Calditol-linked membrane lipids are required for acid tolerance inSulfolobus acidocaldarius

Archaea have many unique physiological features of which the lipid composition of their cellular membranes is the most striking. Archaeal ether-linked isoprenoidal membranes can occur as bilayers or monolayers, possess diverse polar head groups, and a multiplicity of ring structures in the isoprenoidal cores. These lipid structures are proposed to provide protection from the extreme temperature, pH, salinity, and nutrient-starved conditions that many archaea inhabit. However, many questions remain regarding the synthesis and physiological role of some of the more complex archaeal lipids. In this study, we identify a radicalS-adenosylmethionine (SAM) protein inSulfolobus acidocaldariusrequired for the synthesis of a unique cyclopentyl head group, known as calditol. Calditol-linked glycerol dibiphytanyl glycerol tetraethers (GDGTs) are membrane spanning lipids in which calditol is ether bonded to the glycerol backbone and whose production is restricted to a subset of thermoacidophilic archaea of the Sulfolobales order within the Crenarchaeota phylum. Several studies have focused on the enzymatic mechanism for the synthesis of the calditol moiety, but to date no protein that catalyzes this reaction has been discovered. Phylogenetic analyses of this putative calditol synthase (Cds) reveal the genetic potential for calditol–GDGT synthesis in phyla other than the Crenarchaeota, including the Korarchaeota and Marsarchaeota. In addition, we identify Cds homologs in metagenomes predominantly from acidic ecosystems. Finally, we demonstrate that deletion of calditol synthesis rendersS. acidocaldariussensitive to extremely low pH, indicating that calditol plays a critical role in protecting archaeal cells from acidic stress.

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Phosphinothricin Acetyltransferases Identified UsingIn Vivo,In Vitro, and Bioinformatic Analyses

Applied and Environmental Microbiology ◽

10.1128/aem.02604-16 ◽

2016 ◽

Vol 82 (24) ◽

pp. 7041-7051 ◽

Cited By ~ 5

Author(s):

Chelsey M. VanDrisse ◽

Kristy L. Hentchel ◽

Jorge C. Escalante-Semerena

Keyword(s):

Deinococcus Radiodurans ◽

Phylogenetic Analyses ◽

Critical Role ◽

Metabolic Stress ◽

Physiological Role ◽

Methionine Sulfoximine ◽

Acinetobacter Baylyi ◽

Content Type

ABSTRACTAcetylation of small molecules is widespread in nature, and in some cases, cells use this process to detoxify harmful chemicals.Streptomycesspecies utilize aGcn5N-acetyltransferase (GNAT), known as Bar, to acetylate and detoxify a self-produced toxin,phosphinothricin (PPT), a glutamate analogue. Bar homologues, such as MddA fromSalmonella enterica, acetylate methionine analogues such as methionine sulfoximine (MSX) and methionine sulfone (MSO), but not PPT, even though Bar homologues are annotated as PPT acetyltransferases.S. entericawas used as a heterologous host to determine whether or not putative PPT acetyltransferases from various sources could acetylate PPT, MSX, and MSO.In vitroandin vivoanalyses identified substrates acetylated by putative PPT acetyltransferases fromDeinococcus radiodurans(DR_1057 and DR_1182) andGeobacillus kaustophilus(GK0593 and GK2920).In vivo, synthesis of DR_1182, GK0593, and GK2920 blocked the inhibitory effects of PPT, MSX, and MSO. In contrast, DR_1057 did not detoxify any of the above substrates. Results ofin vitrostudies were consistent with thein vivoresults. In addition, phylogenetic analyses were used to predict the functionality of annotated PPT acetyltransferases inBurkholderia xenovorans,Bacillus subtilis,Staphylococcus aureus,Acinetobacter baylyi, andEscherichia coli.IMPORTANCEThe work reported here provides an example of the use of a heterologous system for the identification of enzyme function. Many members of this superfamily of proteins do not have a known function, or it has been annotated solely on the basis of sequence homology to previously characterized enzymes. The critical role ofGcn5N-acetyltransferases (GNATs) in the modulation of central metabolic processes, and in controlling metabolic stress, necessitates approaches that can reveal their physiological role. The combination ofin vivo,in vitro, and bioinformatics approaches reported here identified GNATs that can acetylate and detoxify phosphinothricin.

