. The major phospholipids are the ether lipids phosphatidylglycerophosphate methyl ester and phosphatidylglycerosulfate, while phosphatidylglycerol is barely detectable; in addition the bisphosphatidylglycerol (archaeal cardiolipin) has been detected for the first time in a representative of the genus Halobellus. The structures of glycolipids, including a glycosyl-cardiolipin, have been elucidated by post source decay (PSD) mass spectrometry analysis. Besides the monosulfated diglycosyl diphytanylglyceroldiether, two variants of a bis-sulfated diglycosyl diphytanylglyceroldiether have been identified; furthermore the glycosyl-cardiolipin is found to have the same structure of the analogue present in Halorubrum trapanicum and Haloferax volcanii. The role of the abundant sulfated glycolipids in facing high extracellular salinity is discussed.

Novel ether lipid cardiolipins in archaeal membranes of extreme haloalkaliphiles

The lipidome of two extremely haloalkaliphilic archaea, Natronococcus occultus and Natronococcus amylolyticus, has been examined by means of combined thin-layer chromatography and MALDI-TOF/MS analyses. The detailed investigation of lipid profiles has confirmed the presence of i) ether lipid phosphatidylglycerol and phosphatidylglycerophosphate methyl ester as main lipid components, ii) both C20 and C25 isopranoid chains in the lipid core and yielded new findings on membrane lipids of these unusual organisms. Besides some novel minor or trace phospholipids and glycolipids, data indicate the presence of ether lipid cardiolipin variants constituted by different combinations of C20 and C25 isopranoid chains, never before described in archaea. The role of C25 isopranoid chains in the adaptation to high pH gradients in the presence of very high salt concentrations is discussed.

Salarchaeum japonicum gen. nov., sp. nov., an aerobic, extremely halophilic member of the Archaea isolated from commercial salt

Strain YSM-79(T) was isolated from commercial salt made from seawater in Yonaguni island, Okinawa, Japan. The strain is an aerobic, Gram-negative, chemo-organotrophic and extremely halophilic archaeon. Cells are short rods that lyse in distilled water. Growth occurs at 1.5-5.3 M NaCl (optimum 2.5-3.0 M), pH 5.0-8.8 (optimum pH 5.2-6.3) and 20-55 °C (optimum 40 °C). Mg²⁺ is required for growth, with maximum growth at 200-300 mM Mg²⁺. Polar lipid analysis revealed the presence of phosphatidylglycerol, phosphatidylglycerophosphate methyl ester, sulfated diglycosyl diether-1 and five unidentified glycolipids. The G+C content of the DNA was 64 mol%. On the basis of 16S rRNA gene sequence analysis, strain YSM-79(T) was determined to be a member of the family Halobacteriaceae, with the closest related genus being Halobacterium (94 % sequence identity). In addition, the rpoB' gene sequence of strain YSM-79(T) had <88 % sequence similarity to those of other members of the family Halobacteriaceae. The results of phenotypic, chemotaxonomic and phylogenetic analysis suggested that strain YSM-79(T) should be placed in a new genus, Salarchaeum gen. nov., as a representative of Salarchaeum japonicum sp. nov. The type strain is YSM-79(T) ( = JCM 16327(T) = CECT 7563(T)).

Natronoarchaeum mannanilyticum gen. nov., sp. nov., an aerobic, extremely halophilic archaeon isolated from commercial salt

Strain YSM-123(T) was isolated from commercial salt made from Japanese seawater in Niigata prefecture. Optimal NaCl and Mg(2+) concentrations for growth were 4.0-4.5 M and 5 mM, respectively. The isolate was a mesophilic and slightly alkaliphilic haloarchaeon, whose optimal growth temperature and pH were 37 °C and pH 8.0-9.0. Phylogenetic analysis based on 16S rRNA gene sequence analysis suggested that strain YSM-123(T) is a member of the phylogenetic group defined by the family Halobacteriaceae, but there were low similarities to type strains of other genera of this family (≤90 %); for example, Halococcus (similarity <89 %), Halostagnicola (<89 %), Natronolimnobius (<89 %), Halobiforma (<90 %), Haloterrigena (<90 %), Halovivax (<90 %), Natrialba (<90 %), Natronobacterium (<90 %) and Natronococcus (<90 %). The G+C content of the DNA was 63 mol%. Polar lipid analysis revealed the presence of phosphatidylglycerol, phosphatidylglycerophosphate methyl ester, disulfated diglycosyl diether and an unknown glycolipid. On the basis of the data presented, we propose that strain YSM-123(T) should be placed in a new genus and species, Natronoarchaeum mannanilyticum gen. nov., sp. nov. The type strain of Natronoarchaeum mannanilyticum is strain YSM-123(T) (=JCM 16328(T) =CECT 7565(T)).

Halomicrobium katesii sp. nov., an extremely halophilic archaeon

Two extremely halophilic archaea, strains Al-5(T) and K-1, were isolated from Lake Tebenquiche (Atacama Saltern, Chile) and Ezzemoul sabkha (Algeria), respectively. Cells of the two strains were short-rod-shaped and Gram-negative; colonies were orange-pigmented. They grew optimally at 37-40 degrees C and pH 7.0-7.5 in the presence of 25 % (w/v) NaCl. Magnesium was not required. Polar lipid analysis revealed the presence of phosphatidylglycerol and phosphatidylglycerophosphate methyl ester, the absence of phosphatidylglycerosulfate, and the presence of sulfated diglycosyl diether and diether diglycosyl as the sole glycolipids. DNA G+C contents of strains Al-5(T) and K-1 were 52.4 and 52.9 mol% (T(m) method), respectively. 16S rRNA gene sequence comparison with database sequences showed that strains Al-5(T) and K-1 were most closely related to Halomicrobium mukohataei DSM 12286(T) (similarities of 97.5 and 96.9 %, respectively). DNA-DNA hybridization indicated that strains Al-5(T) and K-1 were members of a single species. However, DNA-DNA relatedness to Halomicrobium mukohataei was 55.7+/-2.5 %. A comparative analysis of phenotypic characteristics and DNA-DNA hybridization between the isolates and Halomicrobium mukohataei DSM 12286(T) supported the conclusion that Al-5(T) and K-1 represent a novel species within the genus Halomicrobium, for which the name Halomicrobium katesii sp. nov. is proposed. The type strain is Al-5(T) (=CECT 7257(T)=DSM 19301(T)).

Lipids of the ultra-thin square halophilic archaeonHaloquadratum walsbyi

The lipid composition of the extremely halophilic archaeon Haloquadratum walsbyi was investigated by thin-layer chromatography and electrospray ionization-mass spectrometry. The analysis of neutral lipids showed the presence of vitamin MK-8, squalene, carotene, bacterioruberin and several retinal isomers. The major polar lipids were phosphatidylglycerophosphate methyl ester, phosphatidylglycerosulfate, phosphatidylglycerol and sulfated diglycosyl diether lipid. Among cardiolipins, the tetra-phytanyl or dimeric phospholipids, only traces of bisphosphatidylglycerol were detected. When the cells were exposed to hypotonic medium, no changes in the membrane lipid composition occurred. Distinguishing it from other extreme halophiles of the Halobacteriaceae family, the osmotic stress did not induce the neo-synthesis of cardiolipins in H. walsbyi. The difference may depend on the three-laminar structure of the cell wall, which differs significantly from that of other Haloarchaea.

Lipid composition of integral purple membrane by 1H and 31P NMR

In the purple membrane (PM) of halobacteria, lipids stabilize the trimeric arrangement of bacteriorhodopsin (BR) molecules and mediate the packing of the trimers in a regular crystalline arrangement. To date, the identification and quantification of these lipids has been based either on lipid extraction procedures or structural models. By directly solubilizing PMs from Halobacterium salinarum in aqueous detergent solutions (SDS or Triton X-100), we avoided any separation or modification steps that might modify the lipid composition or even the lipid molecules themselves. Our analysis of integral PM preparations should resolve partially conflicting literature data on the lipid composition of the PM. Using 31P and 1H NMR of detergent-solubilized but otherwise untreated samples, we found two glycolipids and 6.4 +/- 0.1 phospholipids per BR molecule, 4.4 +/- 0.1 of the latter being the phosphatidylglycerophosphate methyl ester. The only glycolipid detected was S-TGD-1. For an additional glycolipid, glycocardiolipin, that was recently identified in lipid extracts, we show that it was produced mainly during the lipid extraction procedure but also was partially dependent on the preparation of the PM suspensions.

Organic solvent tolerance of halophilic archaea, Haloarcula strains: Effects of NaCl concentration on the tolerance and polar lipid composition

Strains of halophilic archaea, Haloarcula vallismortis and two Haloarcula strains OHF-1 and OHF-2, showed high tolerance to organic solvents at high media NaCl concentrations. For example, the lowest log Pow of the solvent which allowed growth (log Pow is the common logarithm of the partition coefficient of a given solvent in a mixture of n-octanol and water) for H. vallismortis was 5.1 at 20% NaCl and 4.4 at 30% NaCl. The solvent tolerance of Haloarcula argentinensis, on the other hand, was not affected by the NaCl concentration. Cells of strains OHF-1 and OHF-2 were of triangular or irregular morphology but became spherical in cultures in NaCl media overlaid with cyclohexane (log Pow=3.4), but returned to the triangular shape when the organic solvent evaporated from the medium. When cells of strains OHF-1, OHF-2, and H. argentinensis were grown in NaCl media in the presence of n-decane, they contained less phosphatidylglycerol and more phosphatidylglycerosulfate and phosphatidylglycerophosphate methyl ester than when grown without added n-decane. When the solvent was removed from the media after cultivation, the levels of these compounds returned to their initial ones.

Natronolimnobius baerhuensis gen. nov., sp. nov. and Natronolimnobius innermongolicus sp. nov., novel haloalkaliphilic archaea isolated from soda lakes in Inner Mongolia, China

Three novel isolates of haloalkaliphilic archaea, strains IHC-005T, IHC-010, and N-1311T, from soda lakes in Inner Mongolia, China, were characterized to elucidate their taxonomic positions. The three strains were aerobic, Gram-negative chemoorganotrophs growing optimally at 37-45 degrees C, pH 9.0-9.5, and 15-20% NaCl. Cells of strains IHC-005T/IHC-010 were motile rods, while those of strain N-1311T were non-motile pleomorphic flats or cocci. The three strains contained diphytanyl and phytanyl-sesterterpanyl diether derivatives of phosphatidylglycerol and phosphatidylglycerophosphate methyl ester. No glycolipids were detected. On phylogenetic analysis of 16S rRNA gene sequences, they formed an independent cluster in the Natro group of the family Halobacteriaceae. Comparison of their morphological, physiological, and biochemical properties, DNA G + C content and 16S rRNA gene sequences, and DNA-DNA hybridization study support the view that strains IHC-005T/IHC-010 and strain N-1311T represent separate species. Therefore, we propose Natronolimnobius baerhuensis gen. nov., sp. nov. for strains IHC-005T (=CGMCC 1.3597T =JCM 12253T)/IHC-010 (=CGMCC 1.3598 = JCM 12254) and Natronolimnobius innermongolicus sp. nov. for N-1311T (=CGMCC 1.2124T =JCM 12255T).

Lipid-protein stoichiometries in a crystalline biological membrane: NMR quantitative analysis of the lipid extract of the purple membrane

The lipid/protein stoichiometries of a naturally crystalline biological membrane, the purple membrane (PM) of Halobacterium salinarum, have been obtained by a combination of (31)P- and (1)H-NMR analyses of the lipid extract. In total, 10 lipid molecules per retinal were found to be present in the PM lipid extract: 2-3 molecules of phosphatidylglycerophosphate methyl ester (PGP-Me), 3 of glycolipid sulfate, 1 of phosphatidylglycerol, 1 of archaeal glycocardiolipin (GlyC), 2 of squalene plus minor amounts of phosphatidylglycerosulfate (PGS) and bisphosphatidylglycerol (archaeal cardiolipin) (BPG) and a negligible amount of vitamin MK8. The novel data of the present study are necessary to identify the lipids in the electron density map, and to shed light on the structural relationships of the lipid and protein components of the PM.

Natronobacterium nitratireducens sp. nov., a aloalkaliphilic archaeon isolated from a soda lake in China.

Two novel haloalkaliphilic archaea, strains C231T and C42, were isolated from a soda lake in China. Cells of the two strains were rod-shaped and gram-negative and colonies were bright red. They required at least 2.5 M NaCl for growth, with an optimum at 3.5 M NaCl, and grew over a pH range from 8.0 to 10.5, with an optimum at pH 8.5. Hypotonic treatment with less than 1.5 M NaCl caused cell lysis. They had similar polar lipid compositions, possessing the diphytanyl (C20:C20) and phytanyl-sesterterpanyl (C20:C25) diether derivatives of phosphatidylglycerol and phosphatidylglycerophosphate methyl ester and a minor phospholipid, PL1. No glycolipids were detected. Comparison of 16S rDNA sequences and morphological features placed them in the genus Natronobacterium. Detailed phenotypic characterization and DNA-DNA hybridization studies revealed that the two strains belong to a new species in the genus Natronobacterium, for which the name Natronobacterium nitratireducens sp. nov. is proposed. The type strain is C231T (= AS 1.1980T = JCM 10879T).