Acyl Chains Of Membrane Phospholipids Research Articles

Divergent unsaturated fatty acid synthesis in two highly related model pseudomonads.

The genomes of the best-studied pseudomonads, Pseudomonas aeruginosa and Pseudomonas putida, which share 85% of the predicted coding regions, contain a fabA fabB operon (demonstrated in P. aeruginosa, putative in P. putida). The enzymes encoded by the fabA and fabB genes catalyze the introduction of a double bond into a 10-carbon precursor which is elongated to the 16:1Δ9 and 18:1Δ11 unsaturated fatty acyl chains required for functional membrane phospholipids. A detailed analysis of transcription of the P. putida fabA fabB gene cluster showed that fabA and fabB constitute an operon and disclosed an unexpected and essential fabB promoter located within the fabA coding sequence. Inactivation of the fabA fabB operon fails to halt the growth of P. aeruginosa PAO1 but blocks growth of P. putida F1 unless an exogenous unsaturated fatty acid is provided. We report that the asymmetry between these two species is due to the P. aeruginosa PAO1 desA gene which encodes a fatty acid desaturase that introduces double bonds into the 16-carbon acyl chains of membrane phospholipids. Although P. putida F1 encodes a putative DesA homolog that is 84% identical to the P. aeruginosa PAO1, the protein fails to provide sufficient unsaturated fatty acid synthesis for growth when the FabA FabB pathway is inactivated. We report that the P. putida F1 DesA homolog can functionally replace the P. aeruginosa DesA. Hence, the defect in P. putida F1 desaturation is not due to a defective P. putida F1 DesA protein but probably to a weakly active component of the electron transfer process.

Unraveling the Gordian knot of insulin resistance in type 2 diabetes mellitus

<abstract> <p>β-cells play an important role in unraveling the Gordian knot of insulin resistance in type 2 diabetes. Firstly, a key feature of the etiology of type 2 diabetes, which appears in the prediabetic phase, is a reduction in the unsaturation index (number of cis carbon-carbon double bonds per 100 acyl chains of membrane phospholipids) compared to healthy controls, which leads to a lower rate of transmembrane glucose transport, and consequently causes reduced glucose effectiveness. Thus, the amount of glucose entering the β-cell via glucose transporter-2 reduces insulin production, leading to reduced insulin sensitivity. Secondly, after synthesis of monomer insulin, six monomer insulin molecules can join together in the presence of zinc ions. The mature hexamers are packed inside mature intracellular vesicles that are transported to the β-cell plasma membrane. Fusion of the intracellular vesicle membrane with the β-cell plasma membrane creates a fusion pore that allows expulsion of monomer insulin molecules into the blood circulation. The large dimensions of the monomer insulin molecule (30 Å wide and 35 Å high) require substantial flexibility of the vesicle membrane and the β-cell plasma membrane. Reduction in the unsaturation indexes leads to a lower rate of insulin transport into the blood circulation, which results in a further decrease in insulin sensitivity.</p> <p>That brings us to a crucial point. The conceit behind the term “insulin resistance” is wrong. It suggests that cells do not respond well to insulin, but the fact is that this term ignores the essential reduction, compared to the plasma glucose concentration, in the amount of glucose entering the β-cell via glucose transporter-2, resulting in reduced insulin production. We now know that an increase in glucose effectiveness, powered by an increased unsaturation index, reframes fundamentally the mechanisms that participate in the glucose homeostasis during type 2 diabetes mellitus.</p> </abstract>

Fundamentals about onset and progressive disease character of type 2 diabetes mellitus.

ResearchGate is a world wide web for scientists and researchers to share papers, ask and answer questions, and find collaborators. As one of the more than 15 million members, the author uploads research output and reads and responds to some of the questions raised, which are related to type 2 diabetes. In that way, he noticed a serious gap of knowledge of this disease among medical professionals over recent decades. The main aim of the current study is to remedy this situation through providing a comprehensive review on recent developments in biochemistry and molecular biology, which can be helpful for the scientific understanding of the molecular nature of type 2 diabetes. To fill up the shortcomings in the curricula of medical education, and to familiarize the medical community with a new concept of the onset of type 2 diabetes, items are discussed like: Insulin resistance, glucose effectiveness, insulin sensitivity, cell membranes, membrane flexibility, unsaturation index (UI; number of carbon-carbon double bonds per 100 acyl chains of membrane phospholipids), slow-down principle, effects of temperature acclimation on phospholipid membrane composition, free fatty acids, energy transport, onset of type 2 diabetes, metformin, and exercise. Based on the reviewed data, a new model is presented with proposed steps in the development of type 2 diabetes, a disease arising as a result of a hypothetical hereditary anomaly, which causes hyperthermia in and around the mitochondria. Hyperthermia is counterbalanced by the slow-down principle, which lowers the amount of carbon-carbon double bonds of membrane phospholipid acyl chains. The accompanying reduction in the UI lowers membrane flexibility, promotes a redistribution of the lateral pressure in cell membranes, and thereby reduces the glucose transporter protein pore diameter of the transmembrane glucose transport channel of all Class I GLUT proteins. These events will set up a reduction in transmembrane glucose transport. So, a new blood glucose regulation system, effective in type 2 diabetes and its prediabetic phase, is based on variations in the acyl composition of phospholipids and operates independent of changes in insulin and glucose concentration. UI assessment is currently arising as a promising analytical technology for a membrane flexibility analysis. An increase in mitochondrial heat production plays a pivotal role in the existence of this regulation system.

An N-terminal di-proline motif is essential for fatty acid–dependent degradation of Δ9-desaturase in Drosophila

The Δ9-fatty acid desaturase introduces a double bond at the Δ9 position of the acyl moiety of acyl-CoA and regulates the cellular levels of unsaturated fatty acids. However, it is unclear how Δ9-desaturase expression is regulated in response to changes in the levels of fatty acid desaturation. In this study, we found that the degradation of DESAT1, the sole Δ9-desaturase in the Drosophila cell line S2, was significantly enhanced when the amounts of unsaturated acyl chains of membrane phospholipids were increased by supplementation with unsaturated fatty acids, such as oleic and linoleic acids. In contrast, inhibition of DESAT1 activity remarkably suppressed its degradation. Of note, removal of the DESAT1 N-terminal domain abolished the responsiveness of DESAT1 degradation to the level of fatty acid unsaturation. Further truncation and amino acid replacement analyses revealed that two sequential prolines, the second and third residues of DESAT1, were responsible for the unsaturated fatty acid-dependent degradation. Although degradation of mouse stearoyl-CoA desaturase 1 (SCD1) was unaffected by changes in fatty acid unsaturation, introduction of the N-terminal sequential proline residues into SCD1 conferred responsiveness to unsaturated fatty acid-dependent degradation. Furthermore, we also found that the Ca2+-dependent cysteine protease calpain is involved in the sequential proline-dependent degradation of DESAT1. In light of these findings, we designated the sequential prolines at the second and third positions of DESAT1 as a "di-proline motif," which plays a crucial role in the regulation of Δ9-desaturase expression in response to changes in the level of cellular unsaturated fatty acids.

Lipid Composition of Cell Membranes and Its Relevance in Type 2 Diabetes Mellitus

Identifying the causative relationship between the fatty acid composition of cell membranes and type 2 diabetes mellitus fundamentally contributes to the understanding of the basic pathophysiological mechanisms of the disease. Important outcomes of the reviewed studies appear to support the hypotheses that the flexibility of a membrane determined by the ratio of (poly)unsaturated to saturated fatty acyl chains of its phospholipids influences the effectiveness of glucose transport by insulin-independent glucose transporters (GLUTs) and the insulin-dependent GLUT4, and from the prediabetic stage on a shift from unsaturated towards saturated fatty acyl chains of membrane phospholipids directly induces a decrease in glucose effectiveness and insulin sensitivity. In addition, it has become evident that a concomitant increase in stiffness of both plasma and erythrocyte membranes may decrease the microcirculatory flow, leading ultimately to tissue hypoxia, insufficient tissue nutrition, and diabetes-specific microvascular pathology. As to the etiology of type 2 diabetes mellitus, a revised hypothesis that attempts to accommodate the reviewed findings is presented.

Membrane-modifying effect of taurine

The effect of taurine on the membrane-bound processes in rat erythrocytes and peritoneal mast cells was studied. By EPR method using spin probe 5-DS a significant decrease in the order parameter S of erythrocyte membrane phospholipid acyl chains in in vitro experiments after incubation with taurine (10 mM, 1 h) was shown. The increase of erythrocyte membrane response to peroxide hemolysis was also observed, that was apparently induced by decrease of membrane viscosity as a result of lessening the solidity of the erythrocyte membrane phospholipid packing. The different effects of taurine on ionophore A23187 and compound 48/80-stimulated functional activity of the peritoneal rat mast cells by various modes of administration (peroral, intraperitoneal, intramuscular) were shown. This makes it possible to conclude that the mechanism of taurine activity at the organism level seems to be a mediated process and systemic by nature.

Interaction with Lipid II Induces Conformational Changes in Bovicin HC5 Structure

Bovicin HC5 is a lantibiotic produced by Streptococcus bovis HC5 that targets the cell wall precursor lipid II. An understanding of the modes of action against target bacteria can help broadening the clinical applicability of lantibiotics in human and veterinary medicine. In this study, the interaction of bovicin HC5 with lipid II was examined using tryptophan fluorescence and circular dichroism spectroscopy with model membrane systems that do or do not allow pore formation by bovicin HC5. In the presence of lipid II, a blue-shift of 12 nm could be observed for the fluorescence emission maximum of the tryptophan residue for all of the membrane systems tested. This change in fluorescence emission was paralleled by a decrease in accessibility toward acrylamide and phospholipids carrying a spin-label at the acyl chain; the tryptophan residue of bovicin HC5 was located near the twelfth position of the membrane phospholipid acyl chains. Moreover, the binding of lipid II by bovicin HC5 induced remarkable conformational changes in the bovicin HC5 structure. The interaction of bovicin HC5 with lipid II was highly stable even at pH 2.0. These results indicate that bovicin HC5 interacts directly with lipid II and that the topology of this interaction changes under different conditions, which is relevant for the biological activity of the peptide. To our knowledge, bovicin HC5 is the only bacteriocin described thus far that is able to interact with its target in extreme pH values, and this fact might be related to its unique structure and stability.

THE STRUCTURAL BIOLOGY OF TYPE II FATTY ACID BIOSYNTHESIS

The type II fatty acid synthetic pathway is the principal route for the production of membrane phospholipid acyl chains in bacteria and plants. The reaction sequence is carried out by a series of individual soluble proteins that are each encoded by a discrete gene, and the pathway intermediates are shuttled between the enzymes as thioesters of an acyl carrier protein. The Escherichia coli system is the paradigm for the study of this system, and high-resolution X-ray and/or NMR structures of representative members of every enzyme in the type II pathway are now available. The structural biology of these proteins reveals the specific three-dimensional features of the enzymes that explain substrate recognition, chain length specificity, and the catalytic mechanisms that define their roles in producing the multitude of products generated by the type II system. These structures are also a valuable resource to guide antibacterial drug discovery.

Cholesterol in health and disease.

Imagine if an excess amount of a critical, life-sustaining molecule like ATP were, by a perverse series of events involving the lifestyle of modern humans, causally related to a major human disease. The thought of “ATP” being synonymous with poor health and poor living in the minds of the lay public and press, even of health care providers, would be difficult for any selfrespecting scientist to accept. So one might view the biomedical history of cholesterol — indeed, this history might be seen as even stranger than the hypothetical ATP scenario, given that evolution has devoted close to 100 genes to the synthesis, transport, metabolism, and regulation of cholesterol. This structurally fascinating lipid is utterly essential to the proper functioning of cells and organisms. Cholesterol, cholesterol metabolites, and immediate biosynthetic precursors of cholesterol play essential roles in cellular membrane physiology, dietary nutrient absorption, reproductive biology, stress responses, salt and water balance, and calcium metabolism. Indeed, many of the articles in this Perspective series are devoted to the normal physiology of cholesterol. Still, there is little doubt that the disease process responsible for the leading cause of death in the industrialized world — atherosclerosis — is a disorder in which an excess of cholesterol is a major culprit. How did evolution come up with a molecule that is critical for so many aspects of normal physiology, and what went “wrong”? The physiology of cholesterol Cholesterol and cellular membranes: an evolutionary perspective. As organisms became more complex, cells required membranes that would provide the proper conformational environment for a wide variety of integral membrane proteins, such as channels, transporters, and enzymes. Moreover, the cells of these advanced organisms required sophisticated signaling machinery, and this machinery would have to be organized as multiprotein complexes in focal, nonhomogenous areas of cellular membranes. Put simply, these requirements were met by focally increasing the “stiffness” or viscosity of phospholipid bilayers. In theory, this goal could be achieved by increasing the degree of saturation of the fatty acyl moieties of membrane phospholipids. However, unsaturated fatty acids in these phospholipids, particularly in the sn-2 position, are needed for a wide variety of cellular signaling functions, and so an exogenous “stiffening factor” was needed. This factor would have to be able to pack tightly with the long saturated and unsaturated fatty acyl chains of membrane phospholipids through van der Waals interactions. This would require a long, flat, and properly shaped hydrophobic molecule, accompanied by additional features (see below) to help further stabilize this interaction. To meet these needs, nature began with molecules, probably originating from prebiotic anaerobic times, that were formed by the sequential condensation of the two-carbon acetate molecule. Prokaryotes evolved enzymes to synthesize more complex linear molecules from these precursors (e.g., carotenoids), which were able to meet the membrane-organizing requirements of most of these primitive organisms. These primitive molecules, however, lacked the proper shape, rigidity, and amphipathic nature to properly organize the mem

Isolation and characterization of the cis-trans-unsaturated fatty acid isomerase of Pseudomonas oleovorans GPo12.

Pseudomonas oleovorans contains an isomerase which catalyzes the cis-trans conversion of the abundant unsaturated membrane fatty acids 9-cis-hexadecenoic acid (palmitoleic acid) and 11-cis-octadecenoic acid (vaccenic acid). We purified the isomerase from the periplasmic fraction of Pseudomonas oleovorans. The molecular mass of the enzyme was estimated to be 80 kDa under denaturing conditions and 70 kDa under native conditions, suggesting a monomeric structure of the active enzyme. N-terminal sequencing showed that the isomerase derives from a precursor with a signal sequence which is cleaved from the primary translation product in accord with the periplasmic localization of the enzyme. The purified isomerase acted only on free unsaturated fatty acids and not on esterified fatty acids. In contrast to the in vivo cis-trans conversion of lipids, this in vitro isomerization of free fatty acids did not require the addition of organic solvents. Pure phospholipids, even in the presence of organic solvents, could not serve as substrate for the isomerase. However, when crude membranes from Pseudomonas or Escherichia coli cells were used as phospholipid sources, a cis-trans isomerization was detectable which occurred only in the presence of organic solvents. These results indicate that isolated membranes from Pseudomonas or E. coli cells must contain factors which, activated by the addition of organic solvents, enable and control the cis-trans conversion of unsaturated acyl chains of membrane phospholipids by the periplasmic isomerase.

Changes in Erythrocyte Membrane Lipids in Breast Cancer after Radiotherapy and Chemotherapy

CMF and radiation treatment alter the erythrocyte membrane architecture in breast cancer patients by inducing free radical formation in the system. The increased free radicals hydrolyze the unsaturated fatty acyl chains of membrane phospholipids and thus alter the membrane lipid profile. To assess the erythrocyte membrane damage caused by CMF chemotherapy and radiation treatment among breast cancer patients and to find a way which can protect them from the observed toxic manifestations form the basis of this study.

A role for phospholipid polyunsaturation in modulating membrane protein function.

Visual transduction is one of the best characterized G protein--coupled signalling systems. In addition, about 50% of the disk membrane phospholipid acyl chains are 22:6n-3, making this system ideal for determining the role of polyunsaturation in modulating membrane-signalling systems. The extent of formation of metarhodopsin II (MII), the G protein--activating photointermediate of rhodopsin, was studied in phospholipid vesicles composed of a variety of phosphatidylcholines, differing in their acyl chain composition at the sn-2 position. The amount of MII formed increased progressively with the level of acyl chain unsaturation at the sn-2 position. The effect of added cholesterol was to reduce the amount of MII formed. The acyl chain packing free volume of the rhodopsin containing lipid vesicles was characterized by a fractional volume parameter fv derived from measurements of the time-resolved fluorescence anisotropy decay of the hydrophobic membrane probe 1,6-diphenyl-1,3,5-hexatriene. The relationship among sn-2 acyl chain unsaturation, cholesterol content, and MII formation is explained on the basis of variation in fv with bilayer lipid composition and a novel model for the packing of phospholipids containing polyenoic acyl chains, such as 22:6n-3.

Dietary docosahexaenoic acid (22: 6n-3) prevents the development of hypertension in SHRSP.

1. We previously reported that hypertension in stroke-prone spontaneously hypertensive rats (SHRSP) caused renal membrane phospholipid degradation. Renal phospholipase A2 activity increased and membranous phospholipids decreased along with age in SHRSP. Membranous abnormalities induced by membrane fluidity and calcium permeability changes may contribute to the elevation of blood pressure in SHRSP. DHA, a major component of fish oil, constitutes a part of membrane phospholipid acylchains. 2. The purpose of this study was to clarify the effect of DHA on the relationship between the renal function and the development of hypertension in SHRSP. 3. Six week old male SHRSP were fed a semi-purified diet supplemented with DHA (0, 1 and 5%) for 14 weeks. 4. The systolic blood pressure of control SHRSP (DHA 0%) significantly increased from 120.2 mmHg to 202.9 mmHg. This increase in systolic blood pressure was significantly inhibited in a dose-dependent manner by 1 and 5% DHA diet to 167.8 to 149.8 mmHg, respectively. 5. Serum creatinine concentration and blood urea nitrogen (BUN) were significantly lower in DHA (5%)-treated SHRSP than in the control SHRSP. 6. These results indicate that DHA prevents the development of hypertension in SHRSP, which is associated with changes in renal function.

Phospholipase A2 as a mechanosensor

Osmotic swelling of large unilamellar vesicles (LUVs) causes membrane stretching and thus reduces the lateral packing of lipids. This is demonstrated to modulate strongly the catalytic activity of phospholipase A2 (PLA2) toward a fluorescent phospholipid, 1-palmitoyl-2-[(6-pyren-1-yl)]decanoyl-sn-glycero-3-phosphocholine (PPDPC) residing in LUVs composed of different unsaturated and saturated phosphatidylcholines. The magnitude of the osmotic pressure gradient delta omega required for maximal PLA2 activity as well as the extent of activation depend on the degree of saturation of the membrane phospholipid acyl chains. More specifically, delta omega needed for maximal hydrolytic activity increases in the sequence DOPC < SOPC < DMPC in accordance with the increment in the intensity of chain-chain van der Waals interactions. Previous studies on the hydrolysis of substrate monolayers by C. adamanteus and N. naja PLA2 revealed maximal hydrolytic rates for these two enzymes to be achieved at lipid packing densities corresponding to surface pressures of 12 and 18 mN m-1, respectively. In keeping with the above the magnitudes of delta omega producing maximal activity of Crotalus adamanteus and Naja naja toward PPDPC/DMPC LUVs were 40 and 20 mOsm/kg, respectively. Our findings suggest a novel possibility of regulating the activity of PLA2 and perhaps also other lipid packing density-dependent enzymes in vivo by osmotic forces applied on cellular membranes. Importantly, our results reveal serendipitously that the responsiveness of membranes to osmotic stress is modulated by the acyl chain composition of the lipids.

Peroxidation of erythrocytes: FTIR spectroscopy studies of extracted lipids, isolated membranes, and intact cells

AbstractFourier transform infrared (FTIR) spectroscopy was used to directly monitor peroxidative damage to membrane phospholipid acyl chains in erythrocyte membranes. Samples were suspended in a mixed 2H2O/H2O buffer system, thereby producing a “spectral window” in the C‐H stretching region of the infrared spectrum. A decrease in the number of acyl chain CC bonds upon erythrocyte peroxidation was quantitated directly from the spectra of isolated membranes. Second‐derivative spectroscopy permitted the conformationally sensitive membrane acyl chain methylene stretching modes to be separated from the protein (mostly hemoglobin) vibrations that dominate the spectra of intact cells. The sensitivity of these modes in erythrocytes was then determined in a series of thermotropic experiments. The effect of peroxidation upon the membrane acyl chain conformational order was monitored in isolated membranes and intact cells. No change in conformational order was detected upon peroxidation in intact cell and ghost spectra. In contrast, experiments with pure unsaturated phospholipids demonstrated that decreasing the CC bond population results in increased conformational order. The finding of identical results for peroxidation versus control samples in isolated membranes (“ghosts”) validates the results observed for intact cells. FTIR technology permits the direct monitoring of conformational order in the acyl chains of intact cells. © 1995 John Wiley &amp; Sons, Inc.

Membrane structural alterations in malignant and normal avian lymphocytes: a Raman investigation.

Resonance and nonresonance Raman spectra have been obtained from neoplastically transformed and normal avian lymphocytes. The acyl chains of membrane phospholipids of neoplastic cells are more highly unsaturated than those of normal cells. The observation of prominent carotenoid bands in both cell populations indicates the availability of a sensitive, intrinsic probe of membrane potential and local membrane environment.

Thermal regulation of membrane fluidity in Escherichia coli. Effects of overproduction of beta-ketoacyl-acyl carrier protein synthase I.

Multicopy plasmids bearing the structural gene (fabB) for beta-ketoacyl-acyl carrier protein (ACP) synthase I were constructed in vitro and transformed into various Escherichia coli strains. Introduction of these plasmids into fabB strains resulted in a fabB+ phenotype and a large (8- to 10-fold) overproduction of synthase I activity. Strains carrying these plasmids were also unusually resistant to cerulenin (an antibiotic that specifically inhibits beta-ketoacyl-ACP synthase activity) and overproduced cis-vaccenic acid. Strains (fabF-) lacking beta-ketoacyl-ACP synthase II are deficient in both cis-vaccenic acid synthesis and thermal regulation. Introduction of the fabB plasmids into these strains resulted in the restoration of cis-vaccenic acid synthesis. However, the plasmid-engendered cis-vaccenic acid synthesis of these strains was unaffected by temperature. These results demonstrate that synthase II, the product of the fabF gene, is the sole enzyme regulating the temperature-dependent composition of the membrane phospholipid acyl chains.

Interaction of DDT ( [formula omitted] with liposomal phospholipids

The influence of 1,1,1- trichloro-2,2- bis(p- chlorophenyl) ethane (DDT) and several other pesticides on the physical state of membrane phospholipids was investigated using model lipids. The thermal dependence of fluorescence intensity of the probe parinaric acid in dipalmitoylphosphatidylcholine liposomes and lipid vesicles of mixed composition were recorded. DDT was incorporated into the liposomal bilayer. The insecticide lowered the phase transition temperature and broadened the temperature range of the transition. The effects were concentration-dependent. The results may be interpreted as a sort of blurred and facilitated phase transition of bilayer lipids caused by intercalation of DDT between fatty acyl chains of membrane phospholipids.