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Changes in the distribution of membrane lipids during growth of Thermotoga maritima at different temperatures: Indications for the potential mechanism of biosynthesis of ether-bound diabolic acid (membrane-spanning) lipids

Applied and Environmental Microbiology ◽

10.1128/aem.01763-21 ◽

2021 ◽

Author(s):

Diana X. Sahonero-Canavesi ◽

Laura Villanueva ◽

Nicole J. Bale ◽

Jade Bosviel ◽

Michel Koenen ◽

...

Keyword(s):

Membrane Lipids ◽

Lipid Analysis ◽

Thermotoga Maritima ◽

Lipid Level ◽

Condensation Reaction ◽

Physiological Role ◽

Dicarboxylic Acids ◽

Protein Domain ◽

Alkyl Ether ◽

Membrane Spanning

Membrane-spanning lipids are present in a wide variety of archaea but they are rarely in bacteria. Nevertheless, the (hyper)thermophilic members of the order Thermotogales harbor tetraester, tetraether, and mixed ether/ester membrane-spanning lipids mostly composed of core lipids derived from diabolic acids, C 30, C 32 and C 34 dicarboxylic acids with two adjacent mid-chain methyl substituents. Lipid analysis of Thermotoga maritima across growth phases revealed a decrease of the relative abundance of fatty acids together with an increase of diabolic acids with independence of growth temperature. We also identified isomers of C 30 and C 32 diabolic acids, i.e. dicarboxylic acids with only one methyl group at C-15. Their distribution suggests they are products of the condensation reaction but preferably produced when the length of the acyl chains is not optimal. In comparison with growth at the optimal temperature of 80°C, an increase of glycerol ether-derived lipids was observed at 55°C. Besides, our analysis only detected diabolic acid-containing intact polar lipids with phosphoglycerol (PG) headgroups. Considering these findings, we hypothesize a biosynthetic pathway for the synthesis of membrane-spanning lipids based on PG polar lipid formation, suggesting that the protein catalyzing this process could be a membrane protein. We also identified, by genomic and protein domain analyses, a gene coding for a putative plasmalogen synthase homologue in T. maritima , which is also present in other bacteria producing sn 1-alkyl ether lipids but not plasmalogens, suggesting it could be involved in the conversion of the ester to ether bond in the diabolic acids bound in membrane-spanning lipids. Importance Membrane-spanning lipids are unique compounds found in most archaeal membranes, but they are also present in specific bacterial groups like the Thermotogales. The synthesis and physiological role of membrane-spanning lipids in bacteria represent an evolutionary and biochemical open question that points to the differentiation of the membrane lipids composition. Understanding the formation of membrane-spanning lipids is crucial to solving this question and identifying the enzymatic and biochemical mechanism performing this procedure. In the present work, we found changes at the core lipid level, and we propose that the growth phase drives the biosynthesis of these lipids rather than temperature. Our results identified physiological conditions influencing the membrane-spanning lipids biosynthetic process which can further clarify the pathway leading to the biosynthesis of these compounds.

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Novel archaeal macrocyclic diether core membrane lipids in a methane-derived carbonate crust from a mud volcano in the Sorokin Trough, NE Black Sea

Archaea ◽

10.1155/2003/329175 ◽

2003 ◽

Vol 1 (3) ◽

pp. 165-173 ◽

Cited By ~ 35

Author(s):

Alina Stadnitskaia ◽

Marianne Baas ◽

Michael K. Ivanov ◽

Tjeerd C. E. Van Weering ◽

Jaap S. Sinninghe Damsté

Keyword(s):

Black Sea ◽

Mass Spectra ◽

Membrane Lipids ◽

Mud Volcano ◽

The Novel ◽

Mass Spectral Data ◽

Mass Spectral ◽

Carbonate Formation ◽

Carbonate Crust ◽

Archaeal Lipids

A methane-derived carbonate crust was collected from the recently discovered NIOZ mud volcano in the Sorokin Trough, NE Black Sea during the 11th Training-through-Research cruise of the R/V Professor Logachev. Among several specific bacterial and archaeal membrane lipids present in this crust, two novel macrocyclic diphytanyl glycerol diethers, containing one or two cyclopentane rings, were detected. Their structures were tentatively identified based on the interpretation of mass spectra, comparison with previously reported mass spectral data, and a hydrogenation experiment. This macrocyclic type of archaeal core membrane diether lipid has so far been identified only in the deep-sea hydrothermal vent methanogenMethanococcus jannaschii. Here, we provide the first evidence that these macrocyclic diethers can also contain internal cyclopentane rings. The molecular structure of the novel diethers resembles that of dibiphytanyl tetraethers in which biphytane chains, containing one and two pentacyclic rings, also occur. Such tetraethers were abundant in the crust. Compound-specific isotope measurements revealed δ13C values of –104 to –111‰ for these new archaeal lipids, indicating that they are derived from methanotrophic archaea acting within anaerobic methane-oxidizing consortia, which subsequently induce authigenic carbonate formation.

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The Hinge-Helix 1 Region of Peroxisome Proliferator-Activated Receptor γ1 (PPARγ1) Mediates Interaction with Extracellular Signal-Regulated Kinase 5 and PPARγ1 Transcriptional Activation: Involvement in Flow-Induced PPARγ Activation in Endothelial Cells

Molecular and Cellular Biology ◽

10.1128/mcb.24.19.8691-8704.2004 ◽

2004 ◽

Vol 24 (19) ◽

pp. 8691-8704 ◽

Cited By ~ 91

Author(s):

Masashi Akaike ◽

Wenyi Che ◽

Nicole-Lerner Marmarosh ◽

Shinsuke Ohta ◽

Masaki Osawa ◽

...

Keyword(s):

Endothelial Cells ◽

Adhesion Molecule ◽

Critical Role ◽

Physiological Role ◽

Cellular Adhesion ◽

Dominant Negative ◽

Peroxisome Proliferator ◽

Extracellular Signal Regulated Kinase ◽

Extracellular Signal ◽

Cellular Adhesion Molecule

ABSTRACT Peroxisome proliferator-activated receptors (PPAR) are ligand-activated transcription factors that form a subfamily of the nuclear receptor gene family. Since both flow and PPARγ have atheroprotective effects and extracellular signal-regulated kinase 5 (ERK5) kinase activity is significantly increased by flow, we investigated whether ERK5 kinase regulates PPARγ activity. We found that activation of ERK5 induced PPARγ1 activation in endothelial cells (ECs). However, we could not detect PPARγ phosphorylation by incubation with activated ERK5 in vitro, in contrast to ERK1/2 and JNK, suggesting a role for ERK5 as a scaffold. Endogenous PPARγ1 was coimmunoprecipitated with endogenous ERK5 in ECs. By mammalian two-hybrid analysis, we found that PPARγ1 associated with ERK5a at the hinge-helix 1 region of PPARγ1. Expressing a hinge-helix 1 region PPARγ1 fragment disrupted the ERK5a-PPARγ1 interaction, suggesting a critical role for hinge-helix 1 region of PPARγ in the ERK5-PPARγ interaction. Flow increased ERK5 and PPARγ1 activation, and the hinge-helix 1 region of the PPARγ1 fragment and dominant negative MEK5β significantly reduced flow-induced PPARγ activation. The dominant negative MEK5β also prevented flow-mediated inhibition of tumor necrosis factor alpha-mediated NF-κB activation and adhesion molecule expression, including vascular cellular adhesion molecule 1 and E-selectin, indicating a physiological role for ERK5 and PPARγ activation in flow-mediated antiinflammatory effects. We also found that ERK5 kinase activation was required, likely by inducing a conformational change in the NH2-terminal region of ERK5 that prevented association of ERK5 and PPARγ1. Furthermore, association of ERK5a and PPARγ1 disrupted the interaction of SMRT and PPARγ1, thereby inducing PPARγ activation. These data suggest that ERK5 mediates flow- and ligand-induced PPARγ activation via the interaction of ERK5 with the hinge-helix 1 region of PPARγ.

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Constitutive activity of dopamine receptor type 1 (D1R) increases CaV2.2 currents in PFC neurons

The Journal of General Physiology ◽

10.1085/jgp.201912492 ◽

2020 ◽

Vol 152 (5) ◽

Author(s):

Clara Inés McCarthy ◽

Cambria Chou-Freed ◽

Silvia Susana Rodríguez ◽

Agustín Yaneff ◽

Carlos Davio ◽

...

Keyword(s):

Dopamine Receptor ◽

Pyramidal Neurons ◽

Critical Role ◽

Intraperitoneal Administration ◽

Brain Slices ◽

Physiological Role ◽

Receptor Type ◽

Constitutive Activity ◽

Medial Pfc

Alterations in dopamine receptor type 1 (D1R) density are associated with cognitive deficits of aging and schizophrenia. In the prefrontal cortex (PFC), D1R plays a critical role in the regulation of working memory, which is impaired in these cognitive deficit states, but the cellular events triggered by changes in D1R expression remain unknown. A previous report demonstrated that interaction between voltage-gated calcium channel type 2.2 (CaV2.2) and D1R stimulates CaV2.2 postsynaptic surface location in medial PFC pyramidal neurons. Here, we show that in addition to the occurrence of the physical receptor-channel interaction, constitutive D1R activity mediates up-regulation of functional CaV2.2 surface density. We performed patch-clamp experiments on transfected HEK293T cells and wild-type C57BL/6 mouse brain slices, as well as imaging experiments and cAMP measurements. We found that D1R coexpression led to ∼60% increase in CaV2.2 currents in HEK293T cells. This effect was occluded by preincubation with a D1/D5R inverse agonist, chlorpromazine, and by replacing D1R with a D1R mutant lacking constitutive activity. Moreover, D1R-induced increase in CaV2.2 currents required basally active Gs protein, as well as D1R-CaV2.2 interaction. In mice, intraperitoneal administration of chlorpromazine reduced native CaV currents’ sensitivity to ω-conotoxin-GVIA and their size by ∼49% in layer V/VI pyramidal neurons from medial PFC, indicating a selective effect on CaV2.2. Additionally, we found that reducing D1/D5R constitutive activity correlates with a decrease in the agonist-induced D1/D5R inhibitory effect on native CaV currents. Our results could be interpreted as a stimulatory effect of D1R constitutive activity on the number of CaV2.2 channels available for dopamine-mediated modulation. Our results contribute to the understanding of the physiological role of D1R constitutive activity and may explain the noncanonical postsynaptic distribution of functional CaV2.2 in PFC neurons.

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The Halophyte Halostachys caspica AP2/ERF Transcription Factor HcTOE3 Positively Regulates Freezing Tolerance in Arabidopsis

Frontiers in Plant Science ◽

10.3389/fpls.2021.638788 ◽

2021 ◽

Vol 12 ◽

Author(s):

Fangliu Yin ◽

Youling Zeng ◽

Jieyun Ji ◽

Pengju Wang ◽

Yufang Zhang ◽

...

Keyword(s):

Transcription Factor ◽

Abscisic Acid ◽

Freezing Tolerance ◽

Critical Role ◽

Extreme Temperature ◽

Freezing Stress ◽

Antioxidant Enzyme Activities ◽

Transcription Factor Gene ◽

Factor Gene ◽

Halostachys Caspica

The APETALA2 (AP2) and ethylene-responsive element-binding factor (ERF) gene family is one of the largest plant-specific transcription factor gene families, which plays a critical role in plant development and evolution, as well as response to various stresses. The TARGET OF EAT3 (TOE3) gene is derived from Halostachys caspica and belongs to the AP2 subfamily with two AP2 DNA-binding domains. Currently, AP2 family mainly plays crucial roles in plant growth and evolution, yet there are few reports about the role of AP2 in abiotic stress tolerance. Here, we report HcTOE3, a new cold-regulated transcription factor gene, which has an important contribution to freezing tolerance. The main results showed that the expression of HcTOE3 in the H. caspica assimilating branches was strongly induced by different abiotic stresses, including high salinity, drought, and extreme temperature (heat, chilling, and freezing), as well as abscisic acid and methyl viologen treatments. Overexpressing HcTOE3 gene (OE) induced transgenic Arabidopsis plant tolerance to freezing stress. Under freezing treatment, the OE lines showed lower content of malondialdehyde and electrolyte leakage and less accumulation of reactive oxygen species compared with the wild type. However, the survival rates, antioxidant enzyme activities, and contents of osmotic adjustment substance proline were enhanced in transgenic plants. Additionally, the OE lines increased freezing tolerance by up-regulating the transcription level of cold responsive genes (CBF1, CBF2, COR15, COR47, KIN1, and RD29A) and abscisic acid signal transduction pathway genes (ABI1, ABI2, ABI5, and RAB18). Our results suggested that HcTOE3 positively regulated freezing stress and has a great potential as a candidate gene to improve plant freezing tolerance.

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A Critical Role for Natural Killer T Cells in Immunosurveillance of Methylcholanthrene-induced Sarcomas

Journal of Experimental Medicine ◽

10.1084/jem.20020092 ◽

2002 ◽

Vol 196 (1) ◽

pp. 119-127 ◽

Cited By ~ 237

Author(s):

Nadine Y. Crowe ◽

Mark J. Smyth ◽

Dale I. Godfrey

Keyword(s):

T Cells ◽

T Cell ◽

Natural Killer ◽

Critical Role ◽

Physiological Role ◽

Cell Receptor ◽

Interferon Γ ◽

Tumor Rejection ◽

Nk T Cells ◽

Nk T Cell

Natural killer (NK) T cells initiate potent antitumor responses when stimulated by exogenous factors such as interleukin (IL)-12 or α-galactosylceramide (α-GalCer), however, it is not clear whether this reflects a physiological role for these cells in tumor immunity. Through adoptive transfer of NK T cells from wild-type to NK T cell–deficient (T cell receptor [TCR] Jα281−/−) mice, we demonstrate a critical role for NK T cells in immunosurveillance of methylcholanthrene (MCA)-induced fibrosarcomas, in the absence of exogenous stimulatory factors. Using the same approach with gene-targeted and/or antibody-depleted donor or recipient mice, we have shown that this effect depends on CD1d recognition and requires the additional involvement of both NK and CD8+ T cells. Interferon-γ production by both NK T cells and downstream, non-NK T cells, is essential for protection, and perforin production by effector cells, but not NK T cells, is also critical. The protective mechanisms in this more physiologically relevant system are distinct from those associated with α-GalCer–induced, NK T cell–mediated, tumor rejection. This study demonstrates that, in addition to their importance in tumor immunotherapy induced by IL-12 or α-GalCer, NK T cells can play a critical role in tumor immunosurveillance, at least against MCA-induced sarcomas, in the absence of exogenous stimulation.

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GDGT cyclization proteins identify the dominant archaeal sources of tetraether lipids in the ocean

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1909306116 ◽

2019 ◽

Vol 116 (45) ◽

pp. 22505-22511 ◽

Cited By ~ 14

Author(s):

Zhirui Zeng ◽

Xiao-Lei Liu ◽

Kristen R. Farley ◽

Jeremy H. Wei ◽

William W. Metcalf ◽

...

Keyword(s):

Physiological Response ◽

Phylogenetic Analyses ◽

Radical Mechanism ◽

Comprehensive Understanding ◽

Physiological Factors ◽

The Core ◽

Free Radical Mechanism ◽

Tetraether Lipids ◽

Membrane Spanning ◽

Archaeal Communities

Glycerol dibiphytanyl glycerol tetraethers (GDGTs) are distinctive archaeal membrane-spanning lipids with up to eight cyclopentane rings and/or one cyclohexane ring. The number of rings added to the GDGT core structure can vary as a function of environmental conditions, such as changes in growth temperature. This physiological response enables cyclic GDGTs preserved in sediments to be employed as proxies for reconstructing past global and regional temperatures and to provide fundamental insights into ancient climate variability. Yet, confidence in GDGT-based paleotemperature proxies is hindered by uncertainty concerning the archaeal communities contributing to GDGT pools in modern environments and ambiguity in the environmental and physiological factors that affect GDGT cyclization in extant archaea. To properly constrain these uncertainties, a comprehensive understanding of GDGT biosynthesis is required. Here, we identify 2 GDGT ring synthases, GrsA and GrsB, essential for GDGT ring formation in Sulfolobus acidocaldarius. Both proteins are radical S-adenosylmethionine proteins, indicating that GDGT cyclization occurs through a free radical mechanism. In addition, we demonstrate that GrsA introduces rings specifically at the C-7 position of the core GDGT lipid, while GrsB cyclizes at the C-3 position, suggesting that cyclization patterns are differentially controlled by 2 separate enzymes and potentially influenced by distinct environmental factors. Finally, phylogenetic analyses of the Grs proteins reveal that marine Thaumarchaeota, and not Euryarchaeota, are the dominant source of cyclized GDGTs in open ocean settings, addressing a major source of uncertainty in GDGT-based paleotemperature proxy applications.

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Expression of tachykinin receptors (tacr1a and tacr1b) in zebrafish: influence of cocaine and opioid receptors

Journal of Molecular Endocrinology ◽

10.1530/jme-12-0199 ◽

2012 ◽

Vol 50 (2) ◽

pp. 115-129 ◽

Cited By ~ 9

Author(s):

Roger López-Bellido ◽

Katherine Barreto-Valer ◽

Raquel E Rodríguez

Keyword(s):

Embryonic Development ◽

Opioid Receptors ◽

Phylogenetic Analyses ◽

Critical Role ◽

Cocaine Exposure ◽

Peripheral Tissues ◽

Tachykinin Receptors ◽

Hek 293 ◽

293 Cells ◽

Gene Study

Opioid and tachykinin receptors (TACRs) are closely related in addiction and pain processes. In zebrafish, opioid receptors have been cloned and characterized both biochemically and pharmacologically. However, thetacr1gene has not yet been described in zebrafish. The aim of this research was to identify thetacr1gene, study the effects of cocaine ontacr1, and analyze the interaction betweentacr1and opioid receptors. We have identified a duplicate oftacr1gene in zebrafish, designated astacr1aandtacr1b. Phylogenetic analyses revealed an alignment of these receptors in the Tacr1 fish cluster, with a clear distinction from other TACR1s of amphibians, birds, and mammals. Our qPCR results showed thattacr1aandtacr1bmRNAs are expressed during embryonic development. Whole-mountin situhybridization showedtacr1expression in the CNS and in the peripheral tissues. Cocaine (1.5 μM) induced an upregulation oftacr1aandtacr1bat 24 and 48 h post-fertilization (hpf; except fortacr1aat 48 hpf, which was downregulated). By contrast, HEK-293 cells transfected withtacr1aandtacr1band exposed to cocaine showed a downregulation oftacr1s. The knockdown of ZfDOR2 and ZfMOR, opioid receptors, induced a down- and upregulation oftacr1aandtacr1brespectively. In conclusion,tacr1aandtacr1bin zebrafish are widely expressed throughout the CNS and peripherally, suggesting a critical role of thesetacr1sduring embryogenesis.tacr1aandtacr1bmRNA expression is altered by cocaine exposure and by the knockdown of opioid receptors. Thus, zebrafish can provide clues for a better understanding of the relationship between tachykinin and opioid receptors in pain and addiction during embryonic development.

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The Cell Membrane of Sulfolobus spp.—Homeoviscous Adaption and Biotechnological Applications

International Journal of Molecular Sciences ◽

10.3390/ijms21113935 ◽

2020 ◽

Vol 21 (11) ◽

pp. 3935 ◽

Cited By ~ 1

Author(s):

Kerstin Rastädter ◽

David J. Wurm ◽

Oliver Spadiut ◽

Julian Quehenberger

Keyword(s):

Cell Membrane ◽

Environmental Changes ◽

Specific Growth ◽

Membrane Lipids ◽

Environmental Parameters ◽

Coping Mechanism ◽

Biotechnological Applications ◽

Head Groups ◽

Tetraether Lipids ◽

Archaeal Lipids

The microbial cell membrane is affected by physicochemical parameters, such as temperature and pH, but also by the specific growth rate of the host organism. Homeoviscous adaption describes the process of maintaining membrane fluidity and permeability throughout these environmental changes. Archaea, and thereby, Sulfolobus spp. exhibit a unique lipid composition of ether lipids, which are altered in regard to the ratio of diether to tetraether lipids, number of cyclopentane rings and type of head groups, as a coping mechanism against environmental changes. The main biotechnological application of the membrane lipids of Sulfolobus spp. are so called archaeosomes. Archaeosomes are liposomes which are fully or partly generated from archaeal lipids and harbor the potential to be used as drug delivery systems for vaccines, proteins, peptides and nucleic acids. This review summarizes the influence of environmental parameters on the cell membrane of Sulfolobus spp. and the biotechnological applications of their membrane lipids.

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