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Sample records for membrane fatty acid-binding

  1. Natural ligand binding and transfer from liver fatty acid binding protein (LFABP) to membranes.

    PubMed

    De Gerónimo, Eduardo; Hagan, Robert M; Wilton, David C; Córsico, Betina

    2010-09-01

    Liver fatty acid-binding protein (LFABP) is distinctive among fatty acid-binding proteins because it binds more than one molecule of long-chain fatty acid and a variety of diverse ligands. Also, the transfer of fluorescent fatty acid analogues to model membranes under physiological ionic strength follows a different mechanism compared to most of the members of this family of intracellular lipid binding proteins. Tryptophan insertion mutants sensitive to ligand binding have allowed us to directly measure the binding affinity, ligand partitioning and transfer to model membranes of natural ligands. Binding of fatty acids shows a cooperative mechanism, while acyl-CoAs binding presents a hyperbolic behavior. Saturated fatty acids seem to have a stronger partition to protein vs. membranes, compared to unsaturated fatty acids. Natural ligand transfer rates are more than 200-fold higher compared to fluorescently-labeled analogues. Interestingly, oleoyl-CoA presents a markedly different transfer behavior compared to the rest of the ligands tested, probably indicating the possibility of specific targeting of ligands to different metabolic fates. PMID:20541621

  2. Isolation and partial characterization of a fatty acid binding protein in rat liver plasma membranes.

    PubMed Central

    Stremmel, W; Strohmeyer, G; Borchard, F; Kochwa, S; Berk, P D

    1985-01-01

    When [14C]oleate-bovine serum albumin complexes were incubated in vitro with rat liver plasma membranes (LPM), specific, saturable binding of oleate to the membranes was observed. Maximal heat-sensitive (i.e., specific) binding was 3.2 nmol/mg of membrane protein. Oleate-agarose affinity chromatography of Triton X-100-solubilized LPM was used to isolate a single 40-kDa protein with high affinity for oleate. On gel filtration, the protein comigrated with various fatty acids but not with [14C]bilirubin, [35S]sulfobromophthalein, [14C]taurocholate, [14C]phosphatidylcholine, or [14C]cholesteryloleate. A rabbit antibody to this membrane fatty acid-binding protein gave a single precipitin line with the antigen but no reactivity with concentrated cytosolic proteins, LPM bilirubin/sulfobromophthalein-binding protein, or rat albumin or other rat plasma proteins. The antibody selectively inhibited heat-sensitive binding of [14C]oleate to LPM. Immunofluorescence studies localized the antigen in liver-cell plasma membranes as well as in other major sites of fatty acid transport. These data are compatible with the hypothesis that this protein may act as a receptor in a hepatocellular uptake mechanism for fatty acids. Images PMID:3881757

  3. Surface lysine residues modulate the collisional transfer of fatty acid from adipocyte fatty acid binding protein to membranes.

    PubMed

    Herr, F M; Matarese, V; Bernlohr, D A; Storch, J

    1995-09-19

    The transfer of unesterified fatty acids (FA) from adipocyte fatty acid binding protein (A-FABP) to phospholipid membranes is proposed to occur via a collisional mechanism involving transient ionic and hydrophobic interactions [Wootan & Storch (1994) J. Biol. Chem. 269, 10517-10523]. In particular, it was suggested that membrane acidic phospholipids might specifically interact with basic residues on the surface of A-FABP. Here we addressed whether lysine residues on the surface of the protein are involved in this collisional transfer mechanism. Recombinant A-FABP was acetylated to neutralize all positively charged surface lysine residues. Protein fluorescence, CD spectra, and chemical denaturant data indicate that acetylation did not substantially alter the conformational integrity of the protein, and nearly identical affinities were obtained for binding of the fluorescently labeled FA [12-(9-anthroyloxy)oleate] to native and acetylated protein. Transfer of 2-(9-anthroyloxy)palmitate (2AP) from acetylated A-FABP to small unilamellar vesicles (SUV) was 35-fold slower than from native protein. In addition, whereas the 2AP transfer rate from native A-FABP was directly dependent on SUV concentration, 2AP transfer from acetylated protein was independent on the concentration of acceptor membranes. Factors which alter aqueous-phase solubility of FA, such as ionic strength and acyl chain length and saturation, affected the AOFA transfer rate from acetylated but not native A-FABP. Finally, an increase in the negative charge density of the acceptor SUV resulted in a marked increase in the rate of transfer from native A-FABP but did not increase the rate from acetylated A-FABP.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:7547918

  4. Role of portal region lysine residues in electrostatic interactions between heart fatty acid binding protein and phospholipid membranes.

    PubMed

    Herr, F M; Aronson, J; Storch, J

    1996-01-30

    The structure of heart fatty acid binding protein (HFABP) is a flattened beta-barrel comprising 10 antiparallel beta-sheets capped by two alpha-helical segments. The helical cap region is hypothesized to behave as a portal "lid" for the entry and release of ligand from the binding pocket. The transfer of fatty acid from HFABP is thought to occur via effective collisional interactions with membranes, and these interactions are enhanced when transfer is to membranes of net negative charge, thus implying that specific basic residues on the surface of HFABP may govern the transfer process [Wootan, M. G., & Storch, J. (1994) J. Biol. Chem. 269, 10517-10523]. To directly examine the role of charged lysine residues on the HFABP surface in specific interactions with membranes, chemical modification and selective mutagenesis of HFABP were used. All surface lysine residues were neutralized by acetylation of recombinant HFABP with acetic anhydride. In addition, seven mutant HFABPs were generated that resulted in charge alterations in five distinct sites of HFABP. Modification of the protein did not significantly alter the structural or ligand binding properties of HFABP, as assessed by circular dichroism, fluorescence quantum yield, and ligand binding analyses. By using a resonance energy transfer assay, transfer of 2-(9-anthroyloxy)palmitate (2AP) from acetylated HFABP to membranes was significantly slower than transfer from native HFABP. In addition, in distinct contrast to transfer from native protein, the 2AP transfer rate from acetylated HFABP was not increased to acceptor membranes of increased negative charge. Transfer of 2AP from HFABP mutants involving K22, located on alpha-helix I (alpha-I) of the helical cap region, was 3-fold slower than transfer from wild-type protein, whereas rates from a mutant involving the K59 residue, located on the beta 2-turn of the barrel near the helical cap, were 2-fold faster than those of wild type. A double mutant involving K22 and K

  5. The alpha-helical domain of liver fatty acid binding protein is responsible for the diffusion-mediated transfer of fatty acids to phospholipid membranes.

    PubMed

    Córsico, Betina; Liou, Heng Ling; Storch, Judith

    2004-03-30

    Intestinal fatty acid binding protein (IFABP) and liver FABP (LFABP), homologous proteins expressed at high levels in intestinal absorptive cells, employ markedly different mechanisms for the transfer of fatty acids (FAs) to acceptor membranes. Transfer from IFABP occurs during protein-membrane collisional interactions, while for LFABP, transfer occurs by diffusion through the aqueous phase. Earlier, we had shown that the helical domain of IFABP is critical in determining its collisional FA transfer mechanism. In the study presented here, we have engineered a pair of chimeric proteins, one with the "body" (ligand binding domain) of IFABP and the alpha-helical region of LFABP (alphaLbetaIFABP) and the other with the ligand binding pocket of LFABP and the helical domain of IFABP (alphaIbetaLFABP). The objective of this work was to determine whether the change in the alpha-helical domain of each FABP would alter the rate and mechanism of transfer of FA from the chimeric proteins in comparison with those of the wild-type proteins. The fatty acid transfer properties of the FABP chimeras were examined using a fluorescence resonance transfer assay. The results showed a significant modification of the absolute rate of FA transfer from the chimeric proteins compared to that of the wild type, indicating that the slower rate of FA transfer observed for wild-type LFABP relative to that of wild-type IFABP is, in part, determined by the helical domain of the proteins. In addition to these quantitative changes, it was of great interest to observe that the apparent mechanism of FA transfer also changed when the alpha-helical domain was exchanged, with transfer from alphaLbetaIFABP occurring by aqueous diffusion and transfer from alphaIbetaLFABP occurring via protein-membrane collisional interactions. These results demonstrate that the alpha-helical region of LFABP is responsible for its diffusional mechanism of fatty acid transfer to membranes. PMID:15035630

  6. Hepatocellular uptake of oleate is energy dependent, sodium linked, and inhibited by an antibody to a hepatocyte plasma membrane fatty acid binding protein.

    PubMed Central

    Stremmel, W; Strohmeyer, G; Berk, P D

    1986-01-01

    Several studies suggest that a portion of hepatocellular nonesterified fatty acid uptake may be carrier mediated. To further investigate this process, initial rates (Vo) of [14C]oleate uptake into rat hepatocytes, isolated by collagenase perfusion and incubated at 37 degrees C with oleate in the presence of bovine serum albumin, were studied as a function of the concentration of unbound [14C]oleate in the medium. Vo was saturable with increasing unbound oleate concentration (Km = 8.3 X 10(-8) M; Vmax = 197 pmol per min per 5 X 10(4) hepatocytes) and was not inhibited by up to 40 microM sulfobromophthalein, taurocholate, or cholic acid. Oleate uptake was sodium dependent. Vo was significantly diminished when Li+, K+, choline, or sucrose were substituted for Na+ in the incubation medium and was reduced 46% by 1 mM ouabain. Uptake was also markedly reduced after exposure of cells to metabolic inhibitors (e.g., 2,4-dinitrophenol, carbonyl cyanide m-chlorophenylhydrazone, antimycin, KCN). To evaluate the physiologic significance of the previously isolated rat liver plasma membrane fatty acid-binding protein, the effect of an antibody directed against this protein on hepatocellular [14C]oleate uptake was examined. Preincubation of hepatocytes with the IgG fraction of this antiserum inhibited Vo of [14C]oleate by up to 65% in dose-related fashion, without altering Vo for [35S]sulfobromophthalein, [14C]taurocholate, or [3H]cholate. These data indicate that at least a portion of hepatocellular oleate uptake is energy dependent, sodium linked, and mediated by a specific liver plasma membrane-fatty acid-binding protein. PMID:3459144

  7. The integrity of the alpha-helical domain of intestinal fatty acid binding protein is essential for the collision-mediated transfer of fatty acids to phospholipid membranes.

    PubMed

    Franchini, G R; Storch, J; Corsico, B

    2008-04-01

    Intestinal FABP (IFABP) and liver FABP (LFABP), homologous proteins expressed at high levels in intestinal absorptive cells, employ markedly different mechanisms of fatty acid transfer to acceptor model membranes. Transfer from IFABP occurs during protein-membrane collisional interactions, while for LFABP transfer occurs by diffusion through the aqueous phase. In addition, transfer from IFABP is markedly faster than from LFABP. The overall goal of this study was to further explore the structural differences between IFABP and LFABP which underlie their large functional differences in ligand transport. In particular, we addressed the role of the alphaI-helix domain in the unique transport properties of intestinal FABP. A chimeric protein was engineered with the 'body' (ligand binding domain) of IFABP and the alphaI-helix of LFABP (alpha(I)LbetaIFABP), and the fatty acid transfer properties of the chimeric FABP were examined using a fluorescence resonance energy transfer assay. The results showed a significant decrease in the absolute rate of FA transfer from alpha(I)LbetaIFABP compared to IFABP. The results indicate that the alphaI-helix is crucial for IFABP collisional FA transfer, and further indicate the participation of the alphaII-helix in the formation of a protein-membrane "collisional complex". Photo-crosslinking experiments with a photoactivable reagent demonstrated the direct interaction of IFABP with membranes and further support the importance of the alphaI helix of IFABP in its physical interaction with membranes. PMID:18284926

  8. Echinococcus granulosus fatty acid binding proteins subcellular localization.

    PubMed

    Alvite, Gabriela; Esteves, Adriana

    2016-05-01

    Two fatty acid binding proteins, EgFABP1 and EgFABP2, were isolated from the parasitic platyhelminth Echinococcus granulosus. These proteins bind fatty acids and have particular relevance in flatworms since de novo fatty acids synthesis is absent. Therefore platyhelminthes depend on the capture and intracellular distribution of host's lipids and fatty acid binding proteins could participate in lipid distribution. To elucidate EgFABP's roles, we investigated their intracellular distribution in the larval stage by a proteomic approach. Our results demonstrated the presence of EgFABP1 isoforms in cytosolic, nuclear, mitochondrial and microsomal fractions, suggesting that these molecules could be involved in several cellular processes. PMID:26873273

  9. Enterocyte Fatty Acid Binding Proteins (FABPs): Different Functions of Liver- and Intestinal- FABPs in the Intestine

    PubMed Central

    Gajda, Angela M.; Storch, Judith

    2014-01-01

    SUMMARY Fatty acid binding proteins (FABP) are highly abundant cytosolic proteins that are expressed in most mammalian tissues. In the intestinal enterocyte, both Liver- (LFABP; FABP1) and Intestinal-fatty acid binding proteins (IFABP; FABP2) are expressed. These proteins display high affinity binding for long chain fatty acids (FA) and other hydrophobic ligands, thus they are believed to be involved with uptake and trafficking of lipids in the intestine. In vitro studies have identified differences in ligand binding stoichiometry and specificity, and in mechanisms of FA transfer to membranes, and it has been hypothesized that LFABP and IFABP have difference functions in the enterocyte. Studies directly comparing LFABP- and IFABP-null mice have revealed markedly different phenotypes, indicating that these proteins indeed have different functions in intestinal lipid metabolism and whole body energy homeostasis. In this review, we discuss the evolving knowledge of the functions of LFABP and IFABP in the intestinal enterocyte. PMID:25458898

  10. Studies on fatty acid-binding proteins. The diurnal variation shown by rat liver fatty acid-binding protein.

    PubMed Central

    Wilkinson, T C; Wilton, D C

    1987-01-01

    The concentration of fatty acid-binding protein in rat liver was examined by SDS/polyacrylamide-gel electrophoresis, by Western blotting and by quantifying the fluorescence enhancement achieved on the binding of the fluorescent probe 11-(dansylamino)undecanoic acid. A 2-3-fold increase in the concentration of this protein produced by treatment of rats with the peroxisome proliferator tiadenol was readily detected; however, only a small variation in the concentration of the protein due to a diurnal rhythm was observed. This result contradicts the 7-10-fold variation previously reported for this protein [Hargis, Olson, Clarke & Dempsey (1986) J. Biol. Chem. 261, 1988-1991]. Images Fig. 1. Fig. 3. PMID:3593284

  11. Fatty acid induced remodeling within the human liver fatty acid-binding protein.

    PubMed

    Sharma, Ashwani; Sharma, Amit

    2011-09-01

    We crystallized human liver fatty acid-binding protein (LFABP) in apo, holo, and intermediate states of palmitic acid engagement. Structural snapshots of fatty acid recognition, entry, and docking within LFABP support a heads-in mechanism for ligand entry. Apo-LFABP undergoes structural remodeling, where the first palmitate ingress creates the atomic environment for placement of the second palmitate. These new mechanistic insights will facilitate development of pharmacological agents against LFABP. PMID:21757748

  12. Bile salt recognition by human liver fatty acid binding protein.

    PubMed

    Favretto, Filippo; Santambrogio, Carlo; D'Onofrio, Mariapina; Molinari, Henriette; Grandori, Rita; Assfalg, Michael

    2015-04-01

    Fatty acid binding proteins (FABPs) act as intracellular carriers of lipid molecules, and play a role in global metabolism regulation. Liver FABP (L-FABP) is prominent among FABPs for its wide ligand repertoire, which includes long-chain fatty acids as well as bile acids (BAs). In this work, we performed a detailed molecular- and atomic-level analysis of the interactions established by human L-FABP with nine BAs to understand the binding specificity for this important class of cholesterol-derived metabolites. Protein-ligand complex formation was monitored using heteronuclear NMR, steady-state fluorescence spectroscopy, and mass spectrometry. BAs were found to interact with L-FABP with dissociation constants in the narrow range of 0.6-7 μm; however, the diverse substitution patterns of the sterol nucleus and the presence of side-chain conjugation resulted in complexes endowed with various degrees of conformational heterogeneity. Trihydroxylated BAs formed monomeric complexes in which single ligand molecules occupied similar internal binding sites, based on chemical-shift perturbation data. Analysis of NMR line shapes upon progressive addition of taurocholate indicated that the binding mechanism departed from a simple binary association equilibrium, and instead involved intermediates along the binding path. The co-linear chemical shift behavior observed for L-FABP complexes with cholate derivatives added insight into conformational dynamics in the presence of ligands. The observed spectroscopic features of L-FABP/BA complexes, discussed in relation to ligand chemistry, suggest possible molecular determinants of recognition, with implications regarding intracellular BA transport. Our findings suggest that human L-FABP is a poorly selective, universal BA binder. PMID:25639618

  13. Effect of liver fatty acid binding protein on fatty acid movement between liposomes and rat liver microsomes.

    PubMed Central

    McCormack, M; Brecher, P

    1987-01-01

    Although movement of fatty acids between bilayers can occur spontaneously, it has been postulated that intracellular movement is facilitated by a class of proteins named fatty acid binding proteins (FABP). In this study we have incorporated long chain fatty acids into multilamellar liposomes made of phosphatidylcholine, incubated them with rat liver microsomes containing an active acyl-CoA synthetase, and measured formation of acyl-CoA in the absence or presence of FABP purified from rat liver. FABP increased about 2-fold the accumulation of acyl-CoA when liposomes were the fatty acid donor. Using fatty acid incorporated into liposomes made either of egg yolk lecithin or of dipalmitoylphosphatidylcholine, it was found that the temperature dependence of acyl-CoA accumulation in the presence of FABP correlated with both the physical state of phospholipid molecules in the liposomes and the binding of fatty acid to FABP, suggesting that fatty acid must first desorb from the liposomes before FABP can have an effect. An FABP-fatty acid complex incubated with microsomes, in the absence of liposomes, resulted in greater acyl-CoA formation than when liposomes were present, suggesting that desorption of fatty acid from the membrane is rate-limiting in the accumulation of acyl-CoA by this system. Finally, an equilibrium dialysis cell separating liposomes from microsomes on opposite sides of a Nuclepore filter was used to show that liver FABP was required for the movement and activation of fatty acid between the compartments. These studies show that liver FABP interacts with fatty acid that desorbs from phospholipid bilayers, and promotes movement to a membrane-bound enzyme, suggesting that FABP may act intracellularly by increasing net desorption of fatty acid from cell membranes. PMID:3446187

  14. Expression of liver fatty acid binding protein in hepatocellular carcinoma.

    PubMed

    Cho, Soo-Jin; Ferrell, Linda D; Gill, Ryan M

    2016-04-01

    Loss of expression of liver fatty acid binding protein (LFABP) by immunohistochemistry has been shown to be characteristic of a subset of hepatocellular adenomas (HCAs) in which HNF1A is inactivated. Transformation to hepatocellular carcinoma is thought to be a very rare phenomenon in the HNF1A-inactivated variant of HCA. However, we recently observed 2 cases at our institution, 1 definite hepatocellular carcinoma and 1 possible hepatocellular carcinoma, with loss of LFABP staining, raising the possibility that LFABP down-regulation may be associated with hepatocellular carcinogenesis. Our aim was to evaluate hepatocellular carcinomas arising in various backgrounds and with varying degrees of differentiation for loss of LFABP staining. Twenty total cases of hepatocellular carcinoma were examined. Thirteen cases arose in a background of cirrhosis due to hepatitis C (n = 8) or steatohepatitis (n = 5); 7 cases arose in a noncirrhotic background, with 2 cases arising within HNF1A-inactivated variant HCA and 2 cases arising within inflammatory variant HCA. Complete loss of expression of LFABP was seen in 6 of 20 cases, including 2 cases of hepatocellular carcinoma arising within HNF1A-inactivated variant HCA. Thus, loss of staining for LFABP appears to be common in hepatocellular carcinoma and may be seen in well-differentiated hepatocellular carcinoma. Therefore, LFABP loss should not be interpreted as evidence for hepatocellular adenoma over carcinoma, when other features support a diagnosis of hepatocellular carcinoma. The findings raise consideration for a role of HNF1A inactivation in hepatocellular carcinogenesis, particularly in less differentiated tumors. PMID:26997447

  15. Evidence that Chemical Chaperone 4-Phenylbutyric Acid Binds to Human Serum Albumin at Fatty Acid Binding Sites

    PubMed Central

    James, Joel; Shihabudeen, Mohamed Sham; Kulshrestha, Shweta; Goel, Varun; Thirumurugan, Kavitha

    2015-01-01

    Endoplasmic reticulum stress elicits unfolded protein response to counteract the accumulating unfolded protein load inside a cell. The chemical chaperone, 4-Phenylbutyric acid (4-PBA) is a FDA approved drug that alleviates endoplasmic reticulum stress by assisting protein folding. It is found efficacious to augment pathological conditions like type 2 diabetes, obesity and neurodegeneration. This study explores the binding nature of 4-PBA with human serum albumin (HSA) through spectroscopic and molecular dynamics approaches, and the results show that 4-PBA has high binding specificity to Sudlow Site II (Fatty acid binding site 3, subdomain IIIA). Ligand displacement studies, RMSD stabilization profiles and MM-PBSA binding free energy calculation confirm the same. The binding constant as calculated from fluorescence spectroscopic studies was found to be kPBA = 2.69 x 105 M-1. Like long chain fatty acids, 4-PBA induces conformational changes on HSA as shown by circular dichroism, and it elicits stable binding at Sudlow Site II (fatty acid binding site 3) by forming strong hydrogen bonding and a salt bridge between domain II and III of HSA. This minimizes the fluctuation of HSA backbone as shown by limited conformational space occupancy in the principal component analysis. The overall hydrophobicity of W214 pocket (located at subdomain IIA), increases upon occupancy of 4-PBA at any FA site. Descriptors of this pocket formed by residues from other subdomains largely play a role in compensating the dynamic movement of W214. PMID:26181488

  16. Liver fatty acid-binding protein binds monoacylglycerol in vitro and in mouse liver cytosol.

    PubMed

    Lagakos, William S; Guan, Xudong; Ho, Shiu-Ying; Sawicki, Luciana Rodriguez; Corsico, Betina; Kodukula, Sarala; Murota, Kaeko; Stark, Ruth E; Storch, Judith

    2013-07-01

    Liver fatty acid-binding protein (LFABP; FABP1) is expressed both in liver and intestinal mucosa. Mice null for LFABP were recently shown to have altered metabolism of not only fatty acids but also monoacylglycerol, the two major products of dietary triacylglycerol hydrolysis (Lagakos, W. S., Gajda, A. M., Agellon, L., Binas, B., Choi, V., Mandap, B., Russnak, T., Zhou, Y. X., and Storch, J. (2011) Am. J. Physiol. Gastrointest. Liver Physiol. 300, G803-G814). Nevertheless, the binding and transport of monoacylglycerol (MG) by LFABP are uncertain, with conflicting reports in the literature as to whether this single chain amphiphile is in fact bound by LFABP. In the present studies, gel filtration chromatography of liver cytosol from LFABP(-/-) mice shows the absence of the low molecular weight peak of radiolabeled monoolein present in the fractions that contain LFABP in cytosol from wild type mice, indicating that LFABP binds sn-2 MG in vivo. Furthermore, solution-state NMR spectroscopy demonstrates two molecules of sn-2 monoolein bound in the LFABP binding pocket in positions similar to those found for oleate binding. Equilibrium binding affinities are ∼2-fold lower for MG compared with fatty acid. Finally, kinetic studies examining the transfer of a fluorescent MG analog show that the rate of transfer of MG is 7-fold faster from LFABP to phospholipid membranes than from membranes to membranes and occurs by an aqueous diffusion mechanism. These results provide strong support for monoacylglycerol as a physiological ligand for LFABP and further suggest that LFABP functions in the efficient intracellular transport of MG. PMID:23658011

  17. Liver Fatty Acid-binding Protein Binds Monoacylglycerol in Vitro and in Mouse Liver Cytosol*

    PubMed Central

    Lagakos, William S.; Guan, Xudong; Ho, Shiu-Ying; Sawicki, Luciana Rodriguez; Corsico, Betina; Kodukula, Sarala; Murota, Kaeko; Stark, Ruth E.; Storch, Judith

    2013-01-01

    Liver fatty acid-binding protein (LFABP; FABP1) is expressed both in liver and intestinal mucosa. Mice null for LFABP were recently shown to have altered metabolism of not only fatty acids but also monoacylglycerol, the two major products of dietary triacylglycerol hydrolysis (Lagakos, W. S., Gajda, A. M., Agellon, L., Binas, B., Choi, V., Mandap, B., Russnak, T., Zhou, Y. X., and Storch, J. (2011) Am. J. Physiol. Gastrointest. Liver Physiol. 300, G803–G814). Nevertheless, the binding and transport of monoacylglycerol (MG) by LFABP are uncertain, with conflicting reports in the literature as to whether this single chain amphiphile is in fact bound by LFABP. In the present studies, gel filtration chromatography of liver cytosol from LFABP−/− mice shows the absence of the low molecular weight peak of radiolabeled monoolein present in the fractions that contain LFABP in cytosol from wild type mice, indicating that LFABP binds sn-2 MG in vivo. Furthermore, solution-state NMR spectroscopy demonstrates two molecules of sn-2 monoolein bound in the LFABP binding pocket in positions similar to those found for oleate binding. Equilibrium binding affinities are ∼2-fold lower for MG compared with fatty acid. Finally, kinetic studies examining the transfer of a fluorescent MG analog show that the rate of transfer of MG is 7-fold faster from LFABP to phospholipid membranes than from membranes to membranes and occurs by an aqueous diffusion mechanism. These results provide strong support for monoacylglycerol as a physiological ligand for LFABP and further suggest that LFABP functions in the efficient intracellular transport of MG. PMID:23658011

  18. Titration and exchange studies of liver fatty acid-binding protein with 13C-labeled long-chain fatty acids.

    PubMed

    Wang, Hsin; He, Yan; Kroenke, Christopher D; Kodukula, Sarala; Storch, Judith; Palmer, Arthur G; Stark, Ruth E

    2002-04-30

    Uniformly (13)C-labeled long-chain fatty acids were used to probe ligand binding to rat liver fatty acid-binding protein (LFABP), an atypical member of the fatty acid-binding protein (FABP) family that binds more than one molecule of long-chain fatty acid, accommodates a variety of diverse ligands, and exhibits diffusion-mediated lipid transport to membranes. Two sets of (1)H-(13)C resonances were found in a titration series of NMR spectra for oleate-LFABP complexes, indicating that two molecules of the fatty acid are situated in the protein cavity. However, no distinct resonances were observed for the excess fatty acid in solution, suggesting that at least one ligand undergoes rapid exchange with oleate in the bulk solution. An exchange rate of 54 +/- 6 s(-1) between the two sets of resonances was measured directly using (13)C z,z-exchange spectroscopy. In light of these NMR measurements, possible molecular mechanisms for the ligand-exchange process are evaluated and implications for the anomalous fatty acid transport mechanism of LFABP are discussed. PMID:11969406

  19. Molecular Dynamic Simulations Reveal the Structural Determinants of Fatty Acid Binding to Oxy-Myoglobin

    PubMed Central

    Chintapalli, Sree V.; Bhardwaj, Gaurav; Patel, Reema; Shah, Natasha; Patterson, Randen L.; van Rossum, Damian B.; Anishkin, Andriy; Adams, Sean H.

    2015-01-01

    The mechanism(s) by which fatty acids are sequestered and transported in muscle have not been fully elucidated. A potential key player in this process is the protein myoglobin (Mb). Indeed, there is a catalogue of empirical evidence supporting direct interaction of globins with fatty acid metabolites; however, the binding pocket and regulation of the interaction remains to be established. In this study, we employed a computational strategy to elucidate the structural determinants of fatty acids (palmitic & oleic acid) binding to Mb. Sequence analysis and docking simulations with a horse (Equus caballus) structural Mb reference reveals a fatty acid-binding site in the hydrophobic cleft near the heme region in Mb. Both palmitic acid and oleic acid attain a “U” shaped structure similar to their conformation in pockets of other fatty acid-binding proteins. Specifically, we found that the carboxyl head group of palmitic acid coordinates with the amino group of Lys45, whereas the carboxyl group of oleic acid coordinates with both the amino groups of Lys45 and Lys63. The alkyl tails of both fatty acids are supported by surrounding hydrophobic residues Leu29, Leu32, Phe33, Phe43, Phe46, Val67, Val68 and Ile107. In the saturated palmitic acid, the hydrophobic tail moves freely and occasionally penetrates deeper inside the hydrophobic cleft, making additional contacts with Val28, Leu69, Leu72 and Ile111. Our simulations reveal a dynamic and stable binding pocket in which the oxygen molecule and heme group in Mb are required for additional hydrophobic interactions. Taken together, these findings support a mechanism in which Mb acts as a muscle transporter for fatty acid when it is in the oxygenated state and releases fatty acid when Mb converts to deoxygenated state. PMID:26030763

  20. Molecular dynamic simulations reveal the structural determinants of Fatty Acid binding to oxy-myoglobin.

    PubMed

    Chintapalli, Sree V; Bhardwaj, Gaurav; Patel, Reema; Shah, Natasha; Patterson, Randen L; van Rossum, Damian B; Anishkin, Andriy; Adams, Sean H

    2015-01-01

    The mechanism(s) by which fatty acids are sequestered and transported in muscle have not been fully elucidated. A potential key player in this process is the protein myoglobin (Mb). Indeed, there is a catalogue of empirical evidence supporting direct interaction of globins with fatty acid metabolites; however, the binding pocket and regulation of the interaction remains to be established. In this study, we employed a computational strategy to elucidate the structural determinants of fatty acids (palmitic & oleic acid) binding to Mb. Sequence analysis and docking simulations with a horse (Equus caballus) structural Mb reference reveals a fatty acid-binding site in the hydrophobic cleft near the heme region in Mb. Both palmitic acid and oleic acid attain a "U" shaped structure similar to their conformation in pockets of other fatty acid-binding proteins. Specifically, we found that the carboxyl head group of palmitic acid coordinates with the amino group of Lys45, whereas the carboxyl group of oleic acid coordinates with both the amino groups of Lys45 and Lys63. The alkyl tails of both fatty acids are supported by surrounding hydrophobic residues Leu29, Leu32, Phe33, Phe43, Phe46, Val67, Val68 and Ile107. In the saturated palmitic acid, the hydrophobic tail moves freely and occasionally penetrates deeper inside the hydrophobic cleft, making additional contacts with Val28, Leu69, Leu72 and Ile111. Our simulations reveal a dynamic and stable binding pocket in which the oxygen molecule and heme group in Mb are required for additional hydrophobic interactions. Taken together, these findings support a mechanism in which Mb acts as a muscle transporter for fatty acid when it is in the oxygenated state and releases fatty acid when Mb converts to deoxygenated state. PMID:26030763

  1. Role of surface lysine residues of adipocyte fatty acid-binding protein in fatty acid transfer to phospholipid vesicles.

    PubMed

    Liou, H L; Storch, J

    2001-05-29

    The tertiary structure of murine adipocyte fatty acid-binding protein (AFABP) is a flattened 10-stranded beta-barrel capped by a helix-turn-helix segment. This helical domain is hypothesized to behave as a "lid" or portal for ligand entry into and exit from the binding cavity. Previously, we demonstrated that anthroyloxy-labeled fatty acid (AOFA) transfer from AFABP to phospholipid membranes occurs by a collisional process, in which ionic interactions between positively charged lysine residues on the protein surface and negatively charged phospholipid headgroups are involved. In the present study, the role of specific lysine residues located in the portal and other regions of AFABP was directly examined using site-directed mutagenesis. The results showed that isoleucine replacement for lysine in the portal region, including the alphaI- and alphaII-helices and the beta C-D turn, resulted in much slower 2-(9-anthroyloxy)palmitate (2AP) transfer rates to acidic membranes than those of native AFABP. An additive effect was found for mutant K22,59I, displaying the slowest rates of FA transfer. Rates of 2AP transfer from "nonportal" mutants on the beta-G and I strands were affected only moderately; however, a lysine --> isoleucine mutation in the nonportal beta-A strand decreased the 2AP transfer rate. These studies suggest that lysines in the helical cap domain are important for governing ionic interactions between AFABP and membranes. Furthermore, it appears that more than one distinct region, including the alphaI-helix, alphaII-helix, beta C-D turn, and the beta-A strand, is involved in these charge-charge interactions. PMID:11371211

  2. Urinary liver-type fatty acid-binding protein change in gestational diabetes mellitus.

    PubMed

    Fu, Wen-Jin; Wang, Du-Juan; Deng, Ren-Tang; Huang, Zhi-Hong; Chen, Mei-Lian; Jang, You-Ming; Wen, Shu; Yang, Hong-Ling; Huang, Xian-zhang

    2015-09-01

    We compared urinary liver-type fatty acid-binding protein (L-FABP) among non-pregnant and pregnant women with and without gestational diabetes mellitus (GDM). Higher urinary L-FABP was found in pregnant with and without GDM, and considerably higher urinary L-FABP was found in the GDM group compared with the non-GDM group. Hyperglycemia and anemia were related with high urinary L-FABP expression. PMID:26254248

  3. Fatty Acid-Binding Protein 5 Facilitates the Blood-Brain Barrier Transport of Docosahexaenoic Acid.

    PubMed

    Pan, Yijun; Scanlon, Martin J; Owada, Yuji; Yamamoto, Yui; Porter, Christopher J H; Nicolazzo, Joseph A

    2015-12-01

    The brain has a limited ability to synthesize the essential polyunsaturated fatty acid (PUFA) docosahexaenoic acid (DHA) from its omega-3 fatty acid precursors. Therefore, to maintain brain concentrations of this PUFA at physiological levels, plasma-derived DHA must be transported across the blood-brain barrier (BBB). While DHA is able to partition into the luminal membrane of brain endothelial cells, its low aqueous solubility likely limits its cytosolic transfer to the abluminal membrane, necessitating the requirement of an intracellular carrier protein to facilitate trafficking of this PUFA across the BBB. As the intracellular carrier protein fatty acid-binding protein 5 (FABP5) is expressed at the human BBB, the current study assessed the putative role of FABP5 in the brain endothelial cell uptake and BBB transport of DHA in vitro and in vivo, respectively. hFAPB5 was recombinantly expressed and purified from Escherichia coli C41(DE3) cells and the binding affinity of DHA to hFABP5 assessed using isothermal titration calorimetry. The impact of FABP5 siRNA on uptake of (14)C-DHA into immortalized human brain microvascular endothelial (hCMEC/D3) cells was assessed. An in situ transcardiac perfusion method was optimized in C57BL/6 mice and subsequently used to compare the BBB influx rate (Kin) of (14)C-DHA between FABP5-deficient (FABP5(-/-)) and wild-type (FABP5(+/+)) C57BL/6 mice. DHA bound to hFABP5 with an equilibrium dissociation constant of 155 ± 8 nM (mean ± SEM). FABP5 siRNA transfection decreased hCMEC/D3 mRNA and protein expression of FABP5 by 53.2 ± 5.5% and 44.8 ± 13.7%, respectively, which was associated with a 14.1 ± 2.7% reduction in (14)C-DHA cellular uptake. By using optimized conditions for the in situ transcardiac perfusion (a 1 min preperfusion (10 mL/min) followed by perfusion of (14)C-DHA (1 min)), the Kin of (14)C-DHA was 0.04 ± 0.01 mL/g/s. Relative to FABP5(+/+) mice, the Kin of (14)C-DHA decreased 36.7 ± 12.4% in FABP5(-/-) mice

  4. Enterocyte fatty acid-binding proteins (FABPs): different functions of liver and intestinal FABPs in the intestine.

    PubMed

    Gajda, Angela M; Storch, Judith

    2015-02-01

    Fatty acid-binding proteins (FABP) are highly abundant cytosolic proteins that are expressed in most mammalian tissues. In the intestinal enterocyte, both liver- (LFABP; FABP1) and intestinal FABPs (IFABP; FABP2) are expressed. These proteins display high-affinity binding for long-chain fatty acids (FA) and other hydrophobic ligands; thus, they are believed to be involved with uptake and trafficking of lipids in the intestine. In vitro studies have identified differences in ligand-binding stoichiometry and specificity, and in mechanisms of FA transfer to membranes, and it has been hypothesized that LFABP and IFABP have different functions in the enterocyte. Studies directly comparing LFABP- and IFABP-null mice have revealed markedly different phenotypes, indicating that these proteins indeed have different functions in intestinal lipid metabolism and whole body energy homeostasis. In this review, we discuss the evolving knowledge of the functions of LFABP and IFABP in the intestinal enterocyte. PMID:25458898

  5. Local Unfolding of Fatty Acid Binding Protein to Allow Ligand Entry for Binding.

    PubMed

    Xiao, Tianshu; Fan, Jing-Song; Zhou, Hu; Lin, Qingsong; Yang, Daiwen

    2016-06-01

    Fatty acid binding proteins are responsible for the transportation of fatty acids in biology. Despite intensive studies, the molecular mechanism of fatty acid entry to and exit from the protein cavity is still unclear. Here a cap-closed variant of human intestinal fatty acid binding protein was generated by mutagenesis, in which the helical cap is locked to the β-barrel by a disulfide linkage. Structure determination shows that this variant adopts a closed conformation, but still uptakes fatty acids. Stopped-flow experiments indicate that a rate-limiting step exists before the ligand association and this step corresponds to the conversion of the closed form to the open one. NMR relaxation dispersion and H-D exchange data demonstrate the presence of two excited states: one is native-like, but the other adopts a locally unfolded structure. Local unfolding of helix 2 generates an opening for ligands to enter the protein cavity, and thus controls the ligand association rate. PMID:27105780

  6. Studies on fatty acid-binding proteins. The detection and quantification of the protein from rat liver by using a fluorescent fatty acid analogue.

    PubMed Central

    Wilkinson, T C; Wilton, D C

    1986-01-01

    Fatty acid-binding protein from rat liver is shown to bind the fluorescent fatty acid probe dansyl undecanoic acid. Binding is accompanied by a shift in the fluorescence emission maximum from 550 nm to 500 nm and a 60-fold fluorescence enhancement at 500 nm. These spectral properties have allowed the use of this probe to detect and quantify microgram amounts of liver fatty acid-binding protein during purification procedures. In conjunction with h.p.l.c. the method allows the rapid estimation of liver fatty acid-binding protein in biological samples. The validity of the method is demonstrated by measuring the concentration of fatty acid-binding protein in livers from control and hypolipidaemic-drug-treated rats. The dramatic diurnal rhythm previously reported for this protein [Dempsey (1984) Curr. Top. Cell. Regul. 24, 63-86] was not observed with this method. Images Fig. 1. PMID:3800946

  7. Molecular dynamics simulation of ligand dissociation from liver fatty acid binding protein.

    PubMed

    Long, Dong; Mu, Yuguang; Yang, Daiwen

    2009-01-01

    The mechanisms of how ligands enter and leave the binding cavity of fatty acid binding proteins (FABPs) have been a puzzling question over decades. Liver fatty acid binding protein (LFABP) is a unique family member which accommodates two molecules of fatty acids in its cavity and exhibits the capability of interacting with a variety of ligands with different chemical structures and properties. Investigating the ligand dissociation processes of LFABP is thus a quite interesting topic, which however is rather difficult for both experimental approaches and ordinary simulation strategies. In the current study, random expulsion molecular dynamics simulation, which accelerates ligand motions for rapid dissociation, was used to explore the potential egress routes of ligands from LFABP. The results showed that the previously hypothesized "portal region" could be readily used for the dissociation of ligands at both the low affinity site and the high affinity site. Besides, one alternative portal was shown to be highly favorable for ligand egress from the high affinity site and be related to the unique structural feature of LFABP. This result lends strong support to the hypothesis from the previous NMR exchange studies, which in turn indicates an important role for this alternative portal. Another less favored potential portal located near the N-terminal end was also identified. Identification of the dissociation pathways will allow further mechanistic understanding of fatty acid uptake and release by computational and/or experimental techniques. PMID:19564911

  8. Characterization and amino acid sequence of a fatty acid-binding protein from human heart.

    PubMed

    Offner, G D; Brecher, P; Sawlivich, W B; Costello, C E; Troxler, R F

    1988-05-15

    The complete amino acid sequence of a fatty acid-binding protein from human heart was determined by automated Edman degradation of CNBr, BNPS-skatole [3'-bromo-3-methyl-2-(2-nitrobenzenesulphenyl)indolenine], hydroxylamine, Staphylococcus aureus V8 proteinase, tryptic and chymotryptic peptides, and by digestion of the protein with carboxypeptidase A. The sequence of the blocked N-terminal tryptic peptide from citraconylated protein was determined by collisionally induced decomposition mass spectrometry. The protein contains 132 amino acid residues, is enriched with respect to threonine and lysine, lacks cysteine, has an acetylated valine residue at the N-terminus, and has an Mr of 14768 and an isoelectric point of 5.25. This protein contains two short internal repeated sequences from residues 48-54 and from residues 114-119 located within regions of predicted beta-structure and decreasing hydrophobicity. These short repeats are contained within two longer repeated regions from residues 48-60 and residues 114-125, which display 62% sequence similarity. These regions could accommodate the charged and uncharged moieties of long-chain fatty acids and may represent fatty acid-binding domains consistent with the finding that human heart fatty acid-binding protein binds 2 mol of oleate or palmitate/mol of protein. Detailed evidence for the amino acid sequences of the peptides has been deposited as Supplementary Publication SUP 50143 (23 pages) at the British Library Lending Division, Boston Spa, Yorkshire LS23 7BQ, U.K., from whom copies may be obtained as indicated in Biochem. J. (1988) 249, 5. PMID:3421901

  9. The primary structure of fatty-acid-binding protein from nurse shark liver. Structural and evolutionary relationship to the mammalian fatty-acid-binding protein family.

    PubMed

    Medzihradszky, K F; Gibson, B W; Kaur, S; Yu, Z H; Medzihradszky, D; Burlingame, A L; Bass, N M

    1992-02-01

    The primary structure of a fatty-acid-binding protein (FABP) isolated from the liver of the nurse shark (Ginglymostoma cirratum) was determined by high-performance tandem mass spectrometry (employing multichannel array detection) and Edman degradation. Shark liver FABP consists of 132 amino acids with an acetylated N-terminal valine. The chemical molecular mass of the intact protein determined by electrospray ionization mass spectrometry (Mr = 15124 +/- 2.5) was in good agreement with that calculated from the amino acid sequence (Mr = 15121.3). The amino acid sequence of shark liver FABP displays significantly greater similarity to the FABP expressed in mammalian heart, peripheral nerve myelin and adipose tissue (61-53% sequence similarity) than to the FABP expressed in mammalian liver (22% similarity). Phylogenetic trees derived from the comparison of the shark liver FABP amino acid sequence with the members of the mammalian fatty-acid/retinoid-binding protein gene family indicate the initial divergence of an ancestral gene into two major subfamilies: one comprising the genes for mammalian liver FABP and gastrotropin, the other comprising the genes for mammalian cellular retinol-binding proteins I and II, cellular retinoic-acid-binding protein myelin P2 protein, adipocyte FABP, heart FABP and shark liver FABP, the latter having diverged from the ancestral gene that ultimately gave rise to the present day mammalian heart-FABP, adipocyte FABP and myelin P2 protein sequences. The sequence for intestinal FABP from the rat could be assigned to either subfamily, depending on the approach used for phylogenetic tree construction, but clearly diverged at a relatively early evolutionary time point. Indeed, sequences proximately ancestral or closely related to mammalian intestinal FABP, liver FABP, gastrotropin and the retinoid-binding group of proteins appear to have arisen prior to the divergence of shark liver FABP and should therefore also be present in elasmobranchs

  10. Exogenous fatty acid binding protein 4 promotes human prostate cancer cell progression.

    PubMed

    Uehara, Hisanori; Takahashi, Tetsuyuki; Oha, Mina; Ogawa, Hirohisa; Izumi, Keisuke

    2014-12-01

    Epidemiologic studies have found that obesity is associated with malignant grade and mortality in prostate cancer. Several adipokines have been implicated as putative mediating factors between obesity and prostate cancer. Fatty acid binding protein 4 (FABP4), a member of the cytoplasmic fatty acid binding protein multigene family, was recently identified as a novel adipokine. Although FABP4 is released from adipocytes and mean circulating concentrations of FABP4 are linked with obesity, effects of exogenous FABP4 on prostate cancer progression are unclear. In this study, we examined the effects of exogenous FABP4 on human prostate cancer cell progression. FABP4 treatment promoted serum-induced prostate cancer cell invasion in vitro. Furthermore, oleic acid promoted prostate cancer cell invasion only if FABP4 was present in the medium. These promoting effects were reduced by FABP4 inhibitor, which inhibits FABP4 binding to fatty acids. Immunostaining for FABP4 showed that exogenous FABP4 was taken up into DU145 cells in three-dimensional culture. In mice, treatment with FABP4 inhibitor reduced the subcutaneous growth and lung metastasis of prostate cancer cells. Immunohistochemical analysis showed that the number of apoptotic cells, positive for cleaved caspase-3 and cleaved PARP, was increased in subcutaneous tumors of FABP4 inhibitor-treated mice, as compared with control mice. These results suggest that exogenous FABP4 might promote human prostate cancer cell progression by binding with fatty acids. Additionally, exogenous FABP4 activated the PI3K/Akt pathway, independently of binding to fatty acids. Thus, FABP4 might be a key molecule to understand the mechanisms underlying the obesity-prostate cancer progression link. PMID:24740818

  11. Model of β-Sheet of Muscle Fatty Acid Binding Protein of Locusta migratoria Displays Characteristic Topology

    PubMed Central

    Kizilbash, Nadeem A; Hai, Abdul; Alruwaili, Jamal

    2013-01-01

    The β-sheet of muscle fatty acid binding protein of Locusta migratoria (Lm-FABP) was modeled by employing 2-D NMR data and the Rigid Body Assembly method. The model shows the β-sheet to comprise ten β-strands arranged anti-parallel to each other. There is a β-bulge between Ser 13 and Gln 14 which is a difference from the published structure of β-sheet of bovine heart Fatty Acid Binding Protein. Also, a hydrophobic patch consisting of Ile 45, Phe 51, Phe 64 and Phe 66 is present on the surface which is characteristic of most Fatty Acid Binding Proteins. A “gap” is present between βD and βE that provides evidence for the presence of a portal or opening between the polypeptide chains which allows ligand fatty acids to enter the protein cavity and bind to the protein. PMID:24497726

  12. Structural analysis of ibuprofen binding to human adipocyte fatty-acid binding protein (FABP4)

    PubMed Central

    González, Javier M.; Fisher, S. Zoë

    2015-01-01

    Inhibition of human adipocyte fatty-acid binding protein (FABP4) has been proposed as a treatment for type 2 diabetes, fatty liver disease and atherosclerosis. However, FABP4 displays a naturally low selectivity towards hydrophobic ligands, leading to the possibility of side effects arising from cross-inhibition of other FABP isoforms. In a search for structural determinants of ligand-binding selectivity, the binding of FABP4 towards a group of small molecules structurally related to the nonsteroidal anti-inflammatory drug ibuprofen was analyzed through X-ray crystallography. Several specific hydrophobic interactions are shown to enhance the binding affinities of these compounds, whereas an aromatic edge-to-face interaction is proposed to determine the conformation of bound ligands, highlighting the importance of aromatic interactions in hydrophobic environments. PMID:25664790

  13. Biological characterization of liver fatty acid binding gene from miniature pig liver cDNA library.

    PubMed

    Gao, Y H; Wang, K F; Zhang, S; Fan, Y N; Guan, W J; Ma, Y H

    2015-01-01

    Liver fatty acid binding proteins (L-FABP) are a family of small, highly conserved, cytoplasmic proteins that bind to long-chain fatty acids and other hydrophobic ligands. In this study, a full-length enriched cDNA library was successfully constructed from Wuzhishan miniature pig, and then the L-FABP gene was cloned from this cDNA library and an expression vector (pEGFP-N3-L-FABP) was constructed in vitro. This vector was transfected into hepatocytes to test its function. The results of western blotting analysis demonstrated that the L-FABP gene from our full-length enriched cDNA library regulated downstream genes, including the peroxisome proliferator-activated receptor family in hepatocytes. This study provides a theoretical basis and experimental evidence for the application of L-FABP for the treatment of liver injury. PMID:26345909

  14. Intramuscular fat content and genetic variants at fatty acid-binding protein loci in Austrian pigs.

    PubMed

    Nechtelberger, D; Pires, V; Söolknet, J; Stur; Brem, G; Mueller, M; Mueller, S

    2001-11-01

    Intramuscular fat is an important meat quality trait in pig production. Previously, genetic variants of the heart fatty acid-binding protein (H-FABP) gene and the adipocyte fatty acid-binding protein (A-FABP) gene were suggested to be associated with intramuscular fat content. The objective of this investigation was to study these associations in the three most important Austrian breeding populations (Piétrain, Large White, and Landrace). Restriction fragment length polymorphism analysis of the H-FABP gene revealed a new MspI polymorphic site and genetic variation in all three breeds. Microsatellite analysis of the A-FABP locus showed up to nine different microsatellite alleles segregating. In Austrian breeds, no significant influence of the A-FABP and H-FABP gene polymorphisms on intramuscular fat could be detected. We also evaluated possible associations between the genetic variations at the H-FABP and A-FABP loci and other growth and carcass traits (average daily gain, feed conversion ratio, lean meat content, pH values, meat color, and drip loss). With regard to the extent of the effects, these genetic markers cannot be recommended for selection on growth and carcass traits in Austrian breeding populations. PMID:11768107

  15. Liver fatty acid binding protein: species variation and the accommodation of different ligands.

    PubMed

    Thompson, J; Reese-Wagoner, A; Banaszak, L

    1999-11-23

    The crystal structure of rat liver fatty acid binding protein (LFABP) and an alignment of amino acid sequences of all known species have been used to demonstrate two groups or sub-classes. Based on estimates at neutral pH and the electrostatic field calculated using the crystal coordinates, some evidence of changes that occur in going from holo- to apo-forms has been obtained. LFABP belongs to a large family frequently referred to as the intracellular lipid binding proteins or iLBPs. LFABP, unlike other family members, has two fatty acid binding sites. The two cavity sites have been reviewed and arguments for interactions between the sites are presented. Based on the crystal structure of rat LFABP, differences between the A and B groups have been postulated. Last of all, hypothetical models have been built of complexes of LFABP and heme, and LFABP and oleoyl CoA. In both cases, the stoichiometry is one to one and the models show why this is likely. PMID:10570240

  16. Biochemical Roles for Conserved Residues in the Bacterial Fatty Acid-binding Protein Family.

    PubMed

    Broussard, Tyler C; Miller, Darcie J; Jackson, Pamela; Nourse, Amanda; White, Stephen W; Rock, Charles O

    2016-03-18

    Fatty acid kinase (Fak) is a ubiquitous Gram-positive bacterial enzyme consisting of an ATP-binding protein (FakA) that phosphorylates the fatty acid bound to FakB. In Staphylococcus aureus, Fak is a global regulator of virulence factor transcription and is essential for the activation of exogenous fatty acids for incorporation into phospholipids. The 1.2-Å x-ray structure of S. aureus FakB2, activity assays, solution studies, site-directed mutagenesis, and in vivo complementation were used to define the functions of the five conserved residues that define the FakB protein family (Pfam02645). The fatty acid tail is buried within the protein, and the exposed carboxyl group is bound by a Ser-93-fatty acid carboxyl-Thr-61-His-266 hydrogen bond network. The guanidinium of the invariant Arg-170 is positioned to potentially interact with a bound acylphosphate. The reduced thermal denaturation temperatures of the T61A, S93A, and H266A FakB2 mutants illustrate the importance of the hydrogen bond network in protein stability. The FakB2 T61A, S93A, and H266A mutants are 1000-fold less active in the Fak assay, and the R170A mutant is completely inactive. All FakB2 mutants form FakA(FakB2)2 complexes except FakB2(R202A), which is deficient in FakA binding. Allelic replacement shows that strains expressing FakB2 mutants are defective in fatty acid incorporation into phospholipids and virulence gene transcription. These conserved residues are likely to perform the same critical functions in all bacterial fatty acid-binding proteins. PMID:26774272

  17. Role of a liver fatty acid-binding protein gene in lipid metabolism in chicken hepatocytes.

    PubMed

    Gao, G L; Na, W; Wang, Y X; Zhang, H F; Li, H; Wang, Q G

    2015-01-01

    This study investigated the role of the chicken liver fatty acid-binding protein (L-FABP) gene in lipid metabolism in hepatocytes, and the regulatory relationships between L-FABP and genes related to lipid metabolism. The short hairpin RNA (shRNA) interference vector with L-FABP and an eukaryotic expression vector were used. Chicken hepatocytes were subjected to shRNA-mediated knockdown or L-FABP cDNA overexpression. Expression levels of lipid metabolism-related genes and biochemical parameters were detected 24, 36, 48, 60, and 72 h after transfection with the interference or overexpression plasmids for L-FABP, PPARα and L-BABP expression levels, and the total amount of cholesterol, were significantly affected by L-FABP expression. L-FABP may affect lipid metabolism by regulating PPARα and L-BABP in chicken hepatocytes. PMID:25966259

  18. Fatty acid-binding site environments of serum vitamin D-binding protein and albumin are different

    PubMed Central

    Swamy, Narasimha; Ray, Rahul

    2008-01-01

    Vitamin D-binding protein (DBP) and albumin (ALB) are abundant serum proteins and both possess high-affinity binding for saturated and unsaturated fatty acids. However, certain differences exist. We surmised that in cases where serum albumin level is low, DBP presumably can act as a transporter of fatty acids. To explore this possibility we synthesized several alkylating derivatives of 14C-palmitic acid to probe the fatty acid binding pockets of DBP and ALB. We observed that N-ethyl-5-phenylisooxazolium-3′-sulfonate-ester (WRK ester) of 14C-palmitic acid specifically labeled DBP; but p-nitrophenyl- and N-hydroxysuccinimidyl-esters failed to do so. However, p-nitrophenyl ester of 14C-palmitic acid specifically labeled bovine ALB, indicating that the micro-environment of the fatty acid-binding domains of DBP and ALB may be different; and DBP may not replace ALB as a transporter of fatty acids. PMID:18374965

  19. Solution-state molecular structure of apo and oleate-liganded liver fatty acid-binding protein.

    PubMed

    He, Yan; Yang, Xiaomin; Wang, Hsin; Estephan, Rima; Francis, Fouad; Kodukula, Sarala; Storch, Judith; Stark, Ruth E

    2007-11-01

    Rat liver fatty acid-binding protein (LFABP) is distinctive among intracellular lipid-binding proteins (iLBPs): more than one molecule of long-chain fatty acid and a variety of diverse ligands can be bound within its large cavity, and in vitro lipid transfer to model membranes follows a mechanism that is diffusion-controlled rather than mediated by protein-membrane collisions. Because the apoprotein has proven resistant to crystallization, nuclear magnetic resonance spectroscopy offers a unique route to functionally informative comparisons of molecular structure and dynamics for LFABP in free (apo) and liganded (holo) forms. We report herein the solution-state structures determined for apo-LFABP at pH 6.0 and for holoprotein liganded to two oleates at pH 7.0, as well as the structure of the complex including locations of the ligands. 1H, 13C, and 15N resonance assignments revealed very similar types and locations of secondary structural elements for apo- and holo-LFABP as judged from chemical shift indices. The solution-state tertiary structures of the proteins were derived with the CNS/ARIA computational protocol, using distance and angular restraints based on 1H-1H nuclear Overhauser effects (NOEs), hydrogen-bonding networks, 3J(HNHA) coupling constants, intermolecular NOEs, and residual dipolar (NH) couplings. The holo-LFABP solution-state conformation is in substantial agreement with a previously reported X-ray structure [Thompson, J., Winter, N., Terwey, D., Bratt, J., and Banaszak, L. (1997) The crystal structure of the liver fatty acid-binding protein. A complex with two bound oleates, J. Biol. Chem. 272, 7140-7150], including the typical beta-barrel capped by a helix-turn-helix portal. In the solution state, the internally bound oleate has the expected U-shaped conformation and is tethered electrostatically, but the extended portal ligand can adopt a range of conformations based on the computationally refined structures, in contrast to the single

  20. Fatty acid-binding protein 7 regulates function of caveolae in astrocytes through expression of caveolin-1.

    PubMed

    Kagawa, Yoshiteru; Yasumoto, Yuki; Sharifi, Kazem; Ebrahimi, Majid; Islam, Ariful; Miyazaki, Hirofumi; Yamamoto, Yui; Sawada, Tomoo; Kishi, Hiroko; Kobayashi, Sei; Maekawa, Motoko; Yoshikawa, Takeo; Takaki, Eiichi; Nakai, Akira; Kogo, Hiroshi; Fujimoto, Toyoshi; Owada, Yuji

    2015-05-01

    Fatty acid-binding proteins (FABPs) bind and solubilize long-chain fatty acids, controlling intracellular lipid dynamics. FABP7 is expressed by astrocytes in the developing brain, and suggested to be involved in the control of astrocyte lipid homeostasis. In this study, we sought to examine the role of FABP7 in astrocytes, focusing on plasma membrane lipid raft function, which is important for receptor-mediated signal transduction in response to extracellular stimuli. In FABP7-knockout (KO) astrocytes, the ligand-dependent accumulation of Toll-like receptor 4 (TLR4) and glial cell-line-derived neurotrophic factor receptor alpha 1 into lipid raft was decreased, and the activation of mitogen-activated protein kinases and nuclear factor-κB was impaired after lipopolysaccharide (LPS) stimulation when compared with wild-type astrocytes. In addition, the expression of caveolin-1, not cavin-1, 2, 3, caveolin-2, and flotillin-1, was found to be decreased at the protein and transcriptional levels. FABP7 re-expression in FABP7-KO astrocytes rescued the decreased level of caveolin-1. Furthermore, caveolin-1-transfection into FABP7-KO astrocytes significantly increased TLR4 recruitment into lipid raft and tumor necrosis factor-α production after LPS stimulation. Taken together, these data suggest that FABP7 controls lipid raft function through the regulation of caveolin-1 expression and is involved in the response of astrocytes to the external stimuli. GLIA 2015;63:780-794. PMID:25601031

  1. Expression Pattern of Fatty Acid Binding Proteins in Celiac Disease Enteropathy

    PubMed Central

    Bottasso Arias, Natalia M.; García, Marina; Bondar, Constanza; Guzman, Luciana; Redondo, Agustina; Chopita, Nestor; Córsico, Betina; Chirdo, Fernando G.

    2015-01-01

    Celiac disease (CD) is an immune-mediated enteropathy that develops in genetically susceptible individuals following exposure to dietary gluten. Severe changes at the intestinal mucosa observed in untreated CD patients are linked to changes in the level and in the pattern of expression of different genes. Fully differentiated epithelial cells express two isoforms of fatty acid binding proteins (FABPs): intestinal and liver, IFABP and LFABP, respectively. These proteins bind and transport long chain fatty acids and also have other important biological roles in signaling pathways, particularly those related to PPARγ and inflammatory processes. Herein, we analyze the serum levels of IFABP and characterize the expression of both FABPs at protein and mRNA level in small intestinal mucosa in severe enteropathy and normal tissue. As a result, we observed higher levels of circulating IFABP in untreated CD patients compared with controls and patients on gluten-free diet. In duodenal mucosa a differential FABPs expression pattern was observed with a reduction in mRNA levels compared to controls explained by the epithelium loss in severe enteropathy. In conclusion, we report changes in FABPs' expression pattern in severe enteropathy. Consequently, there might be alterations in lipid metabolism and the inflammatory process in the small intestinal mucosa. PMID:26346822

  2. Inhibition of Fatty Acid Binding Proteins Elevates Brain Anandamide Levels and Produces Analgesia

    PubMed Central

    Kaczocha, Martin; Rebecchi, Mario J.; Ralph, Brian P.; Teng, Yu-Han Gary; Berger, William T.; Galbavy, William; Elmes, Matthew W.; Glaser, Sherrye T.; Wang, Liqun; Rizzo, Robert C.; Deutsch, Dale G.; Ojima, Iwao

    2014-01-01

    The endocannabinoid anandamide (AEA) is an antinociceptive lipid that is inactivated through cellular uptake and subsequent catabolism by fatty acid amide hydrolase (FAAH). Fatty acid binding proteins (FABPs) are intracellular carriers that deliver AEA and related N-acylethanolamines (NAEs) to FAAH for hydrolysis. The mammalian brain expresses three FABP subtypes: FABP3, FABP5, and FABP7. Recent work from our group has revealed that pharmacological inhibition of FABPs reduces inflammatory pain in mice. The goal of the current work was to explore the effects of FABP inhibition upon nociception in diverse models of pain. We developed inhibitors with differential affinities for FABPs to elucidate the subtype(s) that contributes to the antinociceptive effects of FABP inhibitors. Inhibition of FABPs reduced nociception associated with inflammatory, visceral, and neuropathic pain. The antinociceptive effects of FABP inhibitors mirrored their affinities for FABP5, while binding to FABP3 and FABP7 was not a predictor of in vivo efficacy. The antinociceptive effects of FABP inhibitors were mediated by cannabinoid receptor 1 (CB1) and peroxisome proliferator-activated receptor alpha (PPARα) and FABP inhibition elevated brain levels of AEA, providing the first direct evidence that FABPs regulate brain endocannabinoid tone. These results highlight FABPs as novel targets for the development of analgesic and anti-inflammatory therapeutics. PMID:24705380

  3. Expression Pattern of Fatty Acid Binding Proteins in Celiac Disease Enteropathy.

    PubMed

    Bottasso Arias, Natalia M; García, Marina; Bondar, Constanza; Guzman, Luciana; Redondo, Agustina; Chopita, Nestor; Córsico, Betina; Chirdo, Fernando G

    2015-01-01

    Celiac disease (CD) is an immune-mediated enteropathy that develops in genetically susceptible individuals following exposure to dietary gluten. Severe changes at the intestinal mucosa observed in untreated CD patients are linked to changes in the level and in the pattern of expression of different genes. Fully differentiated epithelial cells express two isoforms of fatty acid binding proteins (FABPs): intestinal and liver, IFABP and LFABP, respectively. These proteins bind and transport long chain fatty acids and also have other important biological roles in signaling pathways, particularly those related to PPARγ and inflammatory processes. Herein, we analyze the serum levels of IFABP and characterize the expression of both FABPs at protein and mRNA level in small intestinal mucosa in severe enteropathy and normal tissue. As a result, we observed higher levels of circulating IFABP in untreated CD patients compared with controls and patients on gluten-free diet. In duodenal mucosa a differential FABPs expression pattern was observed with a reduction in mRNA levels compared to controls explained by the epithelium loss in severe enteropathy. In conclusion, we report changes in FABPs' expression pattern in severe enteropathy. Consequently, there might be alterations in lipid metabolism and the inflammatory process in the small intestinal mucosa. PMID:26346822

  4. Buffer interference with protein dynamics: a case study on human liver fatty acid binding protein.

    PubMed

    Long, Dong; Yang, Daiwen

    2009-02-18

    Selection of suitable buffer types is often a crucial step for generating appropriate protein samples for NMR and x-ray crystallographic studies. Although the possible interaction between MES buffer (2-(N-morpholino)ethanesulfonic acid) and proteins has been discussed previously, the interaction is usually thought to have no significant effects on the structures of proteins. In this study, we demonstrate the direct, albeit weak, interaction between MES and human liver fatty acid binding protein (hLFABP). Rather than affecting the structure of hLFABP, we found that the dynamics of hLFABP, which were previously proposed to be relevant to its functions, were significantly affected by the binding of hLFABP with MES. Buffer interference with protein dynamics was also demonstrated with Bis-Tris buffer, which is quite different from MES and fatty acids in terms of their molecular structures and properties. This result, to our knowledge, is the first published report on buffer interference with protein dynamics on a microsecond to millisecond timescale and could represent a generic problem in the studies of functionally relevant protein dynamics. Although being a fortuity, our finding of buffer-induced changes in protein dynamics offers a clue to how hLFABP accommodates its ligands. PMID:19217864

  5. Temporal profile of intestinal tissue expression of intestinal fatty acid-binding protein in a rat model of necrotizing enterocolitis

    PubMed Central

    Simões, Ana Leda Bertoncini; Figueira, Rebeca Lopes; Gonçalves, Frances Lilian Lanhellas; Mitidiero, Luís Felipe Tsuyoshi; Silva, Orlando Castro e; Peiró, José Luis; Sbragia, Lourenço

    2016-01-01

    OBJECTIVES: Necrotizing enterocolitis is a severe multifactorial intestinal disorder that primarily affects preterm newborns, causing 20-40% mortality and morbidity. Intestinal fatty acid-binding protein has been reported to be a biomarker for the detection of intestinal injuries. Our aim was to assess intestinal tissue injury and the molecular expression of intestinal fatty acid-binding protein over time in a necrotizing enterocolitis model. METHODS: A total of 144 Newborn rats were divided into two groups: 1) Control, which received breastfeeding (n=72) and 2) Necrotizing Enterocolitis, which received formula feeding and underwent hypoxia and hypothermia (n=72). A total of six time points of ischemia (2 times a day for 3 days; 12 pups for each time point) were examined. Samples were collected for analysis of body weight, morphological and histological characteristics, intestinal weight, intestinal weight/body weight ratio, injury grade, and intestinal fatty acid-binding protein levels. RESULTS: Body and intestinal weights were lower in the Necrotizing Enterocolitis group than in the Control group (p<0.005 and p<0.0005, respectively). The intestinal weight/body weight ratio was higher in the Necrotizing Enterocolitis group than in the Control group (p<0.005) only at the sixth ischemia time point. The Necrotizing Enterocolitis group displayed higher expression of intestinal fatty acid-binding protein (p<0.0005) and showed greater tissue damage than the Control group. CONCLUSION: Intestinal fatty acid-binding protein was an efficient marker of ischemic injury to the intestine and a good correlation was demonstrated between the time of ischemic injury and the grade of intestinal injury. PMID:27464299

  6. Zinc-induced oligomerization of zinc α2 glycoprotein reveals multiple fatty acid-binding sites.

    PubMed

    Zahid, Henna; Miah, Layeque; Lau, Andy M; Brochard, Lea; Hati, Debolina; Bui, Tam T T; Drake, Alex F; Gor, Jayesh; Perkins, Stephen J; McDermott, Lindsay C

    2016-01-01

    Zinc α2 glycoprotein (ZAG) is an adipokine with a class I MHC protein fold and is associated with obesity and diabetes. Although its intrinsic ligand remains unknown, ZAG binds the dansylated C11 fatty acid 11-(dansylamino)undecanoic acid (DAUDA) in the groove between the α1 and α2 domains. The surface of ZAG has approximately 15 weak zinc-binding sites deemed responsible for precipitation from human plasma. In the present study the functional significance of these metal sites was investigated. Analytical ultracentrifugation (AUC) and CD showed that zinc, but not other divalent metals, causes ZAG to oligomerize in solution. Thus ZAG dimers and trimers were observed in the presence of 1 and 2 mM zinc. Molecular modelling of X-ray scattering curves and sedimentation coefficients indicated a progressive stacking of ZAG monomers, suggesting that the ZAG groove may be occluded in these. Using fluorescence-detected sedimentation velocity, these ZAG-zinc oligomers were again observed in the presence of the fluorescent boron dipyrromethene fatty acid C16-BODIPY (4,4-difluoro-5,7-dimethyl-4-bora-3a,4a-diaza-s-indacene-3-hexadecanoic acid). Fluorescence spectroscopy confirmed that ZAG binds C16-BODIPY. ZAG binding to C16-BODIPY, but not to DAUDA, was reduced by increased zinc concentrations. We conclude that the lipid-binding groove in ZAG contains at least two distinct fatty acid-binding sites for DAUDA and C16-BODIPY, similar to the multiple lipid binding seen in the structurally related immune protein CD1c. In addition, because high concentrations of zinc occur in the pancreas, the perturbation of these multiple lipid-binding sites by zinc may be significant in Type 2 diabetes where dysregulation of ZAG and zinc homoeostasis occurs. PMID:26487699

  7. Liver Fatty acid binding protein (L-Fabp) modulates murine stellate cell activation and diet induced nonalcoholic fatty liver disease

    PubMed Central

    Chen, Anping; Tang, Youcai; Davis, Victoria; Hsu, Fong-Fu; Kennedy, Susan M.; Song, Haowei; Turk, John; Brunt, Elizabeth M.; Newberry, Elizabeth P.; Davidson, Nicholas O.

    2013-01-01

    Activation of hepatic stellate cells (HSCs) is crucial to the development of fibrosis in nonalcoholic fatty liver disease. Quiescent HSCs contain lipid droplets (LDs), whose depletion upon activation induces a fibrogenic gene program. Here we show that liver fatty acid-binding protein (L-Fabp), an abundant cytosolic protein that modulates fatty acid (FA) metabolism in enterocytes and hepatocytes also modulates HSC FA utilization and in turn regulates the fibrogenic program. L-Fabp expression decreased 10-fold following HSC activation, concomitant with depletion of LDs. Primary HSCs isolated from L-FABP−/− mice contain fewer LDs than wild type (WT) HSCs, and exhibit upregulated expression of genes involved in HSC activation. Adenoviral L-Fabp transduction inhibited activation of passaged WT HSCs and increased both the expression of prolipogenic genes and also augmented intracellular lipid accumulation, including triglyceride and FA, predominantly palmitate. Freshly isolated HSCs from L-FABP−/− mice correspondingly exhibited decreased palmitate in the free FA pool. To investigate whether L-FABP deletion promotes HSC activation in vivo, we fed L-FABP−/− and WT mice a high fat diet supplemented with trans-fatty acids and fructose (TFF). TFF-fed L-FABP−/− mice exhibited reduced hepatic steatosis along with decreased LD abundance and size compared to WT mice. In addition, TFF-fed L-FABP−/− mice exhibited decreased hepatic fibrosis, with reduced expression of fibrogenic genes, compared to WT mice. Conclusion L-FABP deletion attenuates both diet-induced hepatic steatosis and fibrogenesis, despite the observation that L-Fabp paradoxically promotes FA and LD accumulation and inhibits HSC activation in vitro. These findings highlight the importance of cell-specific modulation of hepatic lipid metabolism in promoting fibrogenesis in nonalcoholic fatty liver disease. PMID:23401290

  8. Medium-chain fatty acid binding to albumin and transfer to phospholipid bilayers

    SciTech Connect

    Hamilton, J.A. )

    1989-04-01

    Temperature-dependent (5-42{degree}C) {sup 13}C NMR spectra of albumin complexes with 90% isotopically substituted (1-{sup 13}C)octanoic or (1-{sup 13}C)decanoic acids showed a single peak at >30{degree}C but three peaks at lower temperatures. The chemical-shift differences result from different ionic and/or hydrogen-bonding interactions between amino acid side chains and the fatty acid carboxyl carbon. Rapid exchange of fatty acid among binding sites obscures these sites at temperatures >30{degree}C. Rate constants for exchange at 33{degree}C were 350 sec{sup {minus}1} for octanoate and 20 sec {sup {minus}1} for decanoate. Temperature-dependent data for octanoate showed an activation energy of 2 kcal/mol for exchange. Spectra of albumin complexes with the 12-carbon saturated fatty acid, lauric acid, had several narrow laurate carboxyl peaks at 35{degree}C, indicating longer lifetimes in the different binding sites. Fatty acid exchange between albumin and model membranes (phosphatidylcholine bilayers) occurred on a time scale comparable to that for exchange among albumin binding sites, following the order octanoate > decanoate > laurate. The equilibrium distribution of fatty acid between lipid bilayers and protein was measured directly from NMR spectra. Decreasing pH increased the relative affinity of fatty acid for the lipid bilayer. The results predict that the relative affinity of octanoic acid for albumin and membranes will be similar to that of long-chain fatty acids, but the rate of equilibration will be {approx} 10{sup 4} faster for octanoic acid.

  9. Exploring and Expanding the Fatty-Acid-Binding Protein Superfamily in Fasciola Species.

    PubMed

    Morphew, Russell M; Wilkinson, Toby J; Mackintosh, Neil; Jahndel, Veronika; Paterson, Steve; McVeigh, Paul; Abbas Abidi, Syed M; Saifullah, Khalid; Raman, Muthusamy; Ravikumar, Gopalakrishnan; LaCourse, James; Maule, Aaron; Brophy, Peter M

    2016-09-01

    The liver flukes Fasciola hepatica and F. gigantica infect livestock worldwide and threaten food security with climate change and problematic control measures spreading disease. Fascioliasis is also a foodborne disease with up to 17 million humans infected. In the absence of vaccines, treatment depends on triclabendazole (TCBZ), and overuse has led to widespread resistance, compromising future TCBZ control. Reductionist biology from many laboratories has predicted new therapeutic targets. To this end, the fatty-acid-binding protein (FABP) superfamily has proposed multifunctional roles, including functions intersecting vaccine and drug therapy, such as immune modulation and anthelmintic sequestration. Research is hindered by a lack of understanding of the full FABP superfamily complement. Although discovery studies predicted FABPs as promising vaccine candidates, it is unclear if uncharacterized FABPs are more relevant for vaccine formulations. We have coupled genome, transcriptome, and EST data mining with proteomics and phylogenetics to reveal a liver fluke FABP superfamily of seven clades: previously identified clades I-III and newly identified clades IV-VII. All new clade FABPs were analyzed using bioinformatics and cloned from both liver flukes. The extended FABP data set will provide new study tools to research the role of FABPs in parasite biology and as therapy targets. PMID:27495901

  10. Identification of novel PTEN-binding partners: PTEN interaction with fatty acid binding protein FABP4.

    PubMed

    Gorbenko, O; Panayotou, G; Zhyvoloup, A; Volkova, D; Gout, I; Filonenko, V

    2010-04-01

    PTEN is a tumor suppressor with dual protein and lipid-phosphatase activity, which is frequently deleted or mutated in many human advanced cancers. Recent studies have also demonstrated that PTEN is a promising target in type II diabetes and obesity treatment. Using C-terminal PTEN sequence in pEG202-NLS as bait, yeast two-hybrid screening on Mouse Embryo, Colon Cancer, and HeLa cDNA libraries was carried out. Isolated positive clones were validated by mating assay and identified through automated DNA sequencing and BLAST database searches. Sequence analysis revealed a number of PTEN-binding proteins linking this phosphatase to a number of different signaling cascades, suggesting that PTEN may perform other functions besides tumor-suppressing activity in different cell types. In particular, the interplay between PTEN function and adipocyte-specific fatty-acid-binding protein FABP4 is of notable interest. The demonstrable tautology of PTEN to FABP4 suggested a role for this phosphatase in the regulation of lipid metabolism and adipocyte differentiation. This interaction was further studied using coimmunoprecipitation and gel-filtration assays. Finally, based on Biacore assay, we have calculated the K(D) of PTEN-FABP4 complex, which is around 2.8 microM. PMID:19911253

  11. Plasma Fatty Acid Binding Protein 4 and Risk of Sudden Cardiac Death in Older Adults

    PubMed Central

    Djoussé, Luc; Maziarz, Marlena; Biggs, Mary L.; Ix, Joachim H.; Zieman, Susan J.; Kizer, Jorge R.; Lemaitre, Rozenn N.; Mozaffarian, Dariush; Tracy, Russell P.; Mukamal, Kenneth J.; Siscovick, David S.; Sotoodehnia, Nona

    2013-01-01

    Although fatty acid binding protein 4 (FABP4) may increase risk of diabetes and exert negative cardiac inotropy, it is unknown whether plasma concentrations of FABP4 are associated with incidence of sudden cardiac death (SCD). We prospectively analyzed data on 4,560 participants of the Cardiovascular Health Study. FABP4 was measured at baseline using ELISA, and SCD events were adjudicated through review of medical records. We used Cox proportional hazards to estimate effect measures. During a median followup of 11.8 years, 146 SCD cases occurred. In a multivariable model adjusting for demographic, lifestyle, and metabolic factors, relative risk of SCD associated with each higher standard deviation (SD) of plasma FABP4 was 1.15 (95% CI: 0.95–1.38), P = 0.15. In a secondary analysis stratified by prevalent diabetes status, FABP4 was associated with higher risk of SCD in nondiabetic participants, (RR per SD higher FABP4: 1.33 (95% CI: 1.07–1.65), P = 0.009) but not in diabetic participants (RR per SD higher FABP4: 0.88 (95% CI: 0.62–1.27), P = 0.50), P for diabetes-FABP4 interaction 0.049. In summary, a single measure of plasma FABP4 obtained later in life was not associated with the risk of SCD in older adults overall. Confirmation of our post-hoc results in nondiabetic people in other studies is warranted. PMID:24455402

  12. Intestinal-fatty acid binding protein and lipid transport in human intestinal epithelial cells

    SciTech Connect

    Montoudis, Alain; Delvin, Edgard; Menard, Daniel

    2006-01-06

    Intestinal-fatty acid binding protein (I-FABP) is a 14-15 kDa cytoplasmic molecule highly expressed in the enterocyte. Although different functions have been proposed for various FABP family members, the specific function of I-FABP in human intestine remains unclear. Here, we studied the role of I-FABP in molecularly modified normal human intestinal epithelial cells (HIEC-6). cDNA transfection resulted in 90-fold I-FABP overexpression compared to cells treated with empty pQCXIP vector. The high-resolution immunogold technique revealed labeling mainly in the cytosol and confirmed the marked phenotype abundance of I-FABP in cDNA transfected cells. I-FABP overexpression was not associated with alterations in cell proliferation and viability. Studies using these transfected cells cultured with [{sup 14}C]oleic acid did not reveal higher efficiency in de novo synthesis or secretion of triglycerides, phospholipids, and cholesteryl esters compared to cells treated with empty pQCXIP vector only. Similarly, the incubation with [{sup 35}S]methionine did not disclose a superiority in the biogenesis of apolipoproteins (apo) A-I, A-IV, B-48, and B-100. Finally, cells transfected with I-FABP did not exhibit an increased production of chylomicrons, VLDL, LDL, and HDL. Our observations establish that I-FABP overexpression in normal HIEC-6 is not related to cell proliferation, lipid esterification, apo synthesis, and lipoprotein assembly, and, therefore, exclude its role in intestinal fat transport.

  13. Association of androgen with gender difference in serum adipocyte fatty acid binding protein levels

    PubMed Central

    Hu, Xiang; Ma, Xiaojing; Pan, Xiaoping; Luo, Yuqi; Xu, Yiting; Xiong, Qin; Bao, Yuqian; Jia, Weiping

    2016-01-01

    Clinical investigations have indicated women have higher levels of adipocyte fatty acid binding protein (A-FABP) than men. The present study aimed to identify factors related to gender difference in serum A-FABP levels. A total of 507 participants (194 men, 132 premenopausal women, and 181 postmenopausal women) were enrolled in the present study. Serum A-FABP levels increased in the order from men to premenopausal women to postmenopausal women in both body mass index categories (<25.0 and ≥25.0 kg/m2; all P < 0.05). Multiple stepwise regression analyses showed that after adjustment for factors related to serum A-FABP levels, the trunk fat mass was an independent and positive factor of serum A-FABP levels. For men, total testosterone was associated independently and inversely with serum A-FABP levels. For pre- and postmenopausal women, bioavailable testosterone and total testosterone were independent and positive factors associated with serum A-FABP levels, respectively. The present study demonstrated that the androgen was correlated with the serum A-FABP levels negatively in men, but positively in women. With these effects on the fat content, especially trunk fat, androgen might contribute to the gender difference in serum A-FABP levels. PMID:27270834

  14. Association of androgen with gender difference in serum adipocyte fatty acid binding protein levels.

    PubMed

    Hu, Xiang; Ma, Xiaojing; Pan, Xiaoping; Luo, Yuqi; Xu, Yiting; Xiong, Qin; Bao, Yuqian; Jia, Weiping

    2016-01-01

    Clinical investigations have indicated women have higher levels of adipocyte fatty acid binding protein (A-FABP) than men. The present study aimed to identify factors related to gender difference in serum A-FABP levels. A total of 507 participants (194 men, 132 premenopausal women, and 181 postmenopausal women) were enrolled in the present study. Serum A-FABP levels increased in the order from men to premenopausal women to postmenopausal women in both body mass index categories (<25.0 and ≥25.0 kg/m(2); all P < 0.05). Multiple stepwise regression analyses showed that after adjustment for factors related to serum A-FABP levels, the trunk fat mass was an independent and positive factor of serum A-FABP levels. For men, total testosterone was associated independently and inversely with serum A-FABP levels. For pre- and postmenopausal women, bioavailable testosterone and total testosterone were independent and positive factors associated with serum A-FABP levels, respectively. The present study demonstrated that the androgen was correlated with the serum A-FABP levels negatively in men, but positively in women. With these effects on the fat content, especially trunk fat, androgen might contribute to the gender difference in serum A-FABP levels. PMID:27270834

  15. Serologic Intestinal-Fatty Acid Binding Protein in Necrotizing Enterocolitis Diagnosis: A Meta-Analysis

    PubMed Central

    Cheng, Shupeng; Yu, Jialin; Zhou, Min; Tu, Yan; Lu, Qi

    2015-01-01

    Background. Previous studies showed that intestinal-fatty acid binding protein (I-FABP) may be a valid and promising serologic biomarker for early diagnosis of necrotizing enterocolitis (NEC). Objective. To investigate the early diagnostic value of serologic I-FABP in NEC for the premature neonates. Methods. All major databases were searched from January 1, 1990, to May 1, 2015. We used Meta-Disc 1.4 and Revman5.0 software to calculate the diagnostic accuracy. Results. Seven studies with 444 subjects were identified. The pooled sensitivity of I-FABP was 0.67 for NEC I, 0.74 for NEC II, and 0.83 for NEC III, and the pooled specificity was 0.84, respectively, which showed a moderate diagnostic accuracy. The area under curve (AUC) for each stage was 0.75 (Q⁎ = 0.69), 0.82 (Q⁎ = 0.76), and 0.91 (Q⁎ = 0.84). The diagnostic threshold analysis showed no significant difference in threshold effect. The metaregression showed that the cut-off value has the largest effect on heterogeneity. The funnel plots indicated the existence of publication bias. Conclusion. I-FABP is a valid serologic biomarker for early diagnosis in NEC for the premature neonates with a moderate accuracy. PMID:26798632

  16. Examination of the Addictive and Behavioral Properties of Fatty Acid-Binding Protein Inhibitor SBFI26.

    PubMed

    Thanos, Panayotis K; Clavin, Brendan H; Hamilton, John; O'Rourke, Joseph R; Maher, Thomas; Koumas, Christopher; Miao, Erick; Lankop, Jessenia; Elhage, Aya; Haj-Dahmane, Samir; Deutsch, Dale; Kaczocha, Martin

    2016-01-01

    The therapeutic properties of cannabinoids have been well demonstrated but are overshadowed by such adverse effects as cognitive and motor dysfunction, as well as their potential for addiction. Recent research on the natural lipid ligands of cannabinoid receptors, also known as endocannabinoids, has shed light on the mechanisms of intracellular transport of the endocannabinoid anandamide by fatty acid-binding proteins (FABPs) and subsequent catabolism by fatty acid amide hydrolase. These findings facilitated the recent development of SBFI26, a pharmacological inhibitor of epidermal- and brain-specific FABP5 and FABP7, which effectively increases anandamide signaling. The goal of this study was to examine this compound for any possible rewarding and addictive properties as well as effects on locomotor activity, working/recognition memory, and propensity for sociability and preference for social novelty (SN) given its recently reported anti-inflammatory and analgesic properties. Male C57BL mice were split into four treatment groups and conditioned with 5.0, 20.0, 40.0 mg/kg SBFI26, or vehicle during a conditioned place preference (CPP) paradigm. Following CPP, mice underwent a battery of behavioral tests [open field, novel object recognition (NOR), social interaction (SI), and SN] paired with acute SBFI26 administration. Results showed that SBFI26 did not produce CPP or conditioned place aversion regardless of dose and did not induce any differences in locomotor and exploratory activity during CPP- or SBFI26-paired open field activity. We also observed no differences between treatment groups in NOR, SI, and SN. In conclusion, as SBFI26 was shown previously by our group to have significant analgesic and anti-inflammatory properties, here we show that it does not pose a risk of dependence or motor and cognitive impairment under the conditions tested. PMID:27092087

  17. Examination of the Addictive and Behavioral Properties of Fatty Acid-Binding Protein Inhibitor SBFI26

    PubMed Central

    Thanos, Panayotis K.; Clavin, Brendan H.; Hamilton, John; O’Rourke, Joseph R.; Maher, Thomas; Koumas, Christopher; Miao, Erick; Lankop, Jessenia; Elhage, Aya; Haj-Dahmane, Samir; Deutsch, Dale; Kaczocha, Martin

    2016-01-01

    The therapeutic properties of cannabinoids have been well demonstrated but are overshadowed by such adverse effects as cognitive and motor dysfunction, as well as their potential for addiction. Recent research on the natural lipid ligands of cannabinoid receptors, also known as endocannabinoids, has shed light on the mechanisms of intracellular transport of the endocannabinoid anandamide by fatty acid-binding proteins (FABPs) and subsequent catabolism by fatty acid amide hydrolase. These findings facilitated the recent development of SBFI26, a pharmacological inhibitor of epidermal- and brain-specific FABP5 and FABP7, which effectively increases anandamide signaling. The goal of this study was to examine this compound for any possible rewarding and addictive properties as well as effects on locomotor activity, working/recognition memory, and propensity for sociability and preference for social novelty (SN) given its recently reported anti-inflammatory and analgesic properties. Male C57BL mice were split into four treatment groups and conditioned with 5.0, 20.0, 40.0 mg/kg SBFI26, or vehicle during a conditioned place preference (CPP) paradigm. Following CPP, mice underwent a battery of behavioral tests [open field, novel object recognition (NOR), social interaction (SI), and SN] paired with acute SBFI26 administration. Results showed that SBFI26 did not produce CPP or conditioned place aversion regardless of dose and did not induce any differences in locomotor and exploratory activity during CPP- or SBFI26-paired open field activity. We also observed no differences between treatment groups in NOR, SI, and SN. In conclusion, as SBFI26 was shown previously by our group to have significant analgesic and anti-inflammatory properties, here we show that it does not pose a risk of dependence or motor and cognitive impairment under the conditions tested. PMID:27092087

  18. Liver fatty acid-binding protein (L-FABP) promotes cellular angiogenesis and migration in hepatocellular carcinoma.

    PubMed

    Ku, Chung-Yu; Liu, Yu-Huei; Lin, Hsuan-Yuan; Lu, Shao-Chun; Lin, Jung-Yaw

    2016-04-01

    Liver fatty acid-binding protein (L-FABP) is abundant in hepatocytes and known to be involved in lipid metabolism. Overexpression of L-FABP has been reported in various cancers; however, its role in hepatocellular carcinoma (HCC) remains unclear. In this study, we investigated L-FABP and its association with vascular endothelial growth factors (VEGFs) in 90 HCC patients. We found that L-FABP was highly expressed in their HCC tissues, and that this expression was positively correlated with that of VEGF-A. Additionally, L-FABP significantly promoted tumor growth and metastasis in a xenograft mouse model. We also assessed the mechanisms of L-FABP activity in tumorigenesis; L-FABP was found to associate with VEGFR2 on membrane rafts and subsequently activate the Akt/mTOR/P70S6K/4EBP1 and Src/FAK/cdc42 pathways, which resulted in up-regulation of VEGF-A accompanied by an increase in both angiogenic potential and migration activity. Our results thus suggest that L-FABP could be a potential target for HCC chemotherapy. PMID:26919097

  19. Liver fatty acid-binding protein (L-FABP) promotes cellular angiogenesis and migration in hepatocellular carcinoma

    PubMed Central

    Ku, Chung-Yu; Liu, Yu-Huei; Lin, Hsuan-Yuan; Lu, Shao-Chun; Lin, Jung-Yaw

    2016-01-01

    Liver fatty acid-binding protein (L-FABP) is abundant in hepatocytes and known to be involved in lipid metabolism. Overexpression of L-FABP has been reported in various cancers; however, its role in hepatocellular carcinoma (HCC) remains unclear. In this study, we investigated L-FABP and its association with vascular endothelial growth factors (VEGFs) in 90 HCC patients. We found that L-FABP was highly expressed in their HCC tissues, and that this expression was positively correlated with that of VEGF-A. Additionally, L-FABP significantly promoted tumor growth and metastasis in a xenograft mouse model. We also assessed the mechanisms of L-FABP activity in tumorigenesis; L-FABP was found to associate with VEGFR2 on membrane rafts and subsequently activate the Akt/mTOR/P70S6K/4EBP1 and Src/FAK/cdc42 pathways, which resulted in up-regulation of VEGF-A accompanied by an increase in both angiogenic potential and migration activity. Our results thus suggest that L-FABP could be a potential target for HCC chemotherapy. PMID:26919097

  20. Adaptive Evolution of Eel Fluorescent Proteins from Fatty Acid Binding Proteins Produces Bright Fluorescence in the Marine Environment

    PubMed Central

    Gruber, David F.; Gaffney, Jean P.; Mehr, Shaadi; DeSalle, Rob; Sparks, John S.; Platisa, Jelena; Pieribone, Vincent A.

    2015-01-01

    We report the identification and characterization of two new members of a family of bilirubin-inducible fluorescent proteins (FPs) from marine chlopsid eels and demonstrate a key region of the sequence that serves as an evolutionary switch from non-fluorescent to fluorescent fatty acid-binding proteins (FABPs). Using transcriptomic analysis of two species of brightly fluorescent Kaupichthys eels (Kaupichthys hyoproroides and Kaupichthys n. sp.), two new FPs were identified, cloned and characterized (Chlopsid FP I and Chlopsid FP II). We then performed phylogenetic analysis on 210 FABPs, spanning 16 vertebrate orders, and including 163 vertebrate taxa. We show that the fluorescent FPs diverged as a protein family and are the sister group to brain FABPs. Our results indicate that the evolution of this family involved at least three gene duplication events. We show that fluorescent FABPs possess a unique, conserved tripeptide Gly-Pro-Pro sequence motif, which is not found in non-fluorescent fatty acid binding proteins. This motif arose from a duplication event of the FABP brain isoforms and was under strong purifying selection, leading to the classification of this new FP family. Residues adjacent to the motif are under strong positive selection, suggesting a further refinement of the eel protein’s fluorescent properties. We present a phylogenetic reconstruction of this emerging FP family and describe additional fluorescent FABP members from groups of distantly related eels. The elucidation of this class of fish FPs with diverse properties provides new templates for the development of protein-based fluorescent tools. The evolutionary adaptation from fatty acid-binding proteins to fluorescent fatty acid-binding proteins raises intrigue as to the functional role of bright green fluorescence in this cryptic genus of reclusive eels that inhabit a blue, nearly monochromatic, marine environment. PMID:26561348

  1. Liver fatty acid binding protein is the mitosis-associated polypeptide target of a carcinogen in rat hepatocytes

    SciTech Connect

    Bassuk, J.A.; Tsichlis, P.N.; Sorof, S.

    1987-11-01

    Hepatocytes in normal rat liver were found previously to contain a cytoplasmic 14,000-dalton polypeptide (p14) that is associated with mitosis and is the principal early covalent target of activated metabolites of the carcinogen N-2-fluorenylacetamide (2-acetylaminofluorene). The level of immunohistochemically detected p14 was low when growth activity of hepatocytes was low, was markedly elevated during mitosis in normal and regenerating livers, but was very high throughout interphase during proliferation of hyperplastic and malignant hepatocytes induced in rat liver by a carcinogen (N-2-fluorenylacetamide or 3'-methyl-4-dimethylaminoazobenzene). The authors report here that p14 is the liver fatty acid binding protein. The nucleotide sequence of p14 cDNA clones, isolated by screening a rat liver cDNA library in bacteriophage lambdagt11 using p14 antiserum, was completely identical to part of the sequence reported for liver fatty acid binding protein. Furthermore, the two proteins shared the following properties: size of mRNA, amino acid composition, molecular size according to NaDodSO/sub 4/ gel electrophoresis, and electrophoretic mobilities in a Triton X-100/acetic acid/urea gel. The two polypeptides bound oleic acid similarly. Finally, identical elevations of cytoplasmic immunostain were detected specifically in mitotic hepatocytes with either antiserum. The collected findings are suggestive that liver fatty acid binding protein may carry ligands that promote hepatocyte division and may transport certain activated chemical carcinogens.

  2. Interaction of perfluoroalkyl acids with human liver fatty acid-binding protein.

    PubMed

    Sheng, Nan; Li, Juan; Liu, Hui; Zhang, Aiqian; Dai, Jiayin

    2016-01-01

    Perfluoroalkyl acids (PFAAs) are highly persistent and bioaccumulative, resulting in their broad distribution in humans and the environment. The liver is an important target for PFAAs, but the mechanisms behind PFAAs interaction with hepatocyte proteins remain poorly understood. We characterized the binding of PFAAs to human liver fatty acid-binding protein (hL-FABP) and identified critical structural features in their interaction. The binding interaction of PFAAs with hL-FABP was determined by fluorescence displacement and isothermal titration calorimetry (ITC) assay. Molecular simulation was conducted to define interactions at the binding sites. ITC measurement revealed that PFOA/PFNA displayed a moderate affinity for hL-FABP at a 1:1 molar ratio, a weak binding affinity for PFHxS and no binding for PFHxA. Moreover, the interaction was mainly mediated by electrostatic attraction and hydrogen bonding. Substitution of Asn111 with Asp caused loss of binding affinity to PFAA, indicating its crucial role for the initial PFAA binding to the outer binding site. Substitution of Arg122 with Gly caused only one molecule of PFAA to bind to hL-FABP. Molecular simulation showed that substitution of Arg122 increased the volume of the outer binding pocket, making it impossible to form intensive hydrophobic stacking and hydrogen bonds with PFOA, and highlighting its crucial role in the binding process. The binding affinity of PFAAs increased significantly with their carbon number. Arg122 and Asn111 played a pivotal role in these interactions. Our findings may help understand the distribution pattern, bioaccumulation, elimination, and toxicity of PFAAs in humans. PMID:25370009

  3. Polymorphisms in Fatty Acid Binding Protein 5 Show Association with Type 2 Diabetes

    PubMed Central

    Bu, Liming; Salto, Lorena M.; De Leon, Kevin J; De Leon, Marino

    2011-01-01

    Genes for the fatty acid binding protein (FABP) family encode small 14–15 kDa cytosolic proteins and can be regulated during type 2 diabetes mellitus (T2DM) and obesity. This study compared association of single nucleotide polymorphisms (SNPs) in FABP1-5 with T2DM in different ethnic groups. Associations with T2DM of SNPs in these proteins were assessed in African American (AA), non-Hispanic White (NHW), and Hispanic American (HA) individuals. A total of 650 DNA samples were genotyped; control samples were obtained from Coriell’s North American Human Variation Panel Repository (NAVP) of apparently healthy individuals and T2DM cases were taken from the American Diabetes Association GENNID Study. The rs454550 SNP of FABP5 showed a significant association with T2DM in NHW (OR: 9.03, 95% CI: 1.13–71.73, p=0.014). Our analysis also identified a new FABP5 SNP (nSNP) that showed a significant association with T2DM in NHW (OR: 0.44, 95% CI: 0.19–0.99, p=0.045) and AA (OR: 0.17, 95% CI: 0.03–0.80, p=0.016). The Ala54Thr FABP2 polymorphism was significantly associated with T2DM in HA individuals only (OR: 1.85, 95% CI: 1.05–3.27, p=0.032). All other FABP SNPs did not show association with T2DM. These findings suggest a potential distinct role of SNPs in FABP5, 2 genes in T2DM in different populations. PMID:21288588

  4. Fatty Acid Binding Protein 4 Deficiency Protects against Oxygen-Induced Retinopathy in Mice

    PubMed Central

    Saint-Geniez, Magali; Ghelfi, Elisa; Liang, Xiaoliang; Yu, Chenwei; Spencer, Carrie; Abend, Stephanie; Hotamisligil, Gokhan; Cataltepe, Sule

    2014-01-01

    Retinopathy of prematurity (ROP) is a leading cause of blindness in children worldwide due to increasing survival rates of premature infants. Initial suppression, followed by increased production of the retinal vascular endothelial growth factor-A (VEGF) expression are key events that trigger the pathological neovascularization in ROP. Fatty acid binding protein 4 (FABP4) is an intracellular lipid chaperone that is induced by VEGF in a subset of endothelial cells. FABP4 exhibits a pro-angiogenic function in cultured endothelial cells and in airway microvasculature, but whether it plays a role in modulation of retinal angiogenesis is not known. We hypothesized that FABP4 deficiency could ameliorate pathological retinal vascularization and investigated this hypothesis using a well-characterized mouse model of oxygen-induced retinopathy (OIR). We found that FABP4 was not expressed in retinal vessels, but was present in resident macrophages/microglial cells and endothelial cells of the hyaloid vasculature in the immature retina. While FABP4 expression was not required for normal development of retinal vessels, FABP4 expression was upregulated and localized to neovascular tufts in OIR. FABP4−/− mice demonstrated a significant decrease in neovessel formation as well as a significant improvement in physiological revascularization of the avascular retinal tissues. These alterations in retinal vasculature were accompanied by reduced endothelial cell proliferation, but no effect on apoptosis or macrophage/microglia recruitment. FABP4−/− OIR samples demonstrated decreased expression of genes involved in angiogenesis, such as Placental Growth Factor, and angiopoietin 2. Collectively, our findings suggest FABP4 as a potential target of pathologic retinal angiogenesis in proliferative retinopathies. PMID:24802082

  5. Fatty acid-binding proteins and peribronchial angiogenesis in bronchopulmonary dysplasia.

    PubMed

    Ghelfi, Elisa; Karaaslan, Cagatay; Berkelhamer, Sara; Akar, Serra; Kozakewich, Harry; Cataltepe, Sule

    2011-09-01

    Inflammation plays a key role in the pathogenesis of bronchopulmonary dysplasia (BPD). Fatty acid-binding proteins (FABPs) 4 and 5 regulate the inflammatory activity of macrophages. Whether FABPs 4 and 5 could play a role in the pathogenesis of BPD via the promotion of macrophage inflammatory activity is unknown. This study sought to examine whether the expression levels of FABP4 and FABP5 were altered in bronchoalveolar lavage fluid and lung tissue in a baboon model of BPD. This study also sought to characterize the cell types that express these proteins. Real-time PCR, immunoblotting, immunohistochemistry, and double immunofluorescence were used to examine the expression of FABPs in samples of BPD. Morphometric analysis was used to quantify FABP4-positive peribronchial blood vessels in lung sections. FABP4 was primarily expressed in macrophages in samples of BPD. In addition, FABP4 was expressed in the endothelial cells of blood vessels in peribronchial areas and the vasa vasorum, but not in the alveolar vasculature in samples of BPD. FABP4 concentrations were significantly increased in lungs and bronchoalveolar lavage fluid samples with BPD. An increased density of FABP4-positive peribronchial blood vessels was evident in both baboon and human BPD sections. FABP5 was expressed in several cell types, including alveolar epithelial cells and macrophages. FABP5 concentrations did not show any significant alterations in BPD. In conclusion, FABP4 but not FABP5 levels are increased in BPD. FABP4 is differentially expressed in endothelial cells of the bronchial microvasculature, which demonstrates a previously unrecognized expansion in BPD. PMID:21177979

  6. Fatty acid binding protein 4 deficiency protects against oxygen-induced retinopathy in mice.

    PubMed

    Saint-Geniez, Magali; Ghelfi, Elisa; Liang, Xiaoliang; Yu, Chenwei; Spencer, Carrie; Abend, Stephanie; Hotamisligil, Gokhan; Cataltepe, Sule

    2014-01-01

    Retinopathy of prematurity (ROP) is a leading cause of blindness in children worldwide due to increasing survival rates of premature infants. Initial suppression, followed by increased production of the retinal vascular endothelial growth factor-A (VEGF) expression are key events that trigger the pathological neovascularization in ROP. Fatty acid binding protein 4 (FABP4) is an intracellular lipid chaperone that is induced by VEGF in a subset of endothelial cells. FABP4 exhibits a pro-angiogenic function in cultured endothelial cells and in airway microvasculature, but whether it plays a role in modulation of retinal angiogenesis is not known. We hypothesized that FABP4 deficiency could ameliorate pathological retinal vascularization and investigated this hypothesis using a well-characterized mouse model of oxygen-induced retinopathy (OIR). We found that FABP4 was not expressed in retinal vessels, but was present in resident macrophages/microglial cells and endothelial cells of the hyaloid vasculature in the immature retina. While FABP4 expression was not required for normal development of retinal vessels, FABP4 expression was upregulated and localized to neovascular tufts in OIR. FABP4-/- mice demonstrated a significant decrease in neovessel formation as well as a significant improvement in physiological revascularization of the avascular retinal tissues. These alterations in retinal vasculature were accompanied by reduced endothelial cell proliferation, but no effect on apoptosis or macrophage/microglia recruitment. FABP4-/- OIR samples demonstrated decreased expression of genes involved in angiogenesis, such as Placental Growth Factor, and angiopoietin 2. Collectively, our findings suggest FABP4 as a potential target of pathologic retinal angiogenesis in proliferative retinopathies. PMID:24802082

  7. Fatty acid-binding protein 5 limits the anti-inflammatory response in murine macrophages.

    PubMed

    Moore, Sherri M; Holt, Vivian V; Malpass, Lillie R; Hines, Ian N; Wheeler, Michael D

    2015-10-01

    The beginning stages of liver damage induced by various etiologies (i.e. high fat diet, alcohol consumption, toxin exposure) are characterized by abnormal accumulation of lipid in liver. Alterations in intracellular lipid transport, storage, and metabolism accompanied by cellular insult within the liver play an important role in the pathogenesis of liver disease, often involving a sustained inflammatory response. The intracellular lipid transporter, fatty acid binding protein 5 (FABP5), is highly expressed in macrophages and may play an important role in the hepatic inflammatory response after endotoxin exposure in mice. This study tested the hypothesis that FABP5 regulates macrophage response to LPS in male C57bl/6 (wild type) and FABP5 knockout mice, both in vitro and in vivo. Treatment with LPS revealed that loss of FABP5 enhances the number of hepatic F4/80(+) macrophages in the liver despite limited liver injury. Conversely, FABP5 knock out mice display higher mRNA levels of anti-inflammatory cytokines IL-10, arginase, YM-1, and Fizz-1 in liver compared to wild type mice. Bone marrow derived macrophages stimulated with inflammatory (LPS and IFN-γ) or anti-inflammatory (IL-4) mediators also showed significantly higher expression of anti-inflammatory/regulatory factors. These findings reveal a regulatory role of FABP5 in the acute inflammatory response to LPS-induced liver injury, which is consistent with the principle finding that FABP5 is a regulator of macrophage phenotype. Specifically, these findings demonstrate that loss of FABP5 promotes a more anti-inflammatory response. PMID:26105806

  8. Clinical significance of urinary liver-type fatty acid binding protein at various stages of nephropathy.

    PubMed

    Viswanathan, V; Sivakumar, S; Sekar, V; Umapathy, D; Kumpatla, S

    2015-01-01

    This cross-sectional study was to evaluate the levels of urinary liver-type fatty acid binding protein (u-LFABP pg/mg urine creatinine ratio) at different stages of diabetic nephropathy and to see its correlation with other clinical parameters in South Indian patients with type 2 diabetes mellitus (T2DM). A total of 65 (M: F; 42:23) T2DM subjects were divided into three groups, and were compared with 13 (M: F; 3:10) nondiabetic controls. The study groups were as follows: normoalbuminuric (n = 22), microalbuminuric (n = 22) and macroalbuminuric (n = 21). Estimated glomerular filtration rate (eGFR) was calculated using Cockcroft and Gault formula. u-LFABP levels in spot urine samples were measured with a solid phase enzyme linked immunosorbent assay. This study showed that u-LFABP levels were undetectable in healthy controls and was very low in the normoalbuminuric subjects. Elevated levels of u-LFABP are evident from the microalbuminuric stage indicating tubular damage. The levels of u-LFABP increased gradually with declining renal function. Geometric mean (95% confidence interval) for normoalbuminuria was 0.65 (0.47-0.97), microalbuminuria was 0.99 (0.55-1.97) and macroalbuminuria was 5.16 (1.8-14.5), (P = 0.005). In conclusion, u-LFABP levels were elevated in patients with reduced eGFR and showed a positive correlation with systolic blood pressure and protein to creatinine ratio in the total study subjects. PMID:26628791

  9. Clinical significance of urinary liver-type fatty acid binding protein at various stages of nephropathy

    PubMed Central

    Viswanathan, V.; Sivakumar, S.; Sekar, V.; Umapathy, D.; Kumpatla, S.

    2015-01-01

    This cross-sectional study was to evaluate the levels of urinary liver-type fatty acid binding protein (u-LFABP pg/mg urine creatinine ratio) at different stages of diabetic nephropathy and to see its correlation with other clinical parameters in South Indian patients with type 2 diabetes mellitus (T2DM). A total of 65 (M: F; 42:23) T2DM subjects were divided into three groups, and were compared with 13 (M: F; 3:10) nondiabetic controls. The study groups were as follows: normoalbuminuric (n = 22), microalbuminuric (n = 22) and macroalbuminuric (n = 21). Estimated glomerular filtration rate (eGFR) was calculated using Cockcroft and Gault formula. u-LFABP levels in spot urine samples were measured with a solid phase enzyme linked immunosorbent assay. This study showed that u-LFABP levels were undetectable in healthy controls and was very low in the normoalbuminuric subjects. Elevated levels of u-LFABP are evident from the microalbuminuric stage indicating tubular damage. The levels of u-LFABP increased gradually with declining renal function. Geometric mean (95% confidence interval) for normoalbuminuria was 0.65 (0.47–0.97), microalbuminuria was 0.99 (0.55–1.97) and macroalbuminuria was 5.16 (1.8–14.5), (P = 0.005). In conclusion, u-LFABP levels were elevated in patients with reduced eGFR and showed a positive correlation with systolic blood pressure and protein to creatinine ratio in the total study subjects. PMID:26628791

  10. Sex Differences in Long Chain Fatty Acid Utilization and Fatty Acid Binding Protein Concentration in Rat Liver

    PubMed Central

    Ockner, Robert K.; Burnett, David A.; Lysenko, Nina; Manning, Joan A.

    1979-01-01

    Female sex and estrogen administration are associated with increased hepatic production of triglyceride-rich lipoproteins; the basis for this has not been fully elucidated. Inasmuch as hepatic lipoprotein production is also influenced by FFA availability and triglyceride biosynthesis, we investigated sex differences in FFA utilization in rat hepatocyte suspensions and in the components of the triglyceride biosynthetic pathway. Isolated adult rat hepatocyte suspensions were incubated with albumin-bound [14C]oleate for up to 15 min. At physiological and low oleate concentrations, cells from females incorporated significantly more 14C into glycerolipids, especially triglycerides, and into oxidation products than did male cells, per milligram cell protein. At 0.44 mM oleate, incorporation into triglycerides in female cells was approximately twice that in male cells. Comparable sex differences were observed in cells from fasted animals and when [14C]-glycerol incorporation was measured. At higher oleate concentrations, i.e., fatty acid:albumin mole ratios in excess of 2:1, these sex differences were no longer demonstrable, suggesting that maximal rates of fatty acid esterification and oxidation were similar in female and male cells. In female and male hepatic microsomes, specific activities of long chain acyl coenzyme A synthetase, phosphatidate phosphohydrolase, and diglyceride acyltransferase were similar, but glycerol-3-phosphate acyltransferase activity was slightly greater in females at certain substrate concentrations. Microsomal incorporation of [14C]oleate into total glycerolipids was not significantly greater in females. In further contrast to intact cells, microsomal incorporation of [14C]oleate into triglycerides, although significantly greater in female microsomes, accounted for only a small fraction of the fatty acid esterified. The binding affinity and stoichiometry of partially purified female hepatic fatty acid binding protein (FABP) were similar to

  11. Fatty acid binding protein 7 and n-3 poly unsaturated fatty acid supply in early rat brain development.

    PubMed

    Maximin, Elise; Langelier, Bénédicte; Aïoun, Josiane; Al-Gubory, Kaïs H; Bordat, Christian; Lavialle, Monique; Heberden, Christine

    2016-03-01

    Fatty acid binding protein 7 (FABP7), abundant in the embryonic brain, binds with the highest affinity to docosahexaenoic acid (DHA) and is expressed in the early stages of embryogenesis. Here, we have examined the consequences of the exposure to different DHA levels and of the in utero depletion of FABP7 on early rat brain development. Neurodevelopment was evaluated through the contents of two proteins, connexin 43 (Cx43) and cyclin-dependent kinase 5 (CDK5), both involved in neuroblast proliferation, differentiation, and migration. The dams were fed with diets presenting different DHA contents, from deficiency to supplementation. DHA brain embryos contents already differed at embryonic day 11.5 and the differences kept increasing with time. Cx43 and CDK5 contents were positively associated with the brain DHA levels. When FABP7 was depleted in vivo by injections of siRNA in the telencephalon, the enhancement of the contents of both proteins was lost in supplemented animals, but FABP7 depletion did not modify phospholipid compositions regardless of the diets. Thus, FABP7 is a necessary mediator of the effect of DHA on these proteins synthesis, but its role in DHA uptake is not critical, although FABP7 is localized in phospholipid-rich areas. Our study shows that high contents of DHA associated with FABP7 are necessary to promote early brain development, which prompted us to recommend DHA supplementation early in pregnancy. PMID:26037116

  12. Effects of fatty acids and growth hormone on liver fatty acid binding protein and PPARalpha in rat liver.

    PubMed

    Carlsson, L; Lindén, D; Jalouli, M; Oscarsson, J

    2001-10-01

    The aim of this study was to investigate the interaction between long-chain fatty acids (LCFA) and growth hormone (GH) in the regulation of liver fatty acid binding protein (LFABP) and peroxisome proliferator-activated receptor-alpha (PPARalpha). Cultured rat hepatocytes were given oleic acid (OA; 500 microM) and GH (100 ng/ml) for 3 days. LFABP mRNA increased 3.6-fold by GH and 5.7-fold by OA, and combined incubation with GH and OA increased LFABP mRNA 17.6-fold. PPARalpha mRNA was decreased 50% by GH, but OA had no effect. Hypophysectomized (Hx) female rats were treated with L-thyroxine, cortisol, GH, and dietary fat for 7 days. PPARalpha mRNA levels were three- to fourfold higher in Hx than in normal female rats. GH decreased PPARalpha mRNA 50% in Hx rats. Dietary triglycerides (10% corn oil) increased LFABP mRNA and cytosolic LFABP about twofold but had no effect on PPARalpha mRNA in Hx rats. GH and dietary triglycerides had an additive effect on LFABP expression. Dietary triglycerides increased mitochondrial hydroxymethylglutaryl-CoA synthase mRNA only in the presence of GH. The diet increased serum triglycerides in Hx rats, and GH treatment prevented this increase. Addition of cholesterol to the diet did not influence LFABP levels but mitigated increased hepatic triglyceride content. In summary, these studies show that GH regulates LFABP expression independently of PPARalpha. Moreover, GH has different effects on PPARalpha-responsive genes and does not counteract the effect of LCFA on the expression of these gene products. PMID:11551854

  13. Concomitant increase in hepatic triacylglycerol biosynthesis and cytosolic fatty-acid-binding-protein content after feeding rats with a cholestyramine-containing diet.

    PubMed Central

    Kempen, H J; Glatz, J F; de Lange, J; Veerkamp, J H

    1983-01-01

    Cholestyramine feeding of rats increased the rate of palmitate and glycerol incorporation into triacylglycerols of isolated hepatocytes. Concomitantly an increase of fatty-acid binding by hepatic cytosolic proteins was observed, which could be attributed to an elevation of the content of the fatty-acid-binding protein (Mr 12000). The involvement of this protein in cholesterol, bile-acid and triacylglycerol metabolism is discussed. PMID:6661214

  14. In vitro bile acid binding and short-chain fatty acid profile of flax fiber and ethanol co-products.

    PubMed

    Fodje, Adele M L; Chang, Peter R; Leterme, Pascal

    2009-10-01

    Fibers from flaxseed and co-products from ethanol production could be potential sources of dietary fiber in human diet. In vitro fermentation and bile acid binding models were used to investigate the metabolic effects of lignaMax (Bioriginal Food and Science Corp., Saskatoon, SK, Canada) flax meal, spent flax meal, soluble flax gum, wheat insoluble fiber (WIF), and rye insoluble fiber (RIF). Wheat and rye bran were used as reference samples. Bile acid binding of substrates was analysed at taurocholate ([(14)C]taurocholate) concentration of 12.5 mM. Soluble flax gum showed the highest bile acid binding (0.57 micromol/mg of fiber) (P acid binding between wheat bran (0.2 micromol/mg of fiber) and WIF (0.26 micromol/mg of fiber). RIF had higher (P acid binding (0.20 micromol/mg of fiber) than rye bran (0.13 micromol/mg of fiber). Substrates were hydrolyzed and incubated with pig fecal samples. Short-chain fatty acid (SCFA) profile and gas accumulation (G(f)) were compared. Soluble flax gum generated the highest amount of acetic and propionic acids. SCFA profiles of wheat/rye brans and WIF/RIF were similar (except for butyric acid). G(f) for soluble flax gum was greater (P < .001) than that of spent flax meal. G(f) values of the wheat samples were similar, whereas the G(f) of the rye bran was higher (P < .001) than that of RIF. Fractional degradation rate (micro(t = T/2)) (P < .001) was also recorded. The highest mu(t = T/2) was observed for the soluble flax gum. Oil-depleted flaxseed fractions and WIF/RIF (co-products from ethanol production) could be potential sources of dietary fiber in human nutrition. PMID:19857071

  15. Serum complements and heart fatty acid binding protein in Bangladeshi patients with acute myocardial infarction

    PubMed Central

    Akhtar, Nayareen; Taher, Abu; Rahman, Rezwanur; Chowdhury, Ashesh Kumar

    2012-01-01

    The complement system is activated following acute myocardial infarction (AMI). Heart fatty acid binding protein (H-FABP) is a sensitive early biomarker of myocardial necrosis that can be used to confirm or exclude a diagnosis of AMI and to monitor recurrent infarction. This study was designed to detect changes in C3, C4 and H-FABP after AMI. Forty patients with AMI and a control group of 40 apparently healthy people were included. Selections were based on inclusion and exclusion criteria. The baseline characteristics were not significantly different between the groups. Patients’ blood samples were collected within 12 h of admission. Significant increases in C3 (AMI group 1.4260+0.04, healthy group 1.26040+0.04; p<0.05), C4 (AMI group 0.29305±0.013, healthy group 0.20860±0.012; p<0.05) and H-FABP (AMI group 12.3±1.69, healthy group 0.16±0.057; p<0.001) were seen in patients with AMI. The correlation between serum C3 and body mass index (BMI, r=0.33; p<0.05), serum C4 and BMI(r=0.313; p<0.05), serum C3 and total cholesterol high density lipoprotein (HDL, r=0.32; p<0.05), serum C4 and HbA1C (r=0.335; p<0.05) and serum C3 and troponin I (r= 0.325p<0.05) was found to be significant. But the correlation between serum C3 and waist:hip ratio (p=0.56), serum C4 and waist:hip ratio (p=0.83), serum C4 and total cholesterol HDL (p=0.993), serum C3 and HbA1C (p=0.440), serum C3 and random blood sugar (p=0.563), serum C4 and random blood sugar (p=0.828) and serum C4 and troponin I (p=0.373) was not significant. The significant complement activation detected in the plasma of patients with AMI indicated that complement plays a part in the pathogenesis of myocardial infarction. A significant increase of H-FABP improves the diagnosis of AMI.

  16. Isolation and immunological characterization of fatty acid binding protein isoforms from Fasciola hepatica.

    PubMed

    Espino, A M; Rodríguez Medina, J R; Hillyer, G V

    2001-10-01

    A combination of molecular sieving chromatography and 2-step preparative isoelectric focusing showed that native Fh12, a fatty acid-binding protein isolated from Fasciola hepatica adult worms, is a protein complex of at least 8 isoforms with identical molecular mass but different isoelectric points. Using enzyme-linked immunosorbent assay (ELISA) and inhibition ELISA assays, immunological differences were observed between native (nFh12) and a recombinant molecule denoted rFh15 that was obtained after screening a cDNA library from F. hepatica adult worms with an anti-Fh12 monospecific polyclonal antibody. It was confirmed that in infected rabbits, antibodies to nFh12 appear by the second week postinfection, whereas antibodies to rFh15 appear much later, by 6 wk postinfection. Four acidic forms (Fh12(1-4)) showed more immunological identity with rFh15 than with nFh12, based on the observation that they inhibited ELISA activity by nearly 50% when they were added to the anti-rFh15 polyclonal antibody at 20 microg/ml of protein concentration. Moreover, the Fh12(1-4) isoforms were poorly reactive with sera from rabbits 2-4 wk postinfection. However, the 2 acidic forms, denoted Fh12(5) and Fh12(6), and the neutral/basic forms, denoted Fh12(7) and Fh12(8), showed more immunological identity with the native nFh12 molecule than with the recombinant rFh15 because they were highly reactive with sera of rabbits with early 2-wk F. hepatica infection and inhibited ELISA activity nearly 50% when they were quantitatively added to the anti-nFh12 polyclonal antibody. These results suggest that rFh15 could be one of the acidic forms of nFh12, and that it, in fact, may be one of the less immunogenic or immunoprotective members, or both, of the nFh12 protein complex. PMID:11695360

  17. Association Between Serum Levels of Adipocyte Fatty Acid-binding Protein and Free Thyroxine

    PubMed Central

    Tseng, Fen-Yu; Chen, Pei-Lung; Chen, Yen-Ting; Chi, Yu-Chao; Shih, Shyang-Ron; Wang, Chih-Yuan; Chen, Chi-Ling; Yang, Wei-Shiung

    2015-01-01

    Abstract Adipocyte fatty acid-binding protein (AFABP) has been shown to be a biomarker of body weight change and atherosclerosis. Changes in thyroid function are associated with changes in body weight and risks of cardiovascular diseases. The association between AFABP and thyroid function status has been seldom evaluated. The aim of this study was to compare the serum AFABP concentrations in hyperthyroid patients and those in euthyroid individuals, and to evaluate the associations between serum AFABP and free thyroxine (fT4) levels. For this study, 30 hyperthyroid patients and 30 euthyroid individuals at a referral medical center were recruited. The patients with hyperthyroidism were treated with antithyroid regimens as clinically indicated. No medication was given to the euthyroid individuals. The body weight, body mass index, thyroid function, serum levels of AFABP, and biochemical data of both groups at baseline and at the 6th month were compared. Associations between AFABP and fT4 levels were also analyzed. At the baseline, the hyperthyroid patients had significantly higher serum AFABP levels than the euthyroid individuals (median [Q1, Q3]: 22.8 [19.4, 30.6] ng/mL vs 18.6 [15.3, 23.2] ng/mL; P = 0.038). With the antithyroid regimens, the AFABP serum levels of the hyperthyroid patients decreased to 16.6 (15.0, 23.9) ng/mL at the 6th month. No difference in the AFABP level was found between the hyperthyroid and the euthyroid groups at the 6th month. At baseline, sex (female vs male, ß = 7.65, P = 0.022) and fT4 level (ß = 2.51, P = 0.018) were significantly associated with AFABP levels in the univariate regression analysis. At the 6th month, sex and fT4 level (ß = 8.09, P < 0.001 and ß = 3.61, P = 0.005, respectively) were also significantly associated with AFABP levels. The associations between sex and fT4 level with AFABP levels remained significant in the stepwise multivariate regression analysis, both at baseline and at

  18. Liver fatty acid binding protein is the mitosis-associated polypeptide target of a carcinogen in rat hepatocytes.

    PubMed Central

    Bassuk, J A; Tsichlis, P N; Sorof, S

    1987-01-01

    Hepatocytes in normal rat liver were found previously to contain a cytoplasmic 14,000-dalton polypeptide (p14) that is associated with mitosis and is the principal early covalent target of activated metabolites of the carcinogen N-2-fluorenylacetamide (2-acetylaminofluorene). The level of immunohistochemically detected p14 was low when growth activity of hepatocytes was low, was markedly elevated during mitosis in normal and regenerating livers, but was very high throughout interphase during proliferation of hyperplastic and malignant hepatocytes induced in rat liver by a carcinogen (N-2-fluorenylacetamide or 3'-methyl-4-dimethylaminoazobenzene). We report here that p14 is the liver fatty acid binding protein. The nucleotide sequence of p14 cDNA clones, isolated by screening a rat liver cDNA library in bacteriophage lambda gt11 using p14 antiserum, was completely identical to part of the sequence reported for liver fatty acid binding protein. Furthermore, the two proteins shared the following properties: size of mRNA, amino acid composition, molecular size according to NaDodSO4 gel electrophoresis, and electrophoretic mobilities in a Triton X-100/acetic acid/urea gel. Their pI values overlapped in 2-dimensional isoelectric focusing/NaDodSO4 gel electrophoresis and showed the same response to delipidation. Either polypeptide reacted with and blocked the antiserum raised against the other polypeptide. The two polypeptides bound oleic acid similarly. Finally, identical elevations of cytoplasmic immunostain were detected specifically in mitotic hepatocytes with either antiserum. The collected findings are suggestive that liver fatty acid binding protein may carry ligands that promote hepatocyte division and may transport certain activated chemical carcinogens. Images PMID:3478711

  19. Proteomic Upregulation of Fatty Acid Synthase and Fatty Acid Binding Protein 5 and Identification of Cancer- and Race-Specific Pathway Associations in Human Prostate Cancer Tissues

    PubMed Central

    Myers, Jennifer S.; von Lersner, Ariana K.; Sang, Qing-Xiang Amy

    2016-01-01

    Protein profiling studies of prostate cancer have been widely used to characterize molecular differences between diseased and non-diseased tissues. When combined with pathway analysis, profiling approaches are able to identify molecular mechanisms of prostate cancer, group patients by cancer subtype, and predict prognosis. This strategy can also be implemented to study prostate cancer in very specific populations, such as African Americans who have higher rates of prostate cancer incidence and mortality than other racial groups in the United States. In this study, age-, stage-, and Gleason score-matched prostate tumor specimen from African American and Caucasian American men, along with non-malignant adjacent prostate tissue from these same patients, were compared. Protein expression changes and altered pathway associations were identified in prostate cancer generally and in African American prostate cancer specifically. In comparing tumor to non-malignant samples, 45 proteins were significantly cancer-associated and 3 proteins were significantly downregulated in tumor samples. Notably, fatty acid synthase (FASN) and epidermal fatty acid-binding protein (FABP5) were upregulated in human prostate cancer tissues, consistent with their known functions in prostate cancer progression. Aldehyde dehydrogenase family 1 member A3 (ALDH1A3) was also upregulated in tumor samples. The Metastasis Associated Protein 3 (MTA3) pathway was significantly enriched in tumor samples compared to non-malignant samples. While the current experiment was unable to detect statistically significant differences in protein expression between African American and Caucasian American samples, differences in overrepresentation and pathway enrichment were found. Structural components (Cytoskeletal Proteins and Extracellular Matrix Protein protein classes, and Biological Adhesion Gene Ontology (GO) annotation) were overrepresented in African American but not Caucasian American tumors. Additionally, 5

  20. Interaction of LY171883 and other peroxisome proliferators with fatty-acid-binding protein isolated from rat liver.

    PubMed Central

    Cannon, J R; Eacho, P I

    1991-01-01

    Fatty-acid-binding protein (FABP) is a 14 kDa protein found in hepatic cytosol which binds and transports fatty acids and other hydrophobic ligands throughout the cell. The purpose of this investigation was to determine whether LY171883, a leukotriene D4 antagonist, and other peroxisome proliferators bind to FABP and displace an endogenous fatty acid. [3H]Oleic acid was used to monitor the elution of FABP during chromatographic purification. [14C]LY171883 had a similar elution profile when substituted in the purification, indicating a common interaction with FABP. LY171883 and its structural analogue, LY189585, as well as the hypolipidaemic peroxisome proliferators clofibric acid, ciprofibrate, bezafibrate and WY14,643, displaced [3H]oleic acid binding to FABP. Analogues of LY171883 that do not induce peroxisome proliferation only weakly displaced oleate binding. [3H]Ly171883 bound directly to FABP with a Kd of 10.8 microM, compared with a Kd of 0.96 microM for [3H]oleate. LY171883 binding was inhibited by LY189585, clofibric acid, ciprofibrate and bezafibrate. These findings demonstrate that peroxisome proliferators, presumably due to their structural similarity to fatty acids, are able to bind to FABP and displace an endogenous ligand from its binding site. Interaction of peroxisome proliferators with FABP may be involved in perturbations of fatty acid metabolism caused by these agents as well as in the development of the pleiotropic response of peroxisome proliferation. Images Fig. 2. PMID:1747111

  1. Fatty Acid Binding Proteins FABP9 and FABP10 Participate in Antibacterial Responses in Chinese Mitten Crab, Eriocheir sinensis

    PubMed Central

    Li, Shuang; Guo, Xiao-Nv; Wang, Juan; Gong, Ya-Nan; He, Lin; Wang, Qun

    2013-01-01

    Invertebrates rely solely on the innate immune system for defense against pathogens and other stimuli. Fatty acid binding proteins (FABP), members of the lipid binding proteins superfamily, play a crucial role in fatty acid transport and lipid metabolism and are also involved in gene expression induced by fatty acids. In the vertebrate immune system, FABP is involved in inflammation regulated by fatty acids through its interaction with peroxidase proliferator activate receptors (PPARs). However, the immune functions of FABP in invertebrates are not well characterized. For this reason, we investigated the immune functionality of two fatty acid binding proteins, Es-FABP9 and Es-FABP10, following lipopolysaccharide (LPS) challenge in the Chinese mitten crab (Eriocheir sinensis). An obvious variation in the expression of Es-FABP9 and Es-FABP10 mRNA in E. sinensis was observed in hepatopancreas, gills, and hemocytes post-LPS challenge. Recombinant proteins rEs-FABP9 and rEs-FABP10 exhibited distinct bacterial binding activity and bacterial agglutination activity against Escherichia coli and Staphylococcus aureus. Furthermore, bacterial growth inhibition assays demonstrated that rEs-FABP9 responds positively to the growth inhibition of Vibrio parahaemolyticuss and S. aureus, while rEs-FABP10 responds positively to the growth inhibition of Aeromonas hydrophila and Bacillus subtilis. Coating of agarose beads with recombinant rEs-FABP9 and rEs-FABP10 dramatically enhanced encapsulation of the beads by crab hemocytes in vitro. In conclusion, the data presented here demonstrate the participation of these two lipid metabolism-related proteins in the innate immune system of E. sinensis. PMID:23365646

  2. Molecular characterization, tissue expression, and polymorphism analysis of liver-type fatty acid binding protein in Landes geese.

    PubMed

    Song, Z; Shao, D; Sun, X X; Niu, J W; Gong, D Q

    2015-01-01

    Liver weight is an important economic trait in the fatty goose liver industry. Liver-type fatty acid binding protein (L-FABP) is involved in the formation and metabolism of fatty acids. Thus, we hypothesized that sequence polymorphisms in L-FABP were associated with fatty liver weight in goose. We first isolated, sequenced, and characterized the goose L-FABP gene, which had not been previously reported. The goose L-FABP gene was 2490 bp and included 4 exons coding for a 126-amino acid protein. Analysis of expression levels of the goose L-FABP gene in different tissues showed that the expression level in the liver tissue was higher than in other tissues, and was significantly higher in the liver tissue of overfed geese than in control geese. Moreover, a single nucleotide polymorphism located at 774 bp in the gene was identified in a Landes goose population. To test whether this single nucleotide polymorphism was associated with fatty liver production, liver weight and the ratio of liver to carcass weights were determined for the 3 genotypes with this single nucleotide polymorphism (TT, TG, GG) in overfed Landes geese. Our data indicate that individuals with the GG genotype had higher values for the variables measured than those with the other 2 genotypes, suggesting that L-FABP can be a selection marker for the trait of fatty liver production in goose. PMID:25729971

  3. A nuclear magnetic resonance-based structural rationale for contrasting stoichiometry and ligand binding site(s) in fatty acid-binding proteins.

    PubMed

    He, Yan; Estephan, Rima; Yang, Xiaomin; Vela, Adriana; Wang, Hsin; Bernard, Cédric; Stark, Ruth E

    2011-03-01

    Liver fatty acid-binding protein (LFABP) is a 14 kDa cytosolic polypeptide, differing from other family members in the number of ligand binding sites, the diversity of bound ligands, and the transfer of fatty acid(s) to membranes primarily via aqueous diffusion rather than direct collisional interactions. Distinct two-dimensional (1)H-(15)N nuclear magnetic resonance (NMR) signals indicative of slowly exchanging LFABP assemblies formed during stepwise ligand titration were exploited, without determining the protein-ligand complex structures, to yield the stoichiometries for the bound ligands, their locations within the protein binding cavity, the sequence of ligand occupation, and the corresponding protein structural accommodations. Chemical shifts were monitored for wild-type LFABP and an R122L/S124A mutant in which electrostatic interactions viewed as being essential to fatty acid binding were removed. For wild-type LFABP, the results compared favorably with the data for previous tertiary structures of oleate-bound wild-type LFABP in crystals and in solution: there are two oleates, one U-shaped ligand that positions the long hydrophobic chain deep within the cavity and another extended structure with the hydrophobic chain facing the cavity and the carboxylate group lying close to the protein surface. The NMR titration validated a prior hypothesis that the first oleate to enter the cavity occupies the internal protein site. In contrast, (1)H and (15)N chemical shift changes supported only one liganded oleate for R122L/S124A LFABP, at an intermediate location within the protein cavity. A rationale based on protein sequence and electrostatics was developed to explain the stoichiometry and binding site trends for LFABPs and to put these findings into context within the larger protein family. PMID:21226535

  4. Water-mediated recognition of simple alkyl chains by heart-type fatty-acid-binding protein.

    PubMed

    Matsuoka, Shigeru; Sugiyama, Shigeru; Matsuoka, Daisuke; Hirose, Mika; Lethu, Sébastien; Ano, Hikaru; Hara, Toshiaki; Ichihara, Osamu; Kimura, S Roy; Murakami, Satoshi; Ishida, Hanako; Mizohata, Eiichi; Inoue, Tsuyoshi; Murata, Michio

    2015-01-26

    Long-chain fatty acids (FAs) with low water solubility require fatty-acid-binding proteins (FABPs) to transport them from cytoplasm to the mitochondria for energy production. However, the precise mechanism by which these proteins recognize the various lengths of simple alkyl chains of FAs with similar high affinity remains unknown. To address this question, we employed a newly developed calorimetric method for comprehensively evaluating the affinity of FAs, sub-Angstrom X-ray crystallography to accurately determine their 3D structure, and energy calculations of the coexisting water molecules using the computer program WaterMap. Our results clearly showed that the heart-type FABP (FABP3) preferentially incorporates a U-shaped FA of C10-C18 using a lipid-compatible water cluster, and excludes longer FAs using a chain-length-limiting water cluster. These mechanisms could help us gain a general understanding of how proteins recognize diverse lipids with different chain lengths. PMID:25491543

  5. Fatty Acid-binding Proteins Interact with Comparative Gene Identification-58 Linking Lipolysis with Lipid Ligand Shuttling.

    PubMed

    Hofer, Peter; Boeszoermenyi, Andras; Jaeger, Doris; Feiler, Ursula; Arthanari, Haribabu; Mayer, Nicole; Zehender, Fabian; Rechberger, Gerald; Oberer, Monika; Zimmermann, Robert; Lass, Achim; Haemmerle, Guenter; Breinbauer, Rolf; Zechner, Rudolf; Preiss-Landl, Karina

    2015-07-24

    The coordinated breakdown of intracellular triglyceride (TG) stores requires the exquisitely regulated interaction of lipolytic enzymes with regulatory, accessory, and scaffolding proteins. Together they form a dynamic multiprotein network designated as the "lipolysome." Adipose triglyceride lipase (Atgl) catalyzes the initiating step of TG hydrolysis and requires comparative gene identification-58 (Cgi-58) as a potent activator of enzyme activity. Here, we identify adipocyte-type fatty acid-binding protein (A-Fabp) and other members of the fatty acid-binding protein (Fabp) family as interaction partners of Cgi-58. Co-immunoprecipitation, microscale thermophoresis, and solid phase assays proved direct protein/protein interaction between A-Fabp and Cgi-58. Using nuclear magnetic resonance titration experiments and site-directed mutagenesis, we located a potential contact region on A-Fabp. In functional terms, A-Fabp stimulates Atgl-catalyzed TG hydrolysis in a Cgi-58-dependent manner. Additionally, transcriptional transactivation assays with a luciferase reporter system revealed that Fabps enhance the ability of Atgl/Cgi-58-mediated lipolysis to induce the activity of peroxisome proliferator-activated receptors. Our studies identify Fabps as crucial structural and functional components of the lipolysome. PMID:25953897

  6. Fatty Acid-binding Proteins Interact with Comparative Gene Identification-58 Linking Lipolysis with Lipid Ligand Shuttling*

    PubMed Central

    Hofer, Peter; Boeszoermenyi, Andras; Jaeger, Doris; Feiler, Ursula; Arthanari, Haribabu; Mayer, Nicole; Zehender, Fabian; Rechberger, Gerald; Oberer, Monika; Zimmermann, Robert; Lass, Achim; Haemmerle, Guenter; Breinbauer, Rolf; Zechner, Rudolf; Preiss-Landl, Karina

    2015-01-01

    The coordinated breakdown of intracellular triglyceride (TG) stores requires the exquisitely regulated interaction of lipolytic enzymes with regulatory, accessory, and scaffolding proteins. Together they form a dynamic multiprotein network designated as the “lipolysome.” Adipose triglyceride lipase (Atgl) catalyzes the initiating step of TG hydrolysis and requires comparative gene identification-58 (Cgi-58) as a potent activator of enzyme activity. Here, we identify adipocyte-type fatty acid-binding protein (A-Fabp) and other members of the fatty acid-binding protein (Fabp) family as interaction partners of Cgi-58. Co-immunoprecipitation, microscale thermophoresis, and solid phase assays proved direct protein/protein interaction between A-Fabp and Cgi-58. Using nuclear magnetic resonance titration experiments and site-directed mutagenesis, we located a potential contact region on A-Fabp. In functional terms, A-Fabp stimulates Atgl-catalyzed TG hydrolysis in a Cgi-58-dependent manner. Additionally, transcriptional transactivation assays with a luciferase reporter system revealed that Fabps enhance the ability of Atgl/Cgi-58-mediated lipolysis to induce the activity of peroxisome proliferator-activated receptors. Our studies identify Fabps as crucial structural and functional components of the lipolysome. PMID:25953897

  7. Heart type fatty acid binding protein response and subsequent development of atherosclerosis in insulin resistant polycystic ovary syndrome patients

    PubMed Central

    2012-01-01

    Background Women with polycystic ovary syndrome (PCOS) have higher risk for cardiovascular disease (CVD). Heart type fatty acid binding protein (HFABP) has been found to be predictive for myocardial ischemia.Wet ested whether HFABP is the predictor for CVD in PCOS patients, who have an increased risk of cardiovascular disease. Methods This was a prospective, cross sectional controlled study conducted in a training and research hospital.The study population consisted of 46 reproductive-age PCOS women and 28 control subjects. We evaluated anthropometric and metabolic parameters, carotid intima media thickness and HFABP levels in both PCOS patients and control group. Results Mean fasting insulin, homeostasis model assessment insulin resistance index (HOMA-IR), triglyceride, total cholesterol, low density lipoprotein cholesterol, free testosterone, total testosterone, carotid intima media thickness (CIMT) levels were significantly higher in PCOS patients. Although HFABP levels were higher in PCOS patients, the difference did not reach statistically significant in early age groups. After adjustment for age and body mass index, HFABP level was positive correlated with hsCRP, free testosterone levels, CIMT and HOMA-IR. Conclusions Heart type free fatty acid binding protein appeared to have an important role in metabolic response and subsequent development of atherosclerosis in insulin resistant, hyperandrogenemic PCOS patients. PMID:23249450

  8. Identification of a functional polymorphism at the Adipose Fatty Acid Binding protein gene (FABP4) and demonstration of its association with cardiovascular disease: A path to follow

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Fatty acid binding proteins (FABPs) are proteins that reversibly bind fatty acids and other lipids. So far, 9 tissue-specific cytoplasmic FABPs have been identified. Adipose tissue FABP (FABP4) has been suggested to be a bridge between inflammation and other pathways related to the metabolic syndrom...

  9. A Photocytes-Associated Fatty Acid-Binding Protein from the Light Organ of Adult Taiwanese Firefly, Luciola cerata

    PubMed Central

    Goh, King-Siang; Li, Chia-Wei

    2011-01-01

    Background Intracellular fatty acid-binding proteins (FABPs) are considered to be an important energy source supplier in lipid metabolism; however, they have never been reported in any bioluminescent tissue before. In this study, we determined the structural and functional characteristics of a novel FABP (lcFABP) from the light organ of adult Taiwanese firefly, Luciola cerata, and showed anatomical association of lcFABP with photocytes. Principal Findings Our results demonstrated the primary structure of lcFABP deduced from the cDNA clone of light organ shares structural homologies with other insect and human FABPs. In vitro binding assay indicated the recombinant lcFABP binds saturated long chain fatty acids (C14-C18) more strongly than other fatty acids and firefly luciferin. In addition, tissue distribution screening assay using a rabbit antiserum specifically against the N-terminal sequence of lcFABP confirmed the light organ-specific expression of lcFABP. In the light organ, the lcFABP constituted about 15% of total soluble proteins, and was detected in both cytosol and nucleus of photocytes. Conclusions The specific localization of abundant lcFABP in the light organ suggests that sustained bioluminescent flashes in the light organ might be a high energy demanding process. In photocytes, lcFABP might play a key role in providing long chain fatty acids to peroxisomes for the luciferase-catalyzed long chain acyl-CoA synthetic reaction. PMID:22242133

  10. CD36 Binds Oxidized Low Density Lipoprotein (LDL) in a Mechanism Dependent upon Fatty Acid Binding*

    PubMed Central

    Jay, Anthony G.; Chen, Alexander N.; Paz, Miguel A.; Hung, Justin P.; Hamilton, James A.

    2015-01-01

    The association of unesterified fatty acid (FA) with the scavenger receptor CD36 has been actively researched, with focuses on FA and oxidized low density lipoprotein (oxLDL) uptake. CD36 has been shown to bind FA, but this interaction has been poorly characterized to date. To gain new insights into the physiological relevance of binding of FA to CD36, we characterized FA binding to the ectodomain of CD36 by the biophysical method surface plasmon resonance. Five structurally distinct FAs (saturated, monounsaturated (cis and trans), polyunsaturated, and oxidized) were pulsed across surface plasmon resonance channels, generating association and dissociation binding curves. Except for the oxidized FA HODE, all FAs bound to CD36, with rapid association and dissociation kinetics similar to HSA. Next, to elucidate the role that each FA might play in CD36-mediated oxLDL uptake, we used a fluorescent oxLDL (Dii-oxLDL) live cell assay with confocal microscopy imaging. CD36-mediated uptake in serum-free medium was very low but greatly increased when serum was present. The addition of exogenous FA in serum-free medium increased oxLDL binding and uptake to levels found with serum and affected CD36 plasma membrane distribution. Binding/uptake of oxLDL was dependent upon the FA dose, except for docosahexaenoic acid, which exhibited binding to CD36 but did not activate the uptake of oxLDL. HODE also did not affect oxLDL uptake. High affinity FA binding to CD36 and the effects of each FA on oxLDL uptake have important implications for protein conformation, binding of other ligands, functional properties of CD36, and high plasma FA levels in obesity and type 2 diabetes. PMID:25555908

  11. Molecular dynamic simulations reveal the structural determinants of fatty acid binding to oxy-myoglobin

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The mechanism(s) by which fatty acids are sequestered and transported in muscle have not been fully elucidated. A potential key player in this process is the protein myoglobin (Mb). Indeed, there is a catalogue of empirical evidence supporting direct interaction of globins with fatty acid metabolite...

  12. The discovery of novel and selective fatty acid binding protein 4 inhibitors by virtual screening and biological evaluation.

    PubMed

    Zhou, Yang; Nie, Tao; Zhang, Yan; Song, Ming; Li, Kuai; Ding, Mengxiao; Ding, Ke; Wu, Donghai; Xu, Yong

    2016-09-15

    Adipocyte fatty acid binding protein (AFABP, FABP4) has been proven to be a potential therapeutic target for diabetes, atherosclerosis and inflammation-related diseases. In this study, a series of new scaffolds of small molecule inhibitors of FABP4 were identified by virtual screening and were validated by a bioassay. Fifty selected compounds were tested, which led to the discovery of seven hits. Structural similarity-based searches were then performed based on the hits and led to the identification of one high affinity compound 33b (Ki=0.29±0.07μM, ΔTm=8.5°C). This compound's effective blockade of inflammatory response was further validated by its ability to suppress pro-inflammatory cytokines induced by lipopolysaccharide (LPS) stimulation. Molecular dynamics simulation (MD) and mutagenesis studies validated key residues for its inhibitory potency and thus provide an important clue for the further development of drugs. PMID:27460668

  13. Fatty acid binding protein 4 is a target of VEGF and a regulator of cell proliferation in endothelial cells

    PubMed Central

    Elmasri, Harun; Karaaslan, Cagatay; Teper, Yaroslav; Ghelfi, Elisa; Weng, MeiQian; Ince, Tan A.; Kozakewich, Harry; Bischoff, Joyce; Cataltepe, Sule

    2009-01-01

    Fatty acid binding protein 4 (FABP4) plays an important role in maintaining glucose and lipid homeostasis. FABP4 has been primarily regarded as an adipocyte- and macrophage-specific protein, but recent studies suggest that it may be more widely expressed. We found strong FABP4 expression in the endothelial cells (ECs) of capillaries and small veins in several mouse and human tissues, including the heart and kidney. FABP4 was also detected in the ECs of mature human placental vessels and infantile hemangiomas, the most common tumor of infancy and ECs. In most of these cases, FABP4 was detected in both the nucleus and cytoplasm. FABP4 mRNA and protein levels were significantly induced in cultured ECs by VEGF-A and bFGF treatment. The effect of VEGF-A on FABP4 expression was inhibited by chemical inhibition or short-hairpin (sh) RNA-mediated knockdown of VEGF-receptor-2 (R2), whereas the VEGFR1 agonists, placental growth factors 1 and 2, had no effect on FABP4 expression. Knockdown of FABP4 in ECs significantly reduced proliferation both under baseline conditions and in response to VEGF and bFGF. Thus, FABP4 emerged as a novel target of the VEGF/VEGFR2 pathway and a positive regulator of cell proliferation in ECs.—Elmasri, H., Karaaslan, C., Teper, Y., Ghelfi, E., Weng, M., Ince, T. A., Kozakewich, H., Bischoff, J., Cataltepe, S. Fatty acid binding protein 4 is a target of VEGF and a regulator of cell proliferation in endothelial cells. PMID:19625659

  14. Fatty acid binding protein 4 expression marks a population of adipocyte progenitors in white and brown adipose tissues

    PubMed Central

    Shan, Tizhong; Liu, Weiyi; Kuang, Shihuan

    2013-01-01

    Adipose tissues regulate metabolism, reproduction, and life span. The development and growth of adipose tissue are due to increases of both adipocyte cell size and cell number; the latter is mediated by adipocyte progenitors. Various markers have been used to identify either adipocyte progenitors or mature adipocytes. The fatty acid binding protein 4 (FABP4), commonly known as adipocyte protein 2 (aP2), has been extensively used as a marker for differentiated adipocytes. However, whether aP2 is expressed in adipogenic progenitors is controversial. Using Cre/LoxP-based cell lineage tracing in mice, we have identified a population of aP2-expressing progenitors in the stromal vascular fraction (SVF) of both white and brown adipose tissues. The aP2-lineage progenitors reside in the adipose stem cell niche and express adipocyte progenitor markers, including CD34, Sca1, Dlk1, and PDGFRα. When isolated and grown in culture, the aP2-expressing SVF cells proliferate and differentiate into adipocytes upon induction. Conversely, ablation of the aP2 lineage greatly reduces the adipogenic potential of SVF cells. When grafted into wild-type mice, the aP2-lineage progenitors give rise to adipose depots in recipient mice. Therefore, the expression of aP2 is not limited to mature adipocytes, but also marks a pool of undifferentiated progenitors associated with the vasculature of adipose tissues. Our finding adds to the repertoire of adipose progenitor markers and points to a new regulator of adipose plasticity.—Shan, T., Liu, W., Kuang, S. Fatty acid-binding protein 4 expression marks a population of adipocyte progenitors in white and brown adipose tissues. PMID:23047894

  15. Peri-operative heart-type fatty acid binding protein is associated with acute kidney injury after cardiac surgery

    PubMed Central

    Schaub, Jennifer A.; Garg, Amit X.; Coca, Steven G.; Testani, Jeffrey M.; Shlipak, Michael G.; Eikelboom, John; Kavsak, Peter; McArthur, Eric; Shortt, Colleen; Whitlock, Richard; Parikh, Chirag R.

    2015-01-01

    Acute Kidney Injury (AKI) is a common complication after cardiac surgery and is associated with worse outcomes. Since heart fatty acid binding protein (H-FABP) is a myocardial protein that detects cardiac injury, we sought to determine if plasma H-FABP was associated with AKI in the TRIBE-AKI cohort; a multi-center cohort of 1219 patients at high risk for AKI who underwent cardiac surgery. The primary outcomes of interest were any AKI (Acute Kidney Injury Network (AKIN) stage 1 or higher) and severe AKI (AKIN stage 2 or higher). The secondary outcome was long-term mortality after discharge. Patients who developed AKI had higher levels of H-FABP pre- and post-operatively than patients who did not have AKI. In analyses adjusted for known AKI risk factors, first post-operative log(H-FABP) was associated with severe AKI (adjusted OR 5.39 [95% CI, 2.87-10.11] per unit increase), while pre-operative log(H-FABP) was associated with any AKI (2.07 [1.48-2.89]) and mortality (1.67 [1.17-2.37]). These relationships persisted after adjustment for change in serum creatinine (for first postoperative log(H-FABP)) and biomarkers of cardiac and kidney injury, including brain natriuretic peptide, cardiac troponin-I, interleukin-18, liver fatty acid binding protein, kidney injury molecule-1, and neutrophil gelatinase associated lipocalin. Thus, peri-operative plasma H-FABP levels may be used for risk-stratification of AKI and mortality following cardiac surgery. PMID:25830762

  16. The human intestinal fatty acid binding protein (hFABP2) gene is regulated by HNF-4{alpha}

    SciTech Connect

    Klapper, Maja . E-mail: klapper@molnut.uni-kiel.de; Boehme, Mike; Nitz, Inke; Doering, Frank

    2007-04-27

    The cytosolic human intestinal fatty acid binding protein (hFABP2) is proposed to be involved in intestinal absorption of long-chain fatty acids. The aim of this study was to investigate the regulation of hFABP2 by the endodermal hepatocyte nuclear factor 4{alpha} (HNF-4{alpha}), involved in regulation of genes of fatty acid metabolism and differentiation. Electromobility shift assays demonstrated that HNF-4{alpha} binds at position -324 to -336 within the hFABP2 promoter. Mutation of this HNF-4 binding site abolished the luciferase reporter activity of hFABP2 in postconfluent Caco-2 cells. In HeLa cells, this mutation reduced the activation of the hFABP2 promoter by HNF-4{alpha} by about 50%. Thus, binding element at position -336/-324 essentially determines the transcriptional activity of promoter and may be important in control of hFABP2 expression by dietary lipids and differentiation. Studying genotype interactions of hFABP2 and HNF-4{alpha}, that are both candidate genes for diabetes type 2, may be a powerful approach.

  17. The liver fatty acid binding protein--comparison of cavity properties of intracellular lipid-binding proteins.

    PubMed

    Thompson, J; Ory, J; Reese-Wagoner, A; Banaszak, L

    1999-02-01

    The crystal and solution structures of all of the intracellular lipid binding proteins (iLBPs) reveal a common beta-barrel framework with only small local perturbations. All existing evidence points to the binding cavity and a poorly delimited 'portal' region as defining the function of each family member. The importance of local structure within the cavity appears to be its influence on binding affinity and specificity for the lipid. The portal region appears to be involved in the regulation of ligand exchange. Within the iLBP family, liver fatty acid binding protein or LFABP, has the unique property of binding two fatty acids within its internalized binding cavity rather than the commonly observed stoichiometry of one. Furthermore, LFABP will bind hydrophobic molecules larger than the ligands which will associate with other iLBPs. The crystal structure of LFABP contains two bound oleate molecules and provides the explanation for its unusual stoichiometry. One of the bound fatty acids is completely internalized and has its carboxylate interacting with an arginine and two serines. The second oleate represents an entirely new binding mode with the carboxylate on the surface of LFABP. The two oleates also interact with each other. Because of this interaction and its inner location, it appears the first oleate must be present before the second more external molecule is bound. PMID:10331654

  18. Saturated fatty-acids regulate retinoic acid signaling and suppress tumorigenesis by targeting fatty-acid-binding protein 5

    PubMed Central

    Levi, Liraz; Wang, Zeneng; Doud, Mary Kathryn; Hazen, Stanley L.; Noy, Noa

    2015-01-01

    Long chain fatty acids (LCFA) serve as energy sources, components of cell membranes, and precursors for signalling molecules. Here we show that these biological compounds also regulate gene expression and that they do so by controlling the transcriptional activities of the retinoic acid (RA)-activated nuclear receptors RAR and PPARβ/δ. The data indicate that these activities of LCFA are mediated by FABP5 which delivers ligands from the cytosol to nuclear PPARβ/δ. Both saturated and unsaturated LCFA (SLCFA, ULCFA) bind to FABP5, thereby displacing RA and diverting it to RAR. However, while SLCFA inhibit, ULCFA activate the FABP5/PPARβ/δ pathway. We show further that, by concomitantly promoting activation of RAR and inhibiting the activation of PPARβ/δ, SLCFA suppress the oncogenic properties of FABP5-expressing carcinoma cells in cultured cells and in vivo. The observations suggest that compounds that inhibit FABP5 may constitute a new class of drugs for therapy of certain types of cancer. PMID:26592976

  19. Saturated fatty acids regulate retinoic acid signalling and suppress tumorigenesis by targeting fatty acid-binding protein 5.

    PubMed

    Levi, Liraz; Wang, Zeneng; Doud, Mary Kathryn; Hazen, Stanley L; Noy, Noa

    2015-01-01

    Long chain fatty acids (LCFA) serve as energy sources, components of cell membranes and precursors for signalling molecules. Here we show that these biological compounds also regulate gene expression and that they do so by controlling the transcriptional activities of the retinoic acid (RA)-activated nuclear receptors RAR and PPARβ/δ. The data indicate that these activities of LCFA are mediated by FABP5, which delivers ligands from the cytosol to nuclear PPARβ/δ. Both saturated and unsaturated LCFA (SLCFA, ULCFA) bind to FABP5, thereby displacing RA and diverting it to RAR. However, while SLCFA inhibit, ULCFA activate the FABP5/PPARβ/δ pathway. We show further that, by concomitantly promoting the activation of RAR and inhibiting the activation of PPARβ/δ, SLCFA suppress the oncogenic properties of FABP5-expressing carcinoma cells in cultured cells and in vivo. The observations suggest that compounds that inhibit FABP5 may constitute a new class of drugs for therapy of certain types of cancer. PMID:26592976

  20. Liver fatty acid-binding protein: specific mediator of the mitogenesis induced by two classes of carcinogenic peroxisome proliferators.

    PubMed Central

    Khan, S H; Sorof, S

    1994-01-01

    Peroxisome proliferators (PP) are a diverse group of chemicals that induce dramatic increases in peroxisomes in rodent hepatocytes, followed by hypertrophy, hepatomegaly, alterations in lipid metabolism, mitogenesis, and finally hepatocarcinomas. Termed nongenotoxic carcinogens, they do not interact with DNA, are not mutagenic in bacterial assays, and fail to elicit many of the phenotypes associated with classic genotoxic carcinogens. We report here that the mitogenesis induced by the major PP class, the amphipathic carboxylates, and by the tetrazole-substituted acetophenones specifically requires liver fatty acid-binding protein (L-FABP) in cultured rat hepatoma cells transfected with the sense cDNA of L-FABP, in contrast to L-FABP-nonexpressing cells transfected with its antisense cDNA. The mitogenic actions of L-FABP were protein-specific, inasmuch as no other protein in the nonexpressing cells could act like L-FABP. L-FABP was previously shown not only (i) to interact covalently with metabolites of the two genotoxic carcinogens 2-acetylaminofluorene and aminoazo dyes during liver carcinogenesis, but also (ii) to bind noncovalently the two classes of PP in vitro with avidities that correlate with their abilities to elicit peroxisomal enzymatic responses, and (iii) together with unsaturated fatty acids, especially linoleic acid, to promote multiplication of the transfected hepatoma cells in culture. The convergence of the two types of genotoxic carcinogens with the two classes of PP nongenotoxic carcinogens, and also with unsaturated fatty acids, at L-FABP actions in inducing mitogenesis allows the following hypothesis. During tumor promotion of carcinogenesis in vivo, these groups of genotoxic and nongenotoxic carcinogens act on the normal process by which L-FABP, functioning as a specific receptor of unsaturated fatty acids or their metabolites, promotes hepatocyte proliferation. Images PMID:8302856

  1. Low abdominal NIRS values and elevated plasma intestinal fatty acid-binding protein in a premature piglet model of necrotizing enterocolitis

    Technology Transfer Automated Retrieval System (TEKTRAN)

    To identify early markers of necrotizing enterocolitis (NEC), we hypothesized that continuous abdominal near-infrared spectroscopy (A-NIRS) measurement of splanchnic tissue oxygen saturation and intermittent plasma intestinal fatty-acid binding protein (pI-FABP) measured every 6 hours can detect NEC...

  2. Liver Fatty Acid Binding Protein Gene-ablation Exacerbates Weight Gain in High-Fat Fed Female Mice

    PubMed Central

    McIntosh, Avery L.; Atshaves, Barbara P.; Landrock, Danilo; Landrock, Kerstin K.; Martin, Gregory G.; Storey, Stephen M.; Kier, Ann B.; Schroeder, Friedhelm

    2013-01-01

    Loss of liver fatty acid binding protein (L-FABP) decreases long chain fatty acid uptake and oxidation in primary hepatocytes and in vivo. On this basis, L-FABP gene ablation would potentiate high-fat diet-induced weight gain and weight gain/energy intake. While this was indeed the case when L-FABP null (−/−) mice on the C57BL/6NCr background were pair-fed high fat diet, whether this would also be observed under high-fat diet fed ad libitum was not known. Therefore, this possibility was examined in female L-FABP (−/−) mice on the same background. L-FABP (−/−) mice consumed equal amounts of defined high-fat or isocaloric control diets fed ad libitum. However, on the ad libitum fed high-fat diet the L-FABP (−/−) mice exhibited: 1) Decreased hepatic long chain fatty acid (LCFA) β-oxidation as indicated by lower serum β–hydroxybutyrate level; 2) Decreased hepatic protein levels of key enzymes mitochondrial (rate limiting carnitine palmitoyl acyltransferase A1, CPT1A; HMG-CoA synthase) and peroxisomal (acyl CoA oxidase 1, ACOX1) LCFA β-oxidation; 3) Increased fat tissue mass (FTM) and FTM/energy intake to the greatest extent; and 4) Exacerbated body weight gain, weight gain/energy intake, liver weight, and liver weight/body weight to the greatest extent. Taken together, these findings showed that L-FABP gene-ablation exacerbated diet-induced weight gain and fat tissue mass gain in mice fed high-fat diet ad libitum—consistent with the known biochemistry and cell biology of L-FABP. PMID:23539345

  3. Comparative study of the fatty acid binding process of a new FABP from Cherax quadricarinatus by fluorescence intensity, lifetime and anisotropy.

    PubMed

    Li, Jiayao; Henry, Etienne; Wang, Lanmei; Delelis, Olivier; Wang, Huan; Simon, Françoise; Tauc, Patrick; Brochon, Jean-Claude; Zhao, Yunlong; Deprez, Eric

    2012-01-01

    Fatty acid-binding proteins (FABPs) are small cytosolic proteins, largely distributed in invertebrates and vertebrates, which accomplish uptake and intracellular transport of hydrophobic ligands such as fatty acids. Although long chain fatty acids play multiple crucial roles in cellular functions (structural, energy metabolism, regulation of gene expression), the precise functions of FABPs, especially those of invertebrate species, remain elusive. Here, we have identified and characterized a novel FABP family member, Cq-FABP, from the hepatopancreas of red claw crayfish Cherax quadricarinatus. We report the characterization of fatty acid-binding affinity of Cq-FABP by four different competitive fluorescence-based assays. In the two first approaches, the fluorescent probe 8-Anilino-1-naphthalenesulfonate (ANS), a binder of internal cavities of protein, was used either by directly monitoring its fluorescence emission or by monitoring the fluorescence resonance energy transfer occurring between the single tryptophan residue of Cq-FABP and ANS. The third and the fourth approaches were based on the measurement of the fluorescence emission intensity of the naturally fluorescent cis-parinaric acid probe or the steady-state fluorescence anisotropy measurements of a fluorescently labeled fatty acid (BODIPY-C16), respectively. The four methodologies displayed consistent equilibrium constants for a given fatty acid but were not equivalent in terms of analysis. Indeed, the two first methods were complicated by the existence of non specific binding modes of ANS while BODIPY-C16 and cis-parinaric acid specifically targeted the fatty acid binding site. We found a relationship between the affinity and the length of the carbon chain, with the highest affinity obtained for the shortest fatty acid, suggesting that steric effects primarily influence the interaction of fatty acids in the binding cavity of Cq-FABP. Moreover, our results show that the binding affinities of several fatty

  4. Comparative Study of the Fatty Acid Binding Process of a New FABP from Cherax quadricarinatus by Fluorescence Intensity, Lifetime and Anisotropy

    PubMed Central

    Li, Jiayao; Henry, Etienne; Wang, Lanmei; Delelis, Olivier; Wang, Huan; Simon, Françoise; Tauc, Patrick; Brochon, Jean-Claude; Zhao, Yunlong; Deprez, Eric

    2012-01-01

    Fatty acid-binding proteins (FABPs) are small cytosolic proteins, largely distributed in invertebrates and vertebrates, which accomplish uptake and intracellular transport of hydrophobic ligands such as fatty acids. Although long chain fatty acids play multiple crucial roles in cellular functions (structural, energy metabolism, regulation of gene expression), the precise functions of FABPs, especially those of invertebrate species, remain elusive. Here, we have identified and characterized a novel FABP family member, Cq-FABP, from the hepatopancreas of red claw crayfish Cherax quadricarinatus. We report the characterization of fatty acid-binding affinity of Cq-FABP by four different competitive fluorescence-based assays. In the two first approaches, the fluorescent probe 8-Anilino-1-naphthalenesulfonate (ANS), a binder of internal cavities of protein, was used either by directly monitoring its fluorescence emission or by monitoring the fluorescence resonance energy transfer occurring between the single tryptophan residue of Cq-FABP and ANS. The third and the fourth approaches were based on the measurement of the fluorescence emission intensity of the naturally fluorescent cis-parinaric acid probe or the steady-state fluorescence anisotropy measurements of a fluorescently labeled fatty acid (BODIPY-C16), respectively. The four methodologies displayed consistent equilibrium constants for a given fatty acid but were not equivalent in terms of analysis. Indeed, the two first methods were complicated by the existence of non specific binding modes of ANS while BODIPY-C16 and cis-parinaric acid specifically targeted the fatty acid binding site. We found a relationship between the affinity and the length of the carbon chain, with the highest affinity obtained for the shortest fatty acid, suggesting that steric effects primarily influence the interaction of fatty acids in the binding cavity of Cq-FABP. Moreover, our results show that the binding affinities of several fatty

  5. Allosteric Sensing of Fatty Acid Binding by NMR: Application to Human Serum Albumin.

    PubMed

    Jafari, Naeimeh; Ahmed, Rashik; Gloyd, Melanie; Bloomfield, Jonathon; Britz-McKibbin, Philip; Melacini, Giuseppe

    2016-08-25

    Human serum albumin (HSA) serves not only as a physiological oncotic pressure regulator and a ligand carrier but also as a biomarker for pathologies ranging from ischemia to diabetes. Moreover, HSA is a biopharmaceutical with a growing repertoire of putative clinical applications from hypovolemia to Alzheimer's disease. A key determinant of the physiological, diagnostic, and therapeutic functions of HSA is the amount of long chain fatty acids (LCFAs) bound to HSA. Here, we propose to utilize (13)C-oleic acid for the NMR-based assessment of albumin-bound LCFA concentration (CONFA). (13)C-Oleic acid primes HSA for a LCFA-dependent allosteric transition that modulates the frequency separation between the two main (13)C NMR peaks of HSA-bound oleic acid (ΔνAB). On the basis of ΔνAB, the overall [(12)C-LCFA]Tot/[HSA]Tot ratio is reproducibly estimated in a manner that is only minimally sensitive to glycation, albumin concentration, or redox potential, unlike other methods to quantify HSA-bound LCFAs such as the albumin-cobalt binding assay. PMID:27429126

  6. Hepatitis B Virus X Protein Induces Hepatic Steatosis by Enhancing the Expression of Liver Fatty Acid Binding Protein

    PubMed Central

    Wu, Yun-li; Peng, Xian-e; Zhu, Yi-bing; Yan, Xiao-li; Chen, Wan-nan

    2015-01-01

    ABSTRACT Hepatitis B virus (HBV) has been implicated as a potential trigger of hepatic steatosis although molecular mechanisms involved in the pathogenesis of HBV-associated hepatic steatosis still remain elusive. Our prior work has revealed that the expression level of liver fatty acid binding protein 1 (FABP1), a key regulator of hepatic lipid metabolism, was elevated in HBV-producing hepatoma cells. In this study, the effects of HBV X protein (HBx) mediated FABP1 regulation on hepatic steatosis and the underlying mechanism were determined. mRNA and protein levels of FABP1 were measured by quantitative RT-PCR (qPCR) and Western blotting. HBx-mediated FABP1 regulation was evaluated by luciferase assay, coimmunoprecipitation, and chromatin immunoprecipitation. Hepatic lipid accumulation was measured by using Oil-Red-O staining and the triglyceride level. It was found that expression of FABP1 was increased in HBV-producing hepatoma cells, the sera of HBV-infected patients, and the sera and liver tissues of HBV-transgenic mice. Ectopic overexpression of HBx resulted in upregulation of FABP1 in HBx-expressing hepatoma cells, whereas HBx abolishment reduced FABP1 expression. Mechanistically, HBx activated the FABP1 promoter in an HNF3β-, C/EBPα-, and PPARα-dependent manner, in which HBx increased the gene expression of HNF3β and physically interacted with C/EBPα and PPARα. On the other hand, knockdown of FABP1 remarkably blocked lipid accumulation both in long-chain free fatty acids treated HBx-expressing HepG2 cells and in a high-fat diet-fed HBx-transgenic mice. Therefore, FABP1 is a key driver gene in HBx-induced hepatic lipid accumulation via regulation of HNF3β, C/EBPα, and PPARα. FABP1 may represent a novel target for treatment of HBV-associated hepatic steatosis. IMPORTANCE Accumulating evidence from epidemiological and experimental studies has indicated that chronic HBV infection is associated with hepatic steatosis. However, the molecular mechanism

  7. Fatty Acid-Binding Protein 4 (FABP4): Pathophysiological Insights and Potent Clinical Biomarker of Metabolic and Cardiovascular Diseases

    PubMed Central

    Furuhashi, Masato; Saitoh, Shigeyuki; Shimamoto, Kazuaki; Miura, Tetsuji

    2014-01-01

    Over the past decade, evidences of an integration of metabolic and inflammatory pathways, referred to as metaflammation in several aspects of metabolic syndrome, have been accumulating. Fatty acid-binding protein 4 (FABP4), also known as adipocyte FABP (A-FABP) or aP2, is mainly expressed in adipocytes and macrophages and plays an important role in the development of insulin resistance and atherosclerosis in relation to metaflammation. Despite lack of a typical secretory signal peptide, FABP4 has been shown to be released from adipocytes in a non-classical pathway associated with lipolysis, possibly acting as an adipokine. Elevation of circulating FABP4 levels is associated with obesity, insulin resistance, diabetes mellitus, hypertension, cardiac dysfunction, atherosclerosis, and cardiovascular events. Furthermore, ectopic expression and function of FABP4 in several types of cells and tissues have been recently demonstrated. Here, we discuss both the significant role of FABP4 in pathophysiological insights and its usefulness as a biomarker of metabolic and cardiovascular diseases. PMID:25674026

  8. Immunodiagnostic monoclonal antibody-based sandwich ELISA of fasciolosis by detection of Fasciola gigantica circulating fatty acid binding protein.

    PubMed

    Anuracpreeda, Panat; Chawengkirttikul, Runglawan; Sobhon, Prasert

    2016-09-01

    Up to now, parasitological diagnosis of fasciolosis is often unreliable and possesses low sensitivity. Hence, the detection of circulating parasite antigens is thought to be a better alternative for diagnosis of fasciolosis, as it reflects the real parasite burden. In the present study, a monoclonal antibody (MoAb) against recombinant Fasciola gigantica fatty acid binding protein (rFgFABP) has been produced. As well, a reliable sandwich enzyme-linked immunosorbent assay (sandwich ELISA) has been developed for the detection of circulating FABP in the sera of mice experimentally and cattle naturally infected with F. gigantica. MoAb 3A3 and biotinylated rabbit anti-recombinant FABP antibody were selected due to their high reactivities and specificities. The lower detection limit of sandwich ELISA was 5 pg mL-1, and no cross-reaction with other parasite antigens was observed. This assay could detect F. gigantica infection from day 1 post infection. In experimental mice, the sensitivity, specificity and accuracy of this assay were 93·3, 100 and 98·2%, while in natural cattle they were 96·7, 100 and 99·1%. Hence, this sandwich ELISA method showed high efficiencies and precisions for diagnosis of fasciolosis by F. gigantica. PMID:27312522

  9. Adipocyte fatty acid-binding protein levels are associated with left ventricular diastolic dysfunction in morbidly obese subjects

    PubMed Central

    Baessler, A; Lamounier-Zepter, V; Fenk, S; Strack, C; Lahmann, C; Loew, T; Schmitz, G; Blüher, M; Bornstein, S R; Fischer, M

    2014-01-01

    Objectives: This study aimed to examine the association of adipocyte fatty acid-binding protein (FABP4) levels with left ventricular diastolic dysfunction (LVDD) in obese subjects with varying degrees of the metabolic syndrome (MetS). Methods: Fifty morbidly obese subjects with LVDD were selected at random and matched by age (±5 years) and sex with 50 morbidly obese with normal left ventricular (LV) function. In addition, 24 healthy lean subjects were included as controls. Results: Median FABP4 levels (interquartile range) in obese subjects with LVDD were significantly higher (42 ng ml−1 (32–53)) than in obese with normal LV function (24 ng ml−1 (36–43), P=0.036), and in normal weight controls (13 ng ml−1 (10–20), P<0.0001). Increasing FABP4 tertiles were significantly associated with parameters of LVDD, the number of LVDD components, physical performance and epicardial fat thickness. In multivariate regression analysis adjusting for age, sex and adiposity, FABP4 levels remained significantly associated with parameters of diastolic function. The association of FABP4 levels with LVDD was mainly observed in subjects with metabolic complications, but not in metabolically healthy obese. Conclusions: FABP4 levels are significantly associated with LVDD in obese subjects, when the MetS is present. Thus, FABP4 may be a link between obesity and cardiometabolic disorders. PMID:24513579

  10. Human liver-type fatty acid-binding protein protects against tubulointerstitial injury in aldosterone-induced renal injury.

    PubMed

    Ichikawa, Daisuke; Kamijo-Ikemori, Atsuko; Sugaya, Takeshi; Shibagaki, Yugo; Yasuda, Takashi; Hoshino, Seiko; Katayama, Kimie; Igarashi-Migitaka, Junko; Hirata, Kazuaki; Kimura, Kenjiro

    2015-01-15

    To demonstrate the renoprotective function of human liver-type fatty acid-binding protein (hL-FABP) expressed in proximal tubules in aldosterone (Aldo)-induced renal injury, hL-FABP chromosomal transgenic (Tg) and wild-type (WT) mice received systemic Aldo infusions (Tg-Aldo and WT-Aldo, respectively) were given 1% NaCl water for 28 days. In this model, elevation of systolic blood pressure, monocyte chemoattractant protein-1 expression, macrophage infiltration in the interstitium, tubulointerstitial damage, and depositions of type I and III collagens were observed. Elevation of systolic blood pressure did not differ in WT-Aldo vs. Tg-Aldo animals, however, renal injury was suppressed in Tg-Aldo compared with WT-Aldo mice. Dihydroethidium fluorescence was used to evaluate reactive oxidative stress, which was suppressed in Tg-Aldo compared with WT-Aldo mice. Gene expression of angiotensinogen in the kidney was upregulated, and excretion of urinary angiotensinogen was increased in WT-Aldo mice. This exacerbation was suppressed in Tg-Aldo mice. Expression of hL-FABP was upregulated in proximal tubules of Tg-Aldo mice. Urinary excretion of hL-FABP was significantly greater in Tg-Aldo than in Tg-control mice. In conclusion, hL-FABP ameliorated the tubulointerstitial damage in Aldo-induced renal injury via reducing oxidative stress and suppressing activation of the intrarenal renin-angiotensin system. PMID:25339700

  11. Endothelial cell-fatty acid binding protein 4 promotes angiogenesis: role of stem cell factor/c-kit pathway.

    PubMed

    Elmasri, Harun; Ghelfi, Elisa; Yu, Chen-wei; Traphagen, Samantha; Cernadas, Manuela; Cao, Haiming; Shi, Guo-Ping; Plutzky, Jorge; Sahin, Mustafa; Hotamisligil, Gokhan; Cataltepe, Sule

    2012-09-01

    Fatty acid binding protein 4 (FABP4) plays an important role in regulation of glucose and lipid homeostasis as well as inflammation through its actions in adipocytes and macrophages. FABP4 is also expressed in a subset of endothelial cells, but its role in this cell type is not known. We found that FABP4-deficient human umbilical vein endothelial cells (HUVECs) demonstrate a markedly increased susceptibility to apoptosis as well as decreased migration and capillary network formation. Aortic rings from FABP4(-/-) mice demonstrated decreased angiogenic sprouting, which was recovered by reconstitution of FABP4. FABP4 was strongly regulated by mTORC1 and inhibited by Rapamycin. FABP4 modulated activation of several important signaling pathways in HUVECs, including downregulation of P38, eNOS, and stem cell factor (SCF)/c-kit signaling. Of these, the SCF/c-kit pathway was found to have a major role in attenuated angiogenic activity of FABP4-deficient ECs as provision of exogenous SCF resulted in a significant recovery in cell proliferation, survival, morphogenesis, and aortic ring sprouting. These data unravel a novel pro-angiogenic role for endothelial cell-FABP4 and suggest that it could be exploited as a potential target for diseases associated with pathological angiogenesis. PMID:22562362

  12. Fatty acid binding protein 4 is a target of VEGF and a regulator of cell proliferation in endothelial cells.

    PubMed

    Elmasri, Harun; Karaaslan, Cagatay; Teper, Yaroslav; Ghelfi, Elisa; Weng, Meiqian; Ince, Tan A; Kozakewich, Harry; Bischoff, Joyce; Cataltepe, Sule

    2009-11-01

    Fatty acid binding protein 4 (FABP4) plays an important role in maintaining glucose and lipid homeostasis. FABP4 has been primarily regarded as an adipocyte- and macrophage-specific protein, but recent studies suggest that it may be more widely expressed. We found strong FABP4 expression in the endothelial cells (ECs) of capillaries and small veins in several mouse and human tissues, including the heart and kidney. FABP4 was also detected in the ECs of mature human placental vessels and infantile hemangiomas, the most common tumor of infancy and ECs. In most of these cases, FABP4 was detected in both the nucleus and cytoplasm. FABP4 mRNA and protein levels were significantly induced in cultured ECs by VEGF-A and bFGF treatment. The effect of VEGF-A on FABP4 expression was inhibited by chemical inhibition or short-hairpin (sh) RNA-mediated knockdown of VEGF-receptor-2 (R2), whereas the VEGFR1 agonists, placental growth factors 1 and 2, had no effect on FABP4 expression. Knockdown of FABP4 in ECs significantly reduced proliferation both under baseline conditions and in response to VEGF and bFGF. Thus, FABP4 emerged as a novel target of the VEGF/VEGFR2 pathway and a positive regulator of cell proliferation in ECs. PMID:19625659

  13. Endothelial cell-fatty acid binding protein 4 promotes angiogenesis: role of stem cell factor/c-kit pathway

    PubMed Central

    Elmasri, Harun; Ghelfi, Elisa; Yu, Chen-wei; Traphagen, Samantha; Cernadas, Manuela; Cao, Haiming; Shi, Guo-Ping; Plutzky, Jorge; Sahin, Mustafa; Hotamisligil, Gokhan; Cataltepe, Sule

    2013-01-01

    Fatty acid binding protein 4 (FABP4) plays an important role in regulation of glucose and lipid homeostasis as well as inflammation through its actions in adipocytes and macrophages. FABP4 is also expressed in a subset of endothelial cells, but its role in this cell type is not known. We found that FABP4-deficient human umbilical vein endothelial cells (HUVECs) demonstrate a markedly increased susceptibility to apoptosis as well as decreased migration and capillary network formation. Aortic rings from FABP4−/− mice demonstrated decreased angiogenic sprouting, which was recovered by reconstitution of FABP4. FABP4 was strongly regulated by mTORC1 and inhibited by Rapamycin. FABP4 modulated activation of several important signaling pathways in HUVECs, including downregulation of P38, eNOS, and stem cell factor (SCF)/c-kit signaling. Of these, the SCF/c-kit pathway was found to have a major role in attenuated angiogenic activity of FABP4-deficient ECs as provision of exogenous SCF resulted in a significant recovery in cell proliferation, survival, morphogenesis, and aortic ring sprouting. These data unravel a novel pro-angiogenic role for endothelial cell-FABP4 and suggest that it could be exploited as a potential target for diseases associated with pathological angiogenesis. PMID:22562362

  14. Urinary Intestinal Fatty Acid-Binding Protein Can Distinguish Necrotizing Enterocolitis from Sepsis in Early Stage of the Disease

    PubMed Central

    Snajdauf, Jiri; Rygl, Michal

    2016-01-01

    Necrotizing enterocolitis (NEC) is severe disease of gastrointestinal tract, yet its early symptoms are nonspecific, easily interchangeable with sepsis. Therefore, reliable biomarkers for early diagnostics are needed in clinical practice. Here, we analyzed if markers of gut mucosa damage, caspase cleaved cytokeratin 18 (ccCK18) and intestinal fatty acid-binding protein (I-FABP), could be used for differential diagnostics of NEC at early stage of disease. We collected paired serum (at enrollment and week later) and urine (collected for two days in 6 h intervals) samples from 42 patients with suspected NEC. These patients were later divided into NEC (n = 24), including 13 after gastrointestinal surgery, and sepsis (n = 18) groups using standard criteria. Healthy infants (n = 12), without any previous gut surgery, served as controls. Both biomarkers were measured by a commercial ELISA assay. There were no statistically significant differences in serum ccCK18 between NEC and sepsis but NEC patients had significantly higher levels of serum and urinary I-FABP than either sepsis patients or healthy infants. Urinary I-FABP has high sensitivity (81%) and specificity (100%) and can even distinguish NEC from sepsis in patients after surgery. Urinary I-FABP can be used to distinguish NEC from neonatal sepsis, including postoperative one, better than abdominal X-ray. PMID:27110575

  15. Fasciola gigantica Fatty Acid Binding Protein (FABP) as a Prophylactic Agent against Schistosoma mansoni Infection in CD1 Mice

    PubMed Central

    Diab, M.; El-Amir, A. M.; Hendawy, M.; Kadry, S.

    2012-01-01

    Although schistosomicidal drugs and other control measures exist, the advent of an efficacious vaccine remains the most potentially powerful means for controlling this disease. In this study, native fatty acid binding protein (FABP) from Fasciola gigantica was purified from the adult worm's crude extract by saturation with ammonium sulphate followed by separation on DEAE-Sephadex A-50 anion exchange chromatography and gel filtration using Sephacryl HR-100, respectively. CD1 mice were immunized with the purified, native F. gigantica FABP in Freund's adjuvant and challenged subcutaneously with 120 Schistosoma mansoni cercariae. Immunization of CD1 mice with F. gigantica FABP has induced heterologous protection against S. mansoni, evidenced by the significant reduction in mean worm burden (72.3%), liver and intestinal egg counts (81.3% and 80.8%, respectively), and hepatic granuloma counts (42%). Also, it elicited mixed IgG1/IgG2b immune responses with predominant IgG1 isotype, suggesting that native F. gigantica FABP is mediated by a mixed Th1/Th2 response. However, it failed to induce any significant differences in the oogram pattern or in the mean granuloma diameter. This indicated that native F. gigantica FABP could be a promising vaccine candidate against S. mansoni infection. PMID:22451732

  16. Fatty acid-binding protein (fabp) genes of spotted green pufferfish (Tetraodon nigroviridis): comparative genomics and spatial transcriptional regulation.

    PubMed

    Thirumaran, Aruloli; Wright, Jonathan M

    2014-05-01

    The fatty acid-binding protein (fabp) genes belong to the multigene family of intracellular lipid-binding proteins. To date, 12 different FABPs have been identified in vertebrate genomes. Owing to the teleost-specific genome duplication event, many fishes have duplicated copies of the fabp genes. Here, we identified and characterized the fabp genes of spotted green pufferfish (Tetraodon nigroviridis). Seven fabp genes were identified, out of which, two were retained in the pufferfish genome as duplicated copies. Each putative pufferfish Fabp protein shares greatest sequence identity and similarity with their teleost and tetrapod orthologs, and clustered together as a distinct clade in phylogenetic analysis. Conserved gene synteny was evident between the pufferfish fabp genes and the orthologs of human, zebrafish, three-spined stickleback, and medaka FABP/fabp genes, providing evidence that the duplicated copies of pufferfish fabp genes most likely arose as a result of the teleost-specific genome duplication event. The differential tissue-specific distribution of pufferfish fabp transcripts suggests divergent spatial regulation of duplicated pairs of fabp genes. PMID:25153522

  17. An Evidence-Based Approach to the Assessment of Heart-Type Fatty Acid Binding Protein in Acute Coronary Syndrome

    PubMed Central

    Viswanathan, Karthik; Hall, Alistair S; Barth, Julian H

    2012-01-01

    Cardiac troponins have been the biomarkers of choice for the diagnosis of acute coronary syndrome (ACS) for over a decade. There has, however, been considerable interest over the last two decades for newer biomarkers that would bring added value to the measurement of troponin such as the provision of prognosis and assistance in the choice of therapeutic interventions. In this manuscript, we review the development of heart-type fatty acid binding protein (H-FABP) in patients with ACS using the evidence-based laboratory medicine format. Phase I studies have established that H-FABP reference intervals and pre-analytical factors influencing H-FABP. Phase II studies have confirmed a) that H-FABP is elevated in patients with established myocardial infarction; b) that its serum concentration is related to the extent of infarction using survival as a surrogate; and c) that its use in chest pain patients can identify ACS patients and also provide prognostic information on survival. Furthermore, it is an independent prognostic marker for patients with suspected ACS who are troponin negative. Phase III studies involving randomised control trials for diagnosis and prognosis have not yet been performed and Phase IV studies await uptake of H-FABP in a routine service. PMID:22363093

  18. Characterization of the comparative drug binding to intra- (liver fatty acid binding protein) and extra- (human serum albumin) cellular proteins.

    PubMed

    Rowland, Andrew; Hallifax, David; Nussio, Matthew R; Shapter, Joseph G; Mackenzie, Peter I; Brian Houston, J; Knights, Kathleen M; Miners, John O

    2015-01-01

    1. This study compared the extent, affinity, and kinetics of drug binding to human serum albumin (HSA) and liver fatty acid binding protein (LFABP) using ultrafiltration and surface plasmon resonance (SPR). 2. Binding of basic and neutral drugs to both HSA and LFABP was typically negligible. Binding of acidic drugs ranged from minor (fu > 0.8) to extensive (fu < 0.1). Of the compounds screened, the highest binding to both HSA and LFABP was observed for the acidic drugs torsemide and sulfinpyrazone, and for β-estradiol (a polar, neutral compound). 3. The extent of binding of acidic drugs to HSA was up to 40% greater than binding to LFABP. SPR experiments demonstrated comparable kinetics and affinity for the binding of representative acidic drugs (naproxen, sulfinpyrazone, and torsemide) to HSA and LFABP. 4. Simulations based on in vitro kinetic constants derived from SPR experiments and a rapid equilibrium model were undertaken to examine the impact of binding characteristics on compartmental drug distribution. Simulations provided mechanistic confirmation that equilibration of intracellular unbound drug with the extracellular unbound drug is attained rapidly in the absence of active transport mechanisms for drugs bound moderately or extensively to HSA and LFABP. PMID:25801059

  19. Fatty acid-binding proteins (FABPs) are intracellular carriers for Δ9-tetrahydrocannabinol (THC) and cannabidiol (CBD).

    PubMed

    Elmes, Matthew W; Kaczocha, Martin; Berger, William T; Leung, KwanNok; Ralph, Brian P; Wang, Liqun; Sweeney, Joseph M; Miyauchi, Jeremy T; Tsirka, Stella E; Ojima, Iwao; Deutsch, Dale G

    2015-04-01

    Δ(9)-Tetrahydrocannabinol (THC) and cannabidiol (CBD) occur naturally in marijuana (Cannabis) and may be formulated, individually or in combination in pharmaceuticals such as Marinol or Sativex. Although it is known that these hydrophobic compounds can be transported in blood by albumin or lipoproteins, the intracellular carrier has not been identified. Recent reports suggest that CBD and THC elevate the levels of the endocannabinoid anandamide (AEA) when administered to humans, suggesting that phytocannabinoids target cellular proteins involved in endocannabinoid clearance. Fatty acid-binding proteins (FABPs) are intracellular proteins that mediate AEA transport to its catabolic enzyme fatty acid amide hydrolase (FAAH). By computational analysis and ligand displacement assays, we show that at least three human FABPs bind THC and CBD and demonstrate that THC and CBD inhibit the cellular uptake and catabolism of AEA by targeting FABPs. Furthermore, we show that in contrast to rodent FAAH, CBD does not inhibit the enzymatic actions of human FAAH, and thus FAAH inhibition cannot account for the observed increase in circulating AEA in humans following CBD consumption. Using computational molecular docking and site-directed mutagenesis we identify key residues within the active site of FAAH that confer the species-specific sensitivity to inhibition by CBD. Competition for FABPs may in part or wholly explain the increased circulating levels of endocannabinoids reported after consumption of cannabinoids. These data shed light on the mechanism of action of CBD in modulating the endocannabinoid tone in vivo and may explain, in part, its reported efficacy toward epilepsy and other neurological disorders. PMID:25666611

  20. Fatty Acid-binding Proteins (FABPs) Are Intracellular Carriers for Δ9-Tetrahydrocannabinol (THC) and Cannabidiol (CBD)*

    PubMed Central

    Elmes, Matthew W.; Kaczocha, Martin; Berger, William T.; Leung, KwanNok; Ralph, Brian P.; Wang, Liqun; Sweeney, Joseph M.; Miyauchi, Jeremy T.; Tsirka, Stella E.; Ojima, Iwao; Deutsch, Dale G.

    2015-01-01

    Δ9-Tetrahydrocannabinol (THC) and cannabidiol (CBD) occur naturally in marijuana (Cannabis) and may be formulated, individually or in combination in pharmaceuticals such as Marinol or Sativex. Although it is known that these hydrophobic compounds can be transported in blood by albumin or lipoproteins, the intracellular carrier has not been identified. Recent reports suggest that CBD and THC elevate the levels of the endocannabinoid anandamide (AEA) when administered to humans, suggesting that phytocannabinoids target cellular proteins involved in endocannabinoid clearance. Fatty acid-binding proteins (FABPs) are intracellular proteins that mediate AEA transport to its catabolic enzyme fatty acid amide hydrolase (FAAH). By computational analysis and ligand displacement assays, we show that at least three human FABPs bind THC and CBD and demonstrate that THC and CBD inhibit the cellular uptake and catabolism of AEA by targeting FABPs. Furthermore, we show that in contrast to rodent FAAH, CBD does not inhibit the enzymatic actions of human FAAH, and thus FAAH inhibition cannot account for the observed increase in circulating AEA in humans following CBD consumption. Using computational molecular docking and site-directed mutagenesis we identify key residues within the active site of FAAH that confer the species-specific sensitivity to inhibition by CBD. Competition for FABPs may in part or wholly explain the increased circulating levels of endocannabinoids reported after consumption of cannabinoids. These data shed light on the mechanism of action of CBD in modulating the endocannabinoid tone in vivo and may explain, in part, its reported efficacy toward epilepsy and other neurological disorders. PMID:25666611

  1. The cancer-promoting gene fatty acid-binding protein 5 (FABP5) is epigenetically regulated during human prostate carcinogenesis.

    PubMed

    Kawaguchi, Koichiro; Kinameri, Ayumi; Suzuki, Shunsuke; Senga, Shogo; Ke, Youqiang; Fujii, Hiroshi

    2016-02-15

    FABPs (fatty-acid-binding proteins) are a family of low-molecular-mass intracellular lipid-binding proteins consisting of ten isoforms. FABPs are involved in binding and storing hydrophobic ligands such as long-chain fatty acids, as well as transporting these ligands to the appropriate compartments in the cell. FABP5 is overexpressed in multiple types of tumours. Furthermore, up-regulation of FABP5 is strongly associated with poor survival in triple-negative breast cancer. However, the mechanisms underlying the specific up-regulation of the FABP5 gene in these cancers remain poorly characterized. In the present study, we determined that FABP5 has a typical CpG island around its promoter region. The DNA methylation status of the CpG island in the FABP5 promoter of benign prostate cells (PNT2), prostate cancer cells (PC-3, DU-145, 22Rv1 and LNCaP) and human normal or tumour tissue was assessed by bisulfite sequencing analysis, and then confirmed by COBRA (combined bisulfite restriction analysis) and qAMP (quantitative analysis of DNA methylation using real-time PCR). These results demonstrated that overexpression of FABP5 in prostate cancer cells can be attributed to hypomethylation of the CpG island in its promoter region, along with up-regulation of the direct trans-acting factors Sp1 (specificity protein 1) and c-Myc. Together, these mechanisms result in the transcriptional activation of FABP5 expression during human prostate carcinogenesis. Importantly, silencing of Sp1, c-Myc or FABP5 expression led to a significant decrease in cell proliferation, indicating that up-regulation of FABP5 expression by Sp1 and c-Myc is critical for the proliferation of prostate cancer cells. PMID:26614767

  2. Hormonal regulation of liver fatty acid-binding protein in vivo and in vitro: effects of growth hormone and insulin.

    PubMed

    Carlsson, L; Nilsson, I; Oscarsson, J

    1998-06-01

    Liver fatty acid-binding protein (LFABP) is an abundant protein in hepatocytes that binds most of the long chain fatty acids present in the cytosol. It is suggested to be of importance for fatty acid uptake and utilization in the hepatocyte. In the present study, the effects of bovine GH (bGH) and other hormones on the expression of LFABP and its messenger RNA (mRNA) were studied in hypophysectomized rats and in vitro using primary cultures of rat hepatocytes. One injection of bGH increased LFABP mRNA levels about 5-fold after 6 h, but there was no effect of this treatment on LFABP levels. However, 7 days of bGH treatment increased both LFABP mRNA and LFABP protein levels 2- to 5-fold. Female rats had higher levels of LFABP than male rats. Hypophysectomy of female rats, but not that of male rats, decreased LFABP levels markedly. Treatment of hypophysectomized rats with bGH for 7 days as two daily injections or as a continuous infusion increased LFABP levels to a similar degree. This finding indicates that the sex difference in the expression of LFABP is not regulated by the sexually dimorphic secretory pattern of GH. Neither insulin nor insulin-like growth factor I treatment of hypophysectomized rats for 6-7 days had any effect on LFABP mRNA or LFABP levels. In vitro, bGH dose-dependently increased the expression of LFABP mRNA, but only in the presence of insulin. Insulin alone had a marked dose-dependent effect on LFABP mRNA levels and was of importance for maintaining the expression of LFABP mRNA during the culture. Incubation with bGH increased LFABP mRNA levels within 3 h. GH had no effect on LFABP mRNA levels in the presence of actinomycin D, indicating a transcriptional effect of GH. Incubation with glucagon in vitro decreased LFABP mRNA levels markedly, indicating that glucagon, in contrast to GH, has an effect opposite that of insulin on LFABP mRNA expression. It is concluded that GH is an important regulator of LFABP in vivo and in vitro. In contrast to

  3. Transfection of L6 myoblasts with adipocyte fatty acid-binding protein cDNA does not affect fatty acid uptake but disturbs lipid metabolism and fusion.

    PubMed Central

    Prinsen, C F; Veerkamp, J H

    1998-01-01

    We studied the involvement of fatty acid-binding protein (FABP) in growth, differentiation and fatty acid metabolism of muscle cells by lipofection of rat L6 myoblasts with rat heart (H) FABP cDNA or with rat adipocyte (A) FABP cDNA in a eukaryotic expression vector which contained a puromycin acetyltransferase cassette. Stable transfectants showed integration into the genome for all constructs and type-specific overexpression at the mRNA and protein level for the clones with H-FABP and A-FABP cDNA constructs. The rate of proliferation of myoblasts transfected with rat A-FABP cDNA was 2-fold higher compared with all other transfected cells. In addition, these myoblasts showed disturbed fusion and differentiation, as assessed by morphological examination and creatine kinase activity. Uptake rates of palmitate were equal for all clone types, in spite of different FABP content and composition. Palmitate oxidation over a 3 h period was similar in all clones from growth medium. After being cultured in differentiation medium, mock- and H-FABP-cDNA-transfected cells showed a lower fatty acid-oxidation rate, in contrast with A-FABP-cDNA-transfected clones. The ratio of [14C]palmitic acid incorporation into phosphatidylcholine and phosphatidylethanolamine of A-FABP-cDNA-transfected clones changed in the opposite direction in differentiation medium from that of mock- and H-FABP-cDNA-transfected clones. In conclusion, transfection of L6 myoblasts with A-FABP cDNA does not affect H-FABP content and fatty acid uptake, but changes fatty acid metabolism. The latter changes may be related to the observed fusion defect. PMID:9425108

  4. Fatty Acid Binding Protein 5 Modulates Docosahexaenoic Acid-Induced Recovery in Rats Undergoing Spinal Cord Injury.

    PubMed

    Figueroa, Johnny D; Serrano-Illan, Miguel; Licero, Jenniffer; Cordero, Kathia; Miranda, Jorge D; De Leon, Marino

    2016-08-01

    Omega-3 polyunsaturated fatty acids (n-3 PUFAs) promote functional recovery in rats undergoing spinal cord injury (SCI). However, the precise molecular mechanism coupling n-3 PUFAs to neurorestorative responses is not well understood. The aim of the present study was to determine the spatiotemporal expression of fatty acid binding protein 5 (FABP5) after contusive SCI and to investigate whether this protein plays a role in n-3 PUFA-mediated functional recovery post-SCI. We found that SCI resulted in a robust spinal cord up-regulation in FABP5 mRNA levels (556 ± 187%) and protein expression (518 ± 195%), when compared to sham-operated rats, at 7 days post-injury (dpi). This upregulation coincided with significant alterations in the metabolism of fatty acids in the injured spinal cord, as revealed by metabolomics-based lipid analyses. In particular, we found increased levels of the n-3 series PUFAs, particularly docosahexaenoic acid (DHA; 22:6 n-3) and eicosapentaenoic acid (EPA; 20:5 n-3) at 7 dpi. Animals consuming a diet rich in DHA and EPA exhibited a significant upregulation in FABP5 mRNA levels at 7 dpi. Immunofluorescence showed low basal FABP5 immunoreactivity in spinal cord ventral gray matter NeuN(+) neurons of sham-operated rats. SCI resulted in a robust induction of FABP5 in glial (GFAP(+), APC(+), and NG2(+)) and precursor cells (DCX(+), nestin(+)). We found that continuous intrathecal administration of FABP5 silencing with small interfering RNA (2 μg) impaired spontaneous open-field locomotion post-SCI. Further, FABP5 siRNA administration hindered the beneficial effects of DHA to ameliorate functional recovery at 7 dpi. Altogether, our findings suggest that FABP5 may be an important player in the promotion of cellular uptake, transport, and/or metabolism of DHA post-SCI. Given the beneficial roles of n-3 PUFAs in ameliorating functional recovery, we propose that FABP5 is an important contributor to basic repair mechanisms in the

  5. Brief Report: Differential Associations of Interleukin 6 and Intestinal Fatty Acid-Binding Protein With Progressive Untreated HIV-1 Infection in Rakai, Uganda.

    PubMed

    Olwenyi, Omalla A; Naluyima, Prossy; Cham, Fatim; Quinn, Thomas C; Serwadda, David; Sewankambo, Nelson K; Gray, Ronald H; Sandberg, Johan K; Michael, Nelson L; Wabwire-Mangen, Fred; Robb, Merlin L; Eller, Michael A

    2016-05-01

    The significance of HIV-associated immune activation and microbial translocation in Sub-Saharan African population remains poorly defined. We assessed biomarkers of inflammation, microbial translocation, and cellular activation and found most factors elevated in Ugandan HIV-1 seroconverters compared with community-matched controls. In contrast to previous findings in Western cohorts, C-reactive protein, neopterin, and intestinal fatty acid binding protein were not elevated. Higher T-cell activation and IL-6 were associated with faster disease progression. Surprisingly, intestinal fatty acid binding protein, indicative of enterocyte turnover, was higher in slow than in fast progressors. These data suggest differential relationships among biomarkers of intestinal barrier integrity and innate immune activation between developed countries and Sub-Saharan Africa. PMID:26630672

  6. Fatty Acid-binding Protein 4, a Point of Convergence for Angiogenic and Metabolic Signaling Pathways in Endothelial Cells*

    PubMed Central

    Harjes, Ulrike; Bridges, Esther; McIntyre, Alan; Fielding, Barbara A.; Harris, Adrian L.

    2014-01-01

    Fatty acid-binding protein 4 (FABP4) is an adipogenic protein and is implicated in atherosclerosis, insulin resistance, and cancer. In endothelial cells, FABP4 is induced by VEGFA, and inhibition of FABP4 blocks most of the VEGFA effects. We investigated the DLL4-NOTCH-dependent regulation of FABP4 in human umbilical vein endothelial cells by gene/protein expression and interaction analyses following inhibitor treatment and RNA interference. We found that FABP4 is directly induced by NOTCH. Stimulation of NOTCH signaling with human recombinant DLL4 led to FABP4 induction, independently of VEGFA. FABP4 induction by VEGFA was reduced by blockade of DLL4 binding to NOTCH or inhibition of NOTCH signal transduction. Chromatin immunoprecipitation of the NOTCH intracellular domain showed increased binding to two specific regions in the FABP4 promoter. The induction of FABP4 gene expression was dependent on the transcription factor FOXO1, which was essential for basal expression of FABP4, and FABP4 up-regulation following stimulation of the VEGFA and/or the NOTCH pathway. Thus, we show that the DLL4-NOTCH pathway mediates endothelial FABP4 expression. This indicates that induction of the angiogenesis-restricting DLL4-NOTCH can have pro-angiogenic effects via this pathway. It also provides a link between DLL4-NOTCH and FOXO1-mediated regulation of endothelial gene transcription, and it shows that DLL4-NOTCH is a nodal point in the integration of pro-angiogenic and metabolic signaling in endothelial cells. This may be crucial for angiogenesis in the tumor environment. PMID:24939870

  7. Serum Fatty Acid-Binding Protein 4 Is a Predictor of Cardiovascular Events in End-Stage Renal Disease

    PubMed Central

    Furuhashi, Masato; Ishimura, Shutaro; Ota, Hideki; Hayashi, Manabu; Nishitani, Takahiro; Tanaka, Marenao; Yoshida, Hideaki; Shimamoto, Kazuaki; Hotamisligil, Gökhan S.; Miura, Tetsuji

    2011-01-01

    Background Fatty acid-binding protein 4 (FABP4/A-FABP/aP2), a lipid chaperone, is expressed in both adipocytes and macrophages. Recent studies have shown that FABP4 is secreted from adipocytes and that FABP4 level is associated with obesity, insulin resistance, and atherosclerosis. However, little is known about the impact of FABP4 concentrations on prognosis. We tested the hypothesis that FABP4 level predicts prognosis of patients with end-stage renal disease (ESRD), a group at high risk for atherosclerosis-associated morbidity and mortality. Methods and Results Biochemical markers including FABP4 were determined in 61 ESRD patients on chronic hemodialysis (HD). Serum FABP4 level in females (404.2±30.5 ng/ml) was significantly higher than that in males (315.8±30.0 ng/ml), and the levels in ESRD patients were about 20-times higher than those in age-, gender- and body mass index (BMI)-matched control subjects with normal renal function. FABP4 level was decreased by 57.2% after HD and was positively correlated with blood pressure, BMI, and levels of lipids and insulin. Multiple regression analysis indicated that HD duration, BMI, and triglycerides level were independent determinants for FABP4 level. ESRD patients with high FABP4 levels had higher cardiovascular mortality during the 7-year follow-up period. Cox proportional hazard regression analysis showed that logarithmically transformed FABP4 level was an independent predictor of cardiovascular death adjusted for age, gender, HD duration, BMI, and triglycerides level (hazard ratio, 7.75; 95% CI, 1.05–25.31). Conclusion These findings suggest that FABP4 level, being related to adiposity and metabolic disorders, is a novel predictor of cardiovascular mortality in ESRD. PMID:22102888

  8. Fatty Acid Binding Protein 4 Regulates VEGF-Induced Airway Angiogenesis and Inflammation in a Transgenic Mouse Model

    PubMed Central

    Ghelfi, Elisa; Yu, Chen-Wei; Elmasri, Harun; Terwelp, Matthew; Lee, Chun G.; Bhandari, Vineet; Comhair, Suzy A.; Erzurum, Serpil C.; Hotamisligil, Gökhan S.; Elias, Jack A.; Cataltepe, Sule

    2014-01-01

    Neovascularization of the airways occurs in several inflammatory lung diseases, including asthma. Vascular endothelial growth factor (VEGF) plays an important role in vascular remodeling in the asthmatic airways. Fatty acid binding protein 4 (FABP4 or aP2) is an intracellular lipid chaperone that is induced by VEGF in endothelial cells. FABP4 exhibits a proangiogenic function in vitro, but whether it plays a role in modulation of angiogenesis in vivo is not known. We hypothesized that FABP4 promotes VEGF-induced airway angiogenesis and investigated this hypothesis with the use of a transgenic mouse model with inducible overexpression of VEGF165 under a CC10 promoter [VEGF-TG (transgenic) mice]. We found a significant increase in FABP4 mRNA levels and density of FABP4-expressing vascular endothelial cells in mouse airways with VEGF overexpression. FABP4−/− mouse airways showed a significant decrease in neovessel formation and endothelial cell proliferation in response to VEGF overexpression. These alterations in airway vasculature were accompanied by attenuated expression of proinflammatory mediators. Furthermore, VEGF-TG/FABP4−/− mice showed markedly decreased expression of endothelial nitric oxide synthase, a well-known mediator of VEGF-induced responses, compared with VEGF-TG mice. Finally, the density of FABP4-immunoreactive vessels in endobronchial biopsy specimens was significantly higher in patients with asthma than in control subjects. Taken together, these data unravel FABP4 as a potential target of pathologic airway remodeling in asthma. PMID:23391391

  9. Increased expression of fatty acid binding protein 4 and leptin in resident macrophages characterises atherosclerotic plaque rupture

    PubMed Central

    Lee, K.; Santibanez-Koref, M.; Polvikoski, T.; Birchall, D.; Mendelow, A.D.; Keavney, B.

    2013-01-01

    Objective Resident macrophages play an important role in atheromatous plaque rupture. The macrophage gene expression signature associated with plaque rupture is incompletely defined due to the complex cellular heterogeneity in the plaque. We aimed to characterise differential gene expression in resident plaque macrophages from ruptured and stable human atheromatous lesions. Methods and results We performed genome-wide expression analyses of isolated macrophage-rich regions of stable and ruptured human atherosclerotic plaques. Plaques present in carotid endarterectomy specimens were designated as stable or ruptured using clinical, radiological and histopathological criteria. Macrophage-rich regions were excised from 5 ruptured and 6 stable plaques by laser micro-dissection. Transcriptional profiling was performed using Affymetrix microarrays. The profiles were characteristic of activated macrophages. At a false discovery rate of 10%, 914 genes were differentially expressed between stable and ruptured plaques. The findings were confirmed in fourteen further stable and ruptured samples for a subset of eleven genes with the highest expression differences (p < 0.05). Pathway analysis revealed that components of the PPAR/Adipocytokine signaling pathway were the most significantly upregulated in ruptured compared to stable plaques (p = 5.4 × 10−7). Two key components of the pathway, fatty-acid binding-protein 4 (FABP4) and leptin, showed nine-fold (p = 0.0086) and five-fold (p = 0.0012) greater expression respectively in macrophages from ruptured plaques. Conclusions We found differences in gene expression signatures between macrophages isolated from stable and ruptured human atheromatous plaques. Our findings indicate the involvement of FABP4 and leptin in the progression of atherosclerosis and plaque rupture, and suggest that down-regulation of PPAR/adipocytokine signaling within plaques may have therapeutic potential. PMID:23122912

  10. Elevation of urinary liver-type fatty acid binding protein after cardiac catheterization related to cardiovascular events

    PubMed Central

    Kamijo-Ikemori, Atsuko; Hashimoto, Nobuyuki; Sugaya, Takeshi; Matsui, Katsuomi; Hisamichi, Mikako; Shibagaki, Yugo; Miyake, Fumihiko; Kimura, Kenjiro

    2015-01-01

    Purpose Contrast medium (CM) induces tubular hypoxia via endothelial damage due to direct cytotoxicity or viscosity. Urinary liver-type fatty acid binding protein (L-FABP) increases along with tubular hypoxia and may be a detector of systemic circulation injury. The aim of this study was to evaluate the clinical usefulness of detecting increases in urinary L-FABP levels due to administration of CM, as a prognostic biomarker for cardiovascular disease in patients without occurrence of CM-induced nephropathy undergoing cardiac catheterization procedure (CCP). Methods Retrospective longitudinal analyses of the relationship between urinary L-FABP levels and occurrence of cardiovascular events were performed (n=29). Urinary L-FABP was measured by ELISA before CCP, and at 6, 12, 24, and 48 hours after CCP. Results Urinary L-FABP levels were significantly higher at 12 hours (P<0.05) and 24 hours (P<0.005) after CCP compared with before CCP, only in the patients with occurrence of cardiovascular events (n=17), but not in those without cardiovascular events (n=12). The parameter with the largest area under the curve (0.816) for predicting the occurrence of cardiovascular events was the change in urinary L-FABP at 24 hours after CCP. The difference in urinary L-FABP levels (ΔL-FABP ≥11.0 μg/g creatinine) between before CCP and at 24 hours after CCP was a risk factor for the occurrence of cardiovascular events (hazard ratio, 4.93; 95% confidence interval, 1.27–19.13; P=0.021). Conclusion Measurement of urinary L-FABP before CCP and at 24 hours after CCP in patients with mild to moderate renal dysfunction may be an important indicator for risk stratification of onset of cardiovascular events. PMID:26316797

  11. Liver fatty acid binding protein gene ablation potentiates hepatic cholesterol accumulation in cholesterol-fed female mice.

    PubMed

    Martin, Gregory G; Atshaves, Barbara P; McIntosh, Avery L; Mackie, John T; Kier, Ann B; Schroeder, Friedhelm

    2006-01-01

    Although liver fatty acid binding protein (L-FABP) is postulated to influence cholesterol homeostasis, the physiological significance of this hypothesis remains to be resolved. This issue was addressed by examining the response of young (7 wk) female mice to L-FABP gene ablation and a cholesterol-rich diet. In control-fed mice, L-FABP gene ablation alone induced hepatic cholesterol accumulation (2.6-fold), increased bile acid levels, and increased body weight gain (primarily as fat tissue mass). In cholesterol-fed mice, L-FABP gene ablation further enhanced the hepatic accumulation of cholesterol (especially cholesterol ester, 12-fold) and potentiated the effects of dietary cholesterol on increased body weight gain, again mainly as fat tissue mass. However, in contrast to the effects of L-FABP gene ablation in control-fed mice, biliary levels of bile acids (as well as cholesterol and phospholipids) were reduced. These phenotypic alterations were not associated with differences in food intake. In conclusion, it was shown for the first time that L-FABP altered cholesterol metabolism and the response of female mice to dietary cholesterol. While the biliary and lipid phenotype of female wild-type L-FABP+/+ mice was sensitive to dietary cholesterol, L-FABP gene ablation dramatically enhanced many of the effects of dietary cholesterol to greatly induce hepatic cholesterol (primarily cholesterol ester) and triacylglycerol accumulation as well as to potentiate body weight gain (primarily as fat tissue mass). Taken together, these data support the hypothesis that L-FABP is involved in the physiological regulation of cholesterol metabolism, body weight gain, and obesity. PMID:16123197

  12. Metformin reduces lipid accumulation in macrophages by inhibiting FOXO1-mediated transcription of fatty acid-binding protein 4

    SciTech Connect

    Song, Jun; Ren, Pingping; Zhang, Lin; Wang, Xing Li; Chen, Li; Shen, Ying H.

    2010-02-26

    Objective: The accumulation of lipids in macrophages contributes to the development of atherosclerosis. Strategies to reduce lipid accumulation in macrophages may have therapeutic potential for preventing and treating atherosclerosis and cardiovascular complications. The antidiabetic drug metformin has been reported to reduce lipid accumulation in adipocytes. In this study, we examined the effects of metformin on lipid accumulation in macrophages and investigated the mechanisms involved. Methods and results: We observed that metformin significantly reduced palmitic acid (PA)-induced intracellular lipid accumulation in macrophages. Metformin promoted the expression of carnitine palmitoyltransferase I (CPT-1), while reduced the expression of fatty acid-binding protein 4 (FABP4) which was involved in PA-induced lipid accumulation. Quantitative real-time PCR showed that metformin regulates FABP4 expression at the transcriptional level. We identified forkhead transcription factor FOXO1 as a positive regulator of FABP4 expression. Inhibiting FOXO1 expression with FOXO1 siRNA significantly reduced basal and PA-induced FABP4 expression. Overexpression of wild-type FOXO1 and constitutively active FOXO1 significantly increased FABP4 expression, whereas dominant negative FOXO1 dramatically decreased FABP4 expression. Metformin reduced FABP4 expression by promoting FOXO1 nuclear exclusion and subsequently inhibiting its activity. Conclusions: Taken together, these results suggest that metformin reduces lipid accumulation in macrophages by repressing FOXO1-mediated FABP4 transcription. Thus, metformin may have a protective effect against lipid accumulation in macrophages and may serve as a therapeutic agent for preventing and treating atherosclerosis in metabolic syndrome.

  13. Heart‐type fatty acid binding protein is a novel prognostic marker in patients with non‐ischaemic dilated cardiomyopathy

    PubMed Central

    Komamura, K; Sasaki, T; Hanatani, A; Kim, J; Hashimura, K; Ishida, Y; Ohkaru, Y; Asayama, K; Tanaka, T; Ogai, A; Nakatani, T; Kitamura, S; Kangawa, K; Miyatake, K; Kitakaze, M

    2006-01-01

    Objective To determine whether concentrations of heart‐type fatty acid binding protein (H‐FABP) measured before hospital discharge predict critical cardiac events in patients with idiopathic dilated cardiomyopathy (DCM). Patients 92 consecutive patients with DCM were enrolled and followed up for four years. Main outcome measures Serum concentrations of H‐FABP, brain natriuretic peptide (BNP), cardiac troponin T before hospital discharge and survival rate. Results 23 patients died of cardiac causes, received a left ventricular assist device or underwent heart transplantation during the four‐year follow up. Univariate analyses showed that New York Heart Association functional class, heart rate, ejection fraction, serum H‐FABP and plasma BNP were significant variables. According to multivariate analysis, serum H‐FABP and plasma BNP concentrations were independent predictors of critical cardiac events. Cardiac troponin T before hospital discharge was not a predictor. The area under the receiver operating characteristic curve for death from critical cardiac events was similar between H‐FABP and BNP. Patients with an H‐FABP concentration at or above the median (⩾ 5.4 ng/ml) had a significantly lower survival rate than those below the median, according to analysis by log rank test (p < 0.0001). When combined with BNP concentration at or above the median (⩾ 138 pg/ml), H‐FABP below the median predicted the worst prognosis among the combinations. Conclusions The concentration of serum H‐FABP before discharge from hospital may be an independent predictor for critical cardiac events in DCM. PMID:16387818

  14. Angiotensin II receptor blockers decrease serum concentration of fatty acid-binding protein 4 in patients with hypertension.

    PubMed

    Furuhashi, Masato; Mita, Tomohiro; Moniwa, Norihito; Hoshina, Kyoko; Ishimura, Shutaro; Fuseya, Takahiro; Watanabe, Yuki; Yoshida, Hideaki; Shimamoto, Kazuaki; Miura, Tetsuji

    2015-04-01

    Elevated circulating fatty acid-binding protein 4 (FABP4/A-FABP/aP2), an adipokine, is associated with obesity, insulin resistance, hypertension and cardiovascular events. However, how circulating FABP4 level is modified by pharmacological agents remains unclear. We here examined the effects of angiotensin II receptor blockers (ARBs) on serum FABP4 level. First, essential hypertensives were treated with ARBs: candesartan (8 mg day(-1); n=7) for 2 weeks, olmesartan (20 mg day(-1); n=9) for 12 weeks, and valsartan (80 mg day(-1); n=94) or telmisartan (40 mg day(-1); n=91) for 8 weeks added to amlodipine (5 mg day(-1)). Treatment with ARBs significantly decreased blood pressure and serum FABP4 concentrations by 8-20% without significant changes in adiposity or lipid variables, though the M value determined by hyperinsulinemic-euglycemic glucose clamp, a sensitive index of insulin sensitivity, was significantly increased by candesartan. Next, alterations in FABP4 secretion from 3T3-L1 adipocytes were examined under several agents. Lipolytic stimulation of the β-adrenoceptor in 3T3-L1 adipocytes by isoproterenol increased FABP4 secretion, and conversely, insulin suppressed FABP4 secretion. However, treatment of 3T3-L1 adipocytes with angiotensin II or ARBs for 2 h had no effect on gene expression or secretion of FABP4 regardless of β-adrenoceptor stimulation. In conclusion, treatment with structurally different ARBs similarly decreases circulating FABP4 concentrations in hypertensive patients as a class effect of ARBs, which is not attributable to blockade of the angiotensin II receptor in adipocytes. Reduction of FABP4 levels by ARBs might be involved in suppression of cardiovascular events. PMID:25672659

  15. Gender difference in plasma fatty-acid-binding protein 4 levels in patients with chronic obstructive pulmonary disease

    PubMed Central

    Zhang, Xue; Li, Diandian; Wang, Hao; Pang, Caishuang; Wu, Yanqiu; Wen, Fuqiang

    2016-01-01

    COPD (chronic obstructive pulmonary disease) is characterized by airway inflammation and increases the likelihood of the development of atherosclerosis. Recent studies have indicated that FABP4 (fatty-acid-binding protein 4), an intracellular lipid chaperone of low molecular mass, plays an important role in the regulation of inflammation and atherosclerosis. We carried out a preliminary clinical study aiming at investigating the relationships between circulating FABP4 levels in patients with COPD and inflammation and lung function. We enrolled 50 COPD patients and 39 healthy controls in the study. Lung function tests were performed in all subjects. Plasma levels of FABP4 and adiponectin, TNFα (tumour necrosis factor α) and CRP (C-reactive protein) were measured. The correlations between FABP4 and lung function, adipokine (adiponectin), inflammatory factors and BMI (body mass index) were analysed. Compared with both males with COPD and healthy females, plasma FABP4 levels in females with COPD were significantly increased. Adiponectin and CRP levels were significantly higher in patients with COPD. Furthermore, we found that FABP4 levels were inversely correlated with FEV1% predicted (FEV1 is forced expiratory volume in 1 s) and positively correlated with adiponectin and TNFα in COPD patients. In addition, a positive correlation between plasma FABP4 and CRP was found in females with COPD. However, FABP4 levels were not correlated with BMI. Our results underline a gender difference in FABP4 secretion in stable COPD patients. Further studies are warranted to clarify the exact role of FABP4 in the pathogenesis of COPD. PMID:26823558

  16. 1H, 15N and 13C resonance assignments of light organ-associated fatty acid-binding protein of Taiwanese fireflies.

    PubMed

    Tseng, Kai-Li; Lee, Yi-Zong; Chen, Yun-Ru; Lyu, Ping-Chiang

    2016-04-01

    Fatty acid-binding proteins (FABPs) are a family of proteins that modulate the transfer of various fatty acids in the cytosol and constitute a significant portion in many energy-consuming cells. The ligand binding properties and specific functions of a particular type of FABP seem to be diverse and depend on the respective binding cavity as well as the cell type from which this protein is derived. Previously, a novel FABP (lcFABP; lc: Luciola cerata) was identified in the light organ of Taiwanese fireflies. The lcFABP was proved to possess fatty acids binding capabilities, especially for fatty acids of length C14-C18. However, the structural details are unknown, and the structure-function relationship has remained to be further investigated. In this study, we finished the (1)H, (15)N and (13)C chemical shift assignments of (15)N/(13)C-enriched lcFABP by solution NMR spectroscopy. In addition, the secondary structure distribution was revealed based on the backbone N, H, Cα, Hα, C and side chain Cβ assignments. These results can provide the basis for further structural exploration of lcFABP. PMID:26373428

  17. Analysis of the regulation of fatty acid binding protein 7 expression in human renal carcinoma cell lines

    PubMed Central

    2011-01-01

    Background Improving the treatment of renal cell carcinoma (RCC) will depend on the development of better biomarkers for predicting disease progression and aiding the design of appropriate therapies. One such marker may be fatty acid binding protein 7 (FABP7), also known as B-FABP and BLBP, which is expressed normally in radial glial cells of the developing central nervous system and cells of the mammary gland. Melanomas, glioblastomas, and several types of carcinomas, including RCC, overexpress FABP7. The abundant expression of FABP7 in primary RCCs compared to certain RCC-derived cell lines may allow the definition of the molecular components of FABP7's regulatory system. Results We determined FABP7 mRNA levels in six RCC cell lines. Two were highly expressed, whereas the other and the embryonic kidney cell line (HEK293) were weakly expressed FABP7 transcripts. Western blot analysis of the cell lines detected strong FABP7 expression only in one RCC cell line. Promoter activity in the RCC cell lines was 3- to 21-fold higher than that of HEK293. Deletion analysis demonstrated that three FABP7 promoter regions contributed to upregulated expression in RCC cell lines, but not in the HEK293 cell. Competition analysis of gel shifts indicated that OCT1, OCT6, and nuclear factor I (NFI) bound to the FABP7 promoter region. Supershift experiments indicated that BRN2 (POU3F2) and NFI bound to the FABP7 promoter region as well. There was an inverse correlation between FABP7 promoter activity and BRN2 mRNA expression. The FABP7-positive cell line's NFI-DNA complex migrated faster than in other cell lines. Levels of NFIA mRNA were higher in the HEK293 cell line than in any of the six RCC cell lines. In contrast, NFIC mRNA expression was lower in the HEK293 cell line than in the six RCC cell lines. Conclusions Three putative FABP7 promoter regions drive reporter gene expression in RCC cell lines, but not in the HEK293 cell line. BRN2 and NFI may be key factors regulating the

  18. Identification of the messenger RNA for human cutaneous fatty acid-binding protein as a metastasis inducer.

    PubMed

    Jing, C; Beesley, C; Foster, C S; Rudland, P S; Fujii, H; Ono, T; Chen, H; Smith, P H; Ke, Y

    2000-05-01

    Using our recently developed systematic differential display and complete comparison of gene expression approaches combined with other methods, we have identified a large number of mRNAs that are expressed differentially between benign and malignant human cells. One such mRNA that is common to prostate and breast carcinoma cell lines encodes the human cutaneous fatty acid-binding protein (C-FABP). Northern and slot blot analyses confirm that the expression levels of C-FABP mRNA in the malignant prostate and breast carcinoma cell lines are 4.9+/-0.9- to 16.9+/-2.1-fold higher than those expressed in the benign cell lines. A similar difference between the benign and malignant cell lines was also detected at the protein level. In situ hybridization experiments have detected overexpression of the mRNA for C-FABP in human prostate carcinoma tissues. Transfection of a C-FABP expression construct into the benign, nonmetastatic rat mammary epithelial cell line Rama 37 and inoculation of the C-FABP expression transfectants into syngeneic Wistar-Furth rats produce a significant number (P < 0.05) of animals with metastases (6 of 26 animals), whereas the control transfectants generated by the vector alone yield no such metastases. Measurements of mRNA and protein levels with Northern and Western blotting show that C-FABP is not expressed in the control transfectant cells produced by the vector alone but is highly expressed in the pool of C-FABP transfectants and-the sublines established from their metastases. Immunocytochemical staining with antibodies to C-FABP shows that C-FABP is not expressed in the primary tumors developed from the control transfectants that have failed to metastasize, but it is expressed in both the primary tumors developed from the C-FABP transfectants and their metastases. Reinoculation of the sublines established from metastases in syngeneic rats has produced a higher proportion (50%) of animals (7 of 14 animals) with metastases than that obtained in

  19. A Study on the Role of Heart Type Fatty Acid Binding Protein in the Diagnosis of Acute Myocardial Infarction

    PubMed Central

    Kabekkodu, Shama Prakash; Mananje, Sudhindra Rao

    2016-01-01

    Introduction Heart type Fatty Acid Binding Protein (H-FABP) has been proposed as an early cardiac biomarker for the diagnosis of acute myocardial Infarction (AMI) using animal models and clinical samples. Aim The study aimed to evaluate the role of H-FABP in early detection of AMI by comparing its sensitivity, specificity and predictive value with Creatinine Kinase-MB (CK-MB) and Cardiac Troponin I (cTnI). Materials and Methods This is a cross-sectional descriptive study of 50 patients admitted with the diagnosis of AMI at a tertiary care hospital in South India. The study group was categorised in to those coming to the hospital within four hours of symptom onset and those coming in between 4 to 12 hours. H-FABP was compared with those of troponin T and myoglobin tests. Results Among patients presenting within four hours of symptom onset, the sensitivity of H-FABP was 60% and was significantly higher than that of cardiac Troponin I (cTnI, 18.8%) and Creatinine Kinase (CK)-MB (12.5%). But specificity was only 23.53% and was less than that of cTnI (66.67%) and CK-MB (100%). In patients presenting during 4 to 12 hours of symptom onset, the sensitivity of H-FABP was 86.96% which was comparable to that of cTnI (90.9%) and CK-MB (77.3%). The specificity was 60% in the 4-12 hours group which was comparable to that of cTnI (50%) and CK-MB (50%). Conclusion The H-FABP is a sensitive biomarker for the diagnosis of AMI in the initial hours after symptom onset when the standard biomarkers may not be elevated, but it is less specific. During 4-12 hours of symptom onset it is as sensitive and specific as standard cardiac biomarkers troponin and CK-MB. Due to these factors H-FABP can be considered as a promising cardiac biomarker which can be used along with troponins and CK-MB at present. PMID:26894106

  20. The effect of charge reversal mutations in the alpha-helical region of liver fatty acid binding protein on the binding of fatty-acyl CoAs, lysophospholipids and bile acids.

    PubMed

    Hagan, Robert M; Davies, Joanna K; Wilton, David C

    2002-10-01

    Liver fatty acid binding protein (LFABP) is unique among the various types of FABPs in that it can bind a variety of ligands in addition to fatty acids. LFABP is able to bind long chain fatty acids with a 2:1 stoichiometry and the crystal structure has identified two fatty acid binding sites in the binding cavity. The presumed primary site (site 1) involves the fatty acid binding with the carboxylate group buried in the cavity whereas the fatty acid at site 2 has the carboxylate group solvent-exposed within the ligand portal region and in the vicinity of alpha-helix II. The alpha-helical region contains three cationic residues, K20, K31, K33 and modelling studies suggest that K31 on alpha-helix II could make an electrostatic contribution to anionic ligands binding to site 2. The preparation of three charge reversal mutants of LFABP, K20E, K31E and K33E has allowed an investigation of the role of site 2 in ligand binding, particularly those ligands with a bulky anionic head group. The binding of oleoyl CoA, lysophosphatidic acid, lysophosphatidylcholine, lithocholic acid and taurolithocholate 3-sulphate to LFABP has been studied using the alpha-helical mutants. The results support the concept that such ligands bind at site 2 of LFABP where solvent exposure allows the accommodation of their bulky anionic group. PMID:12479568

  1. Liver fatty acid binding protein (L-Fabp) modifies intestinal fatty acid composition and adenoma formation in ApcMin/+ mice

    PubMed Central

    Dharmarajan, Sekhar; Newberry, Elizabeth P.; Montenegro, Grace; Nalbantoglu, ILKe; Davis, Victoria R.; Clanahan, Michael J.; Blanc, Valerie; Xie, Yan; Luo, Jianyang; Fleshman, James W.; Kennedy, Susan; Davidson, Nicholas O.

    2013-01-01

    Evidence suggests a relationship between dietary fat intake, obesity and colorectal cancer, implying a role for fatty acid (FA) metabolism in intestinal tumorigenesis that is incompletely understood. Liver fatty acid binding protein (L-Fabp), a dominant intestinal FA binding protein, regulates intestinal FA trafficking and metabolism and L-Fabp deletion attenuates diet-induced obesity. Here we examined whether changes in intestinal FA metabolism following L-Fabp deletion modify adenoma development in ApcMin/+ mice. Compound L-Fabp−/−ApcMin/+ mice were generated and fed a 10% fat diet balanced equally between saturated, monounsaturated and polyunsaturated fat. L-Fabp−/−ApcMin/+ mice displayed significant reductions in adenoma number and total polyp area compared to ApcMin/+controls, reflecting a significant shift in distribution toward smaller polyps. Adenomas from L-Fabp−/−ApcMin/+ mice exhibited reductions in cellular proliferation, high-grade dysplasia and nuclear β-catenin translocation. Intestinal FA content was increased in L-Fabp−/−ApcMin/+ mice and lipidomic profiling of intestinal mucosa revealed significant shifts to polyunsaturated FA species with reduced saturated FA species. L-Fabp−/−ApcMin/+mice also demonstrated corresponding changes in mRNA expression of enzymes involved in FA elongation and desaturation. Furthermore, adenomas from L-Fabp−/−ApcMin/+mice displayed significant reductions in mRNA abundance of nuclear hormone receptors involved in cellular proliferation and in enzymes involved in lipogenesis. These findings collectively implicate L-Fabp as an important genetic modifier of intestinal tumorigenesis and identify FA trafficking and metabolic compartmentalization as an important pathway linking dietary fat intake, obesity and intestinal tumor formation. PMID:23921281

  2. Uncoupling of Obesity from Insulin Resistance Through a Targeted Mutation in aP2, the Adipocyte Fatty Acid Binding Protein

    NASA Astrophysics Data System (ADS)

    Hotamisligil, Gokhan S.; Johnson, Randall S.; Distel, Robert J.; Ellis, Ramsey; Papaioannou, Virginia E.; Spiegelman, Bruce M.

    1996-11-01

    Fatty acid binding proteins (FABPs) are small cytoplasmic proteins that are expressed in a highly tissue-specific manner and bind to fatty acids such as oleic and retinoic acid. Mice with a null mutation in aP2, the gene encoding the adipocyte FABP, were developmentally and metabolically normal. The aP2-deficient mice developed dietary obesity but, unlike control mice, they did not develop insulin resistance or diabetes. Also unlike their obese wild-type counterparts, obese aP2-/- animals failed to express in adipose tissue tumor necrosis factor-α (TNF-α), a molecule implicated in obesity-related insulin resistance. These results indicate that aP2 is central to the pathway that links obesity to insulin resistance, possibly by linking fatty acid metabolism to expression of TNF-α.

  3. N-Benzyl-indolo carboxylic acids: Design and synthesis of potent and selective adipocyte fatty-acid binding protein (A-FABP) inhibitors.

    PubMed

    Barf, Tjeerd; Lehmann, Fredrik; Hammer, Kristin; Haile, Saba; Axen, Eva; Medina, Carmen; Uppenberg, Jonas; Svensson, Stefan; Rondahl, Lena; Lundbäck, Thomas

    2009-03-15

    Small molecule inhibitors of adipocyte fatty-acid binding protein (A-FABP) have gained renewed interest following the recent publication of pharmacologically beneficial effects of such inhibitors. Despite the potential utility of selective A-FABP inhibitors within the fields of metabolic disease, inflammation and atherosclerosis, there are few examples of useful A-FABP inhibitors in the public domain. Herein, we describe the optimization of N-benzyl-tetrahydrocarbazole derivatives through the use of co-crystal structure guided medicinal chemistry efforts. This led to the identification of a potent and selective class of A-FABP inhibitors as illustrated by N-benzyl-hexahydrocyclohepta[b]indole 30. PMID:19217286

  4. Development of the Brazilian Anti Schistosomiasis Vaccine Based on the Recombinant Fatty Acid Binding Protein Sm14 Plus GLA-SE Adjuvant.

    PubMed

    Tendler, Miriam; Almeida, Marilia; Simpson, Andrew

    2015-01-01

    Data herein reported and discussed refer to vaccination with the recombinant fatty acid binding protein (FABP) family member of the schistosomes, called Sm14. This antigen was discovered and developed under a Brazilian platform led by the Oswaldo Cruz Foundation, from the Health Ministry in Brazil, and was assessed for safety and immunogenicity in healthy volunteers. This paper reviews past and recent outcomes of developmental phases of the Sm14-based anti schistosomiasis vaccine addressed to, ultimately, impact transmission of the second most prevalent parasitic endemic disease worldwide. PMID:26029206

  5. Characterization of the sources of protein-ligand affinity: 1-sulfonato-8-(1')anilinonaphthalene binding to intestinal fatty acid binding protein.

    PubMed Central

    Kirk, W R; Kurian, E; Prendergast, F G

    1996-01-01

    1-Sulfonato-8-(1')anilinonaphthalene (1,8-ANS) was employed as a fluorescent probe of the fatty acid binding site of recombinant rat intestinal fatty acid binding protein (1-FABP). The enhancement of fluorescence upon binding allowed direct determination of binding affinity by fluorescence titration experiments, and measurement of the effects on that affinity of temperature, pH, and ionic strength. Solvent isotope effects were also determined. These data were compared to results from isothermal titration calorimetry. We obtained values for the enthalpy and entropy of this interaction at a variety of temperatures, and hence determined the change in heat capacity of the system consequent upon binding. The ANS-1-FABP is enthalpically driven; above approximately 14 degrees C it is entropically opposed, but below this temperature the entropy makes a positive contribution to the binding. The changes we observe in both enthalpy and entropy of binding with temperature can be derived from the change in heat capacity upon binding by integration, which demonstrates the internal consistency of our results. Bound ANS is displaced by fatty acids and can itself displace fatty acids bound to I-FABP. The binding site for ANS appears to be inside the solvent-containing cavity observed in the x-ray crystal structure, the same cavity occupied by fatty acid. From the fluorescence spectrum and from an inversion of the Debye-Hueckel formula for the activity coefficients as a function of added salt, we inferred that this cavity is fairly polar in character, which is in keeping with inferences drawn from the x-ray structure. The binding affinity of ANS is considered to be a consequence of both electrostatic and conditional hydrophobic effects. We speculate that the observed change in heat capacity is produced mainly by the displacement of strongly hydrogen-bonded waters from the protein cavity. PMID:8770188

  6. A Novel Fatty Acid-Binding Protein-Like Carotenoid-Binding Protein from the Gonad of the New Zealand Sea Urchin Evechinus chloroticus

    PubMed Central

    Pilbrow, Jodi; Sabherwal, Manya; Garama, Daniel; Carne, Alan

    2014-01-01

    A previously uncharacterized protein with a carotenoid-binding function has been isolated and characterized from the gonad of the New Zealand sea urchin Evechinus chloroticus. The main carotenoid bound to the protein was determined by reversed phase-high performance liquid chromatography to be 9′-cis-echinenone and hence this 15 kDa protein has been called an echinenone-binding protein (EBP). Purification of the EBP in quantity from the natural source proved to be challenging. However, analysis of EBP by mass spectrometry combined with information from the Strongylocentrotus purpuratus genome sequence and the recently published E. chloroticus transcriptome database, enabled recombinant expression of wild type EBP and also of a cysteine61 to serine mutant that had improved solubility characteristics. Circular dichroism data and ab initio structure prediction suggests that the EBP adopts a 10-stranded β-barrel fold consistent with that of fatty acid-binding proteins. Therefore, EBP may represent the first report of a fatty acid-binding protein in complex with a carotenoid. PMID:25192378

  7. A novel fatty acid-binding protein-like carotenoid-binding protein from the gonad of the New Zealand sea urchin Evechinus chloroticus.

    PubMed

    Pilbrow, Jodi; Sabherwal, Manya; Garama, Daniel; Carne, Alan

    2014-01-01

    A previously uncharacterized protein with a carotenoid-binding function has been isolated and characterized from the gonad of the New Zealand sea urchin Evechinus chloroticus. The main carotenoid bound to the protein was determined by reversed phase-high performance liquid chromatography to be 9'-cis-echinenone and hence this 15 kDa protein has been called an echinenone-binding protein (EBP). Purification of the EBP in quantity from the natural source proved to be challenging. However, analysis of EBP by mass spectrometry combined with information from the Strongylocentrotus purpuratus genome sequence and the recently published E. chloroticus transcriptome database, enabled recombinant expression of wild type EBP and also of a cysteine61 to serine mutant that had improved solubility characteristics. Circular dichroism data and ab initio structure prediction suggests that the EBP adopts a 10-stranded β-barrel fold consistent with that of fatty acid-binding proteins. Therefore, EBP may represent the first report of a fatty acid-binding protein in complex with a carotenoid. PMID:25192378

  8. Human cutaneous fatty acid-binding protein induces metastasis by up-regulating the expression of vascular endothelial growth factor gene in rat Rama 37 model cells.

    PubMed

    Jing, C; Beesley, C; Foster, C S; Chen, H; Rudland, P S; West, D C; Fujii, H; Smith, P H; Ke, Y

    2001-06-01

    Human cutaneous fatty acid-binding protein (C-FABP) gene is capable of inducing the metastatic phenotype when overexpressed in nonmetastatic rat Rama 37 cells. However, the mechanism of how it induces metastasis is not clear. Northern and slot blot analyses revealed that expression of the endogenous vascular endothelial growth factor (VEGF) gene was increased by 3.8-5.2-fold in the C-FABP-transfected cells (pSV-CFABP-R37) and in their metastatic sublines (e.g., Met-1) when compared with that in the nonmetastatic control transfectant pSV-R37 cells generated by transfection of only plasmid DNA. Higher levels of VEGF immunoreactive protein were also secreted from the malignant C-FABP-expressing cells. Reverse transcription-PCR detected two VEGF transcript isoforms, VEGF(164) and VEGF(188), in both the nonmetastatic control transfectant pSV-R37 cells and the malignant metastatic Met-1 cells. Chick chorioallantoic membrane assays showed that the conditioned medium of the control pSV-R37 cells possessed only very weak angiogenic activity, whereas conditioned media from the metastatic C-FABP transfectants and their sublines were strongly angiogenic and could be inhibited by antibodies to VEGF. Transfection of VEGF(164) cDNA in an expression vector into nonmetastatic Rama 37 cells produced a cell clone (R37-VEGF-2) that expressed high levels of VEGF. Inoculation of R37-VEGF-2 cells into syngeneic Wistar Furth rats produced metastases in a significant number (Fisher's exact test, P < 0.01) of animals (18 of 31 animals), whereas the control, vector alone-transfected R37-PSV cells produced no metastases (0 of 30 animals). Immunocytochemical methods demonstrated a strong positive staining for VEGF and an increased microvessel density in the primary tumors produced from PSV-VEGF-2 cells in comparison with tumors produced from control transfectants. Immunocytochemical staining for factor VIII detected a 3.5-fold increase in microvessel density of the primary tumors produced by

  9. Characterization of Naphthaleneacetic Acid Binding to Receptor Sites on Cellular Membranes of Maize Coleoptile Tissue 1

    PubMed Central

    Ray, Peter M.; Dohrmann, Ulrike; Hertel, Rainer

    1977-01-01

    Characteristics of and optimum conditions for saturable (“specific”) binding of [14C]naphthaleneacetic acid to sites located on membranous particles from maize (Zea mays L.) coleoptiles are described. Most, if not all, of the specific binding appears to be due to a single kinetic class of binding sites having a KD of 5 to 7 × 10−7m for naphthalene-1-acetic acid (NAA). Binding of NAA is insensitive to high monovalent salt concentrations, indicating that binding is not primarily ionic. However, specific binding is inhibited by Mg2+ or Ca2+ above 5 mm. Specific binding is improved by organic acids, especially citrate. Binding is heat-labile and is sensitive to agents that act either on proteins or on lipids. Specific binding is reversibly inactivated by reducing agents such as dithioerythritol; a reducible group, possibly a disulfide group, may be located at the binding site and required for its function. The affinity of the specific binding sites for auxins is modified by an unidentified dialyzable, heat-stable, apparently amphoteric, organic factor (“supernatant factor”) found in maize tissue. PMID:16659851

  10. Sulfate inhibits ( sup 14 C)phosphonoformic acid binding to renal brush-border membranes

    SciTech Connect

    Tenenhouse, H.S.; Lee, J. )

    1990-08-01

    To examine the specificity of the phosphonoformic acid (PFA) interaction with the Na(+)-dependent phosphate transporter of mouse renal brush-border membrane vesicles, we compared the effects of anions on Na(+)-dependent (14C)PFA binding and Na(+)-dependent phosphate transport. Inhibition of PFA binding was achieved by PFA (% control = 0 +/- 1), sulfate (15 +/- 2), arsenate (35 +/- 1), phosphate (59 +/- 2), and nitrate (68 +/- 4), whereas inhibition of phosphate transport was only apparent with phosphate (0 +/- 1), PFA (22 +/- 4), and arsenate (37 +/- 5). Succinate and gluconate had no effect on either Na(+)-dependent process. Under conditions where Na(+)-dependent PFA binding was maximally inhibited by phosphate (% control = 65 +/- 4), further inhibition could be achieved by sulfate (26 +/- 5%). Na(+)-dependent PFA binding was competitively inhibited by phosphate (apparent Ki = 8.9 +/- 1.2 mM) and noncompetitively inhibited by sulfate (apparent Ki = 2.6 +/- 0.5 mM). We found that PFA inhibited Na(+)-dependent sulfate transport (50% inhibition at 9 mM PFA) as well as Na(+)-dependent phosphate transport (50% inhibition at 0.5 mM PFA). We also examined the pH dependence of Na(+)-dependent PFA binding and Na(+)-dependent phosphate and sulfate transport. PFA binding was optimal at pH = 7.4, whereas phosphate transport increased with increasing pH, and sulfate transport increased with decreasing pH.

  11. Different functions of intestinal and liver-type fatty acid-binding proteins in intestine and in whole body energy homeostasis.

    PubMed

    Lagakos, William Stacy; Gajda, Angela Marie; Agellon, Luis; Binas, Bert; Choi, Victor; Mandap, Bernadette; Russnak, Timothy; Zhou, Yin Xiu; Storch, Judith

    2011-05-01

    It has long been known that mammalian enterocytes coexpress two members of the fatty acid-binding protein (FABP) family, the intestinal FABP (IFABP) and the liver FABP (LFABP). Both bind long-chain fatty acids and have similar though not identical distributions in the intestinal tract. While a number of in vitro properties suggest the potential for different functions, the underlying reasons for expression of both proteins in the same cells are not known. Utilizing mice genetically lacking either IFABP or LFABP, we directly demonstrate that each of the enterocyte FABPs participates in specific pathways of intestinal lipid metabolism. In particular, LFABP appears to target fatty acids toward oxidative pathways and dietary monoacylglycerols toward anabolic pathways, while IFABP targets dietary fatty acids toward triacylglycerol synthesis. The two FABP-null models also displayed differences in whole body response to fasting, with LFABP-null animals losing less fat-free mass and IFABP-null animals losing more fat mass relative to wild-type mice. The metabolic changes observed in both null models appear to occur by nontranscriptional mechanisms, supporting the hypothesis that the enterocyte FABPs are specifically trafficking their ligands to their respective metabolic fates. PMID:21350192

  12. Molecular cloning and functional analysis of the fatty acid-binding protein (Sp-FABP) gene in the mud crab (Scylla paramamosain)

    PubMed Central

    Zeng, Xianglan; Ye, Haihui; Yang, Ya’nan; Wang, Guizhong; Huang, Huiyang

    2013-01-01

    Intracellular fatty acid-binding proteins (FABPs) are multifunctional cytosolic lipid-binding proteins found in vertebrates and invertebrates. In this work, we used RACE to obtain a full-length cDNA of Sp-FABP from the mud crab Scylla paramamosain. The open reading frame of the full length cDNA (886 bp) encoded a 136 amino acid polypeptide that showed high homology with related genes from other species. Real-time quantitative PCR identified variable levels of Sp-FABP transcripts in epidermis, eyestalk, gill, heart, hemocytes, hepatopancreas, muscle, ovary, stomach and thoracic ganglia. In ovaries, Sp-FABP expression increased gradually from stage I to stage IV of development and decreased in stage V. Sp-FABP transcripts in the hepatopancreas and hemocytes were up-regulated after a bacterial challenge with Vibrio alginnolyficus. These results suggest that Sp-FABP may be involved in the growth, reproduction and immunity of the mud crab. PMID:23569421

  13. Role of Cardiac Myocytes Heart Fatty Acid Binding Protein Depletion (H-FABP) in Early Myocardial Infarction in Human Heart (Autopsy Study)

    PubMed Central

    Shabaiek, Amany; Ismael, Nour El-Hoda; Elsheikh, Samar; Amin, Hebat Allah

    2016-01-01

    BACKGROUND: Many immunohistochemical markers have been used in the postmortem detection of early myocardial infarction. AIM: In the present study we examined the role of Heart-type fatty acid binding protein (H-FABP), in the detection of early myocardial infarction. MATERIAL AND METHODS: We obtained samples from 40 human autopsy hearts with/without histopathological signs of ischemia. RESULTS: All cases of definite and probable myocardial infarction showed a well-defined area of H-FABP depletion. All of the control cases showed strong H-FABP expression, except two markedly autolysed myocardial samples that showed affected antigenicity. CONCLUSION: Thus, we suggest H-FABP as being one of the valuable tools facing the problem of postmortem detection of early myocardial infarction/ischemia, but not in autolysis.

  14. Long-Term Effect of Docosahexaenoic Acid Feeding on Lipid Composition and Brain Fatty Acid-Binding Protein Expression in Rats

    PubMed Central

    Elsherbiny, Marwa E.; Goruk, Susan; Monckton, Elizabeth A.; Richard, Caroline; Brun, Miranda; Emara, Marwan; Field, Catherine J.; Godbout, Roseline

    2015-01-01

    Arachidonic (AA) and docosahexaenoic acid (DHA) brain accretion is essential for brain development. The impact of DHA-rich maternal diets on offspring brain fatty acid composition has previously been studied up to the weanling stage; however, there has been no follow-up at later stages. Here, we examine the impact of DHA-rich maternal and weaning diets on brain fatty acid composition at weaning and three weeks post-weaning. We report that DHA supplementation during lactation maintains high DHA levels in the brains of pups even when they are fed a DHA-deficient diet for three weeks after weaning. We show that boosting dietary DHA levels for three weeks after weaning compensates for a maternal DHA-deficient diet during lactation. Finally, our data indicate that brain fatty acid binding protein (FABP7), a marker of neural stem cells, is down-regulated in the brains of six-week pups with a high DHA:AA ratio. We propose that elevated levels of DHA in developing brain accelerate brain maturation relative to DHA-deficient brains. PMID:26506385

  15. Epidermal Fatty Acid Binding Protein (E-FABP) Is Not Required for the Generation or Maintenance of Effector and Memory T Cells following Infection with Listeria monocytogenes.

    PubMed

    Li, Bing; Schmidt, Nathan W

    2016-01-01

    Following activation of naïve T cells there are dynamic changes in the metabolic pathways used by T cells to support both the energetic needs of the cell and the macromolecules required for growth and proliferation. Among other changes, lipid metabolism undergoes dynamic transitions between fatty acid oxidation and fatty acid synthesis as cells progress from naïve to effector and effector to memory T cells. The hydrophobic nature of lipids requires that they be bound to protein chaperones within a cell. Fatty acid binding proteins (FABPs) represent a large class of lipid chaperones, with epidermal FABP (E-FABP) expressed in T cells. The objective of this study was to determine the contribution of E-FABP in antigen-specific T cell responses. Following infection with Listeria monocytogenes, we observed similar clonal expansion, contraction and formation of memory CD8 T cells in WT and E-FABP-/- mice, which also exhibited similar phenotypic and functional characteristics. Analysis of Listeria-specific CD4 T cells also revealed no defect in the expansion, contraction, and formation of memory CD4 T cells in E-FABP-/- mice. These data demonstrate that E-FABP is dispensable for antigen-specific T cell responses following a bacterial infection. PMID:27588422

  16. Direct comparison of mice null for liver or intestinal fatty acid-binding proteins reveals highly divergent phenotypic responses to high fat feeding.

    PubMed

    Gajda, Angela M; Zhou, Yin Xiu; Agellon, Luis B; Fried, Susan K; Kodukula, Sarala; Fortson, Walter; Patel, Khamoshi; Storch, Judith

    2013-10-18

    The enterocyte expresses two fatty acid-binding proteins (FABP), intestinal FABP (IFABP; FABP2) and liver FABP (LFABP; FABP1). LFABP is also expressed in liver. Despite ligand transport and binding differences, it has remained uncertain whether these intestinally coexpressed proteins, which both bind long chain fatty acids (FA), are functionally distinct. Here, we directly compared IFABP(-/-) and LFABP(-/-) mice fed high fat diets containing long chain saturated or unsaturated fatty acids, reasoning that providing an abundance of dietary lipid would reveal unique functional properties. The results showed that mucosal lipid metabolism was indeed differentially modified, with significant decreases in FA incorporation into triacylglycerol (TG) relative to phospholipid (PL) in IFABP(-/-) mice, whereas LFABP(-/-) mice had reduced monoacylglycerol incorporation in TG relative to PL, as well as reduced FA oxidation. Interestingly, striking differences were found in whole body energy homeostasis; LFABP(-/-) mice fed high fat diets became obese relative to WT, whereas IFABP(-/-) mice displayed an opposite, lean phenotype. Fuel utilization followed adiposity, with LFABP(-/-) mice preferentially utilizing lipids, and IFABP(-/-) mice preferentially metabolizing carbohydrate for energy production. Changes in body weight and fat may arise, in part, from altered food intake; mucosal levels of the endocannabinoids 2-arachidonoylglycerol and arachidonoylethanolamine were elevated in LFABP(-/-), perhaps contributing to increased energy intake. This direct comparison provides evidence that LFABP and IFABP have distinct roles in intestinal lipid metabolism; differential intracellular functions in intestine and in liver, for LFABP(-/-) mice, result in divergent downstream effects at the systemic level. PMID:23990461

  17. Effect of endurance training and/or fish oil supplemented diet on cytoplasmic fatty acid binding protein in rat skeletal muscles and heart.

    PubMed

    Clavel, Stéphan; Farout, L; Briand, M; Briand, Y; Jouanel, P

    2002-07-01

    Endurance training and/or a fish oil supplemented diet affect cytoplasmic fatty acid binding protein (FABP(c)) content in rat skeletal muscles and heart. After 8 weeks of swimming, trained rats exhibited higher FABP(c) content in the extensor digitorum longus (EDL) and in the gastrocnemius than did control rats (30%). The FABP(c) increase was associated with an increase of citrate synthase activity (85% and 93%, respectively, in the two muscles), whereas lactate dehydrogenase activity decreased significantly. In contrast, in the soleus and in the heart we did not observe any effect of exercise either on FABP(c) or on the metabolic profile. Therefore, increasing oxidative capacities of muscle by exercise resulted in a concomitant increase of the FABP(c) content. Giving a polyunsaturated fatty acid (omega-3) supplemented diet for eight weeks induced a large rise of the FABP(c) in EDL (300%), gastrocnemius (250%), soleus (50%) and heart (15%) without a concurrent accumulation of intramuscular triglycerides or modification of the citrate synthase activity, suggesting that polyunsaturated fatty acids may increase FABP(c) content by up-regulating fatty acid metabolism genes via peroxisome proliferator-activated receptor alpha activation. Endurance trained rats fed with an omega-3 diet had similar FABP(c) content in the gastrocnemius muscle when compared to sedentary omega-3 fed rats, whereas an additive effect of exercise and diet was observed in the EDL. The FABP(c) in the soleus and in the heart of rats fed with omega-3 supplements remained constant whether rats performed exercise or not. As a result, both exercise and omega-3-enriched diet influenced FABP(c) content in muscle. These two physiological treatments presumably acted on FABP(c) content by increasing fatty acid flux within the cell. PMID:12111278

  18. Molecular cloning and tissue expression of the fatty acid-binding protein (Es-FABP) gene in female Chinese mitten crab (Eriocheir sinensis)

    PubMed Central

    2010-01-01

    Background Fatty acid-binding proteins (FABPs), small cytosolic proteins that function in the uptake and utilization of fatty acids, have been extensively studied in higher vertebrates while invertebrates have received little attention despite similar nutritional requirements during periods of reproductive activity. Results Therefore, a cDNA encoding Eriocheir sinensis FABP (Es-FABP) was cloned based upon EST analysis of a hepatopancreas cDNA library. The full length cDNA was 750 bp and encoded a 131 aa polypeptide that was highly homologous to related genes reported in shrimp. The 9108 bp Es-FABP gene contained four exons that were interrupted by three introns, a genomic organization common among FABP multigene family members in vertebrates. Gene expression analysis, as determined by RT-PCR, revealed the presence of Es-FABP transcripts in hepatopancreas, hemocytes, ovary, gills, muscle, thoracic ganglia, heart, and intestine, but not stomach or eyestalk. Real-time quantitative RT-PCR analysis revealed that Es-FABP expression in ovary, hemocytes, and hepatopancreas was dependent on the status of ovarian development, with peak expression observed in January. Conclusions Evidence provided in the present report supports a role of Es-FABP in lipid transport during the period of rapid ovarian growth in E. sinensis, and indirectly confirms the participation of the hepatopancreas, ovary, and hemocytes in lipid nutrient absorption and utilization processes. PMID:20846381

  19. Structural basis for the ligand-binding specificity of fatty acid-binding proteins (pFABP4 and pFABP5) in gentoo penguin.

    PubMed

    Lee, Chang Woo; Kim, Jung Eun; Do, Hackwon; Kim, Ryeo-Ok; Lee, Sung Gu; Park, Hyun Ho; Chang, Jeong Ho; Yim, Joung Han; Park, Hyun; Kim, Il-Chan; Lee, Jun Hyuck

    2015-09-11

    Fatty acid-binding proteins (FABPs) are involved in transporting hydrophobic fatty acids between various aqueous compartments of the cell by directly binding ligands inside their β-barrel cavities. Here, we report the crystal structures of ligand-unbound pFABP4, linoleate-bound pFABP4, and palmitate-bound pFABP5, obtained from gentoo penguin (Pygoscelis papua), at a resolution of 2.1 Å, 2.2 Å, and 2.3 Å, respectively. The pFABP4 and pFABP5 proteins have a canonical β-barrel structure with two short α-helices that form a cap region and fatty acid ligand binding sites in the hydrophobic cavity within the β-barrel structure. Linoleate-bound pFABP4 and palmitate-bound pFABP5 possess different ligand-binding modes and a unique ligand-binding pocket due to several sequence dissimilarities (A76/L78, T30/M32, underlining indicates pFABP4 residues) between the two proteins. Structural comparison revealed significantly different conformational changes in the β3-β4 loop region (residues 57-62) as well as the flipped Phe60 residue of pFABP5 than that in pFABP4 (the corresponding residue is Phe58). A ligand-binding study using fluorophore displacement assays shows that pFABP4 has a relatively strong affinity for linoleate as compared to pFABP5. In contrast, pFABP5 exhibits higher affinity for palmitate than that for pFABP4. In conclusion, our high-resolution structures and ligand-binding studies provide useful insights into the ligand-binding preferences of pFABPs based on key protein-ligand interactions. PMID:26206084

  20. Direct Comparison of Mice Null for Liver or Intestinal Fatty Acid-binding Proteins Reveals Highly Divergent Phenotypic Responses to High Fat Feeding*

    PubMed Central

    Gajda, Angela M.; Zhou, Yin Xiu; Agellon, Luis B.; Fried, Susan K.; Kodukula, Sarala; Fortson, Walter; Patel, Khamoshi; Storch, Judith

    2013-01-01

    The enterocyte expresses two fatty acid-binding proteins (FABP), intestinal FABP (IFABP; FABP2) and liver FABP (LFABP; FABP1). LFABP is also expressed in liver. Despite ligand transport and binding differences, it has remained uncertain whether these intestinally coexpressed proteins, which both bind long chain fatty acids (FA), are functionally distinct. Here, we directly compared IFABP−/− and LFABP−/− mice fed high fat diets containing long chain saturated or unsaturated fatty acids, reasoning that providing an abundance of dietary lipid would reveal unique functional properties. The results showed that mucosal lipid metabolism was indeed differentially modified, with significant decreases in FA incorporation into triacylglycerol (TG) relative to phospholipid (PL) in IFABP−/− mice, whereas LFABP−/− mice had reduced monoacylglycerol incorporation in TG relative to PL, as well as reduced FA oxidation. Interestingly, striking differences were found in whole body energy homeostasis; LFABP−/− mice fed high fat diets became obese relative to WT, whereas IFABP−/− mice displayed an opposite, lean phenotype. Fuel utilization followed adiposity, with LFABP−/− mice preferentially utilizing lipids, and IFABP−/− mice preferentially metabolizing carbohydrate for energy production. Changes in body weight and fat may arise, in part, from altered food intake; mucosal levels of the endocannabinoids 2-arachidonoylglycerol and arachidonoylethanolamine were elevated in LFABP−/−, perhaps contributing to increased energy intake. This direct comparison provides evidence that LFABP and IFABP have distinct roles in intestinal lipid metabolism; differential intracellular functions in intestine and in liver, for LFABP−/− mice, result in divergent downstream effects at the systemic level. PMID:23990461

  1. Fatty Acid Binding Protein-1 (FABP1) and the Human FABP1 T94A Variant: Roles in the Endocannabinoid System and Dyslipidemias.

    PubMed

    Schroeder, Friedhelm; McIntosh, Avery L; Martin, Gregory G; Huang, Huan; Landrock, Danilo; Chung, Sarah; Landrock, Kerstin K; Dangott, Lawrence J; Li, Shengrong; Kaczocha, Martin; Murphy, Eric J; Atshaves, Barbara P; Kier, Ann B

    2016-06-01

    The first discovered member of the mammalian FABP family, liver fatty acid binding protein (FABP1, L-FABP), occurs at high cytosolic concentration in liver, intestine, and in the case of humans also in kidney. While the rat FABP1 is well studied, the extent these findings translate to human FABP1 is not clear-especially in view of recent studies showing that endocannabinoids and cannabinoids represent novel rat FABP1 ligands and FABP1 gene ablation impacts the hepatic endocannabinoid system, known to be involved in non-alcoholic fatty liver (NAFLD) development. Although not detectable in brain, FABP1 ablation nevertheless also impacts brain endocannabinoids. Despite overall tertiary structure similarity, human FABP1 differs significantly from rat FABP1 in secondary structure, much larger ligand binding cavity, and affinities/specificities for some ligands. Moreover, while both mouse and human FABP1 mediate ligand induction of peroxisome proliferator activated receptor-α (PPARα), they differ markedly in pattern of genes induced. This is critically important because a highly prevalent human single nucleotide polymorphism (SNP) (26-38 % minor allele frequency and 8.3 ± 1.9 % homozygous) results in a FABP1 T94A substitution that further accentuates these species differences. The human FABP1 T94A variant is associated with altered body mass index (BMI), clinical dyslipidemias (elevated plasma triglycerides and LDL cholesterol), atherothrombotic cerebral infarction, and non-alcoholic fatty liver disease (NAFLD). Resolving human FABP1 and the T94A variant's impact on the endocannabinoid and cannabinoid system is an exciting challenge due to the importance of this system in hepatic lipid accumulation as well as behavior, pain, inflammation, and satiety. PMID:27117865

  2. Effect of liver fatty acid binding protein (FABP) T94A missense mutation on plasma lipoprotein responsiveness to treatment with fenofibrate.

    PubMed

    Brouillette, Charles; Bossé, Yohan; Pérusse, Louis; Gaudet, Daniel; Vohl, Marie-Claude

    2004-01-01

    Fenofibrate, a peroxisome proliferated activated receptor alpha (PPARalpha) agonist, has been shown to decrease plasma triglyceride (TG) and increase plasma high-density lipoprotein (HDL) cholesterol levels despite a large interindividual variation in the response. Fenofibrate-activated PPARalpha binds to a DNA sequence element termed PPAR response element (PPRE) present in regulatory regions of target genes. A PPRE has been identified in the proximal 5' flanking region of the gene encoding the liver fatty acid binding protein (LFABP). LFABP is a small cytosolic protein of 14 kDa present in the liver and the intestine and is a member of the superfamily of the fatty acid binding proteins (FABPs). FABPs play a role in the solubilization of long-chain fatty acids (LCFAs) and their CoA-ester to various intracellular organelles. FABPs serves as intracellular acceptors of LCFAs, and they may also have an impact in ligand-dependent transactivation of PPARs in trafficking LCFAs to the nucleus. Since PPARs are known to regulate the transcription of many genes involved in lipid metabolism, the importance of LFABP in fatty acid uptake has to be considered. The aim of this study was to verify whether genetic variations in the LFABP gene may impact on plasma lipoprotein/lipid levels in the fasting state as well as on the response to a lipid-lowering therapy with fenofibrate on plasma lipids and obesity variables. We also wanted to verify whether the presence of the PPARalpha L162V mutation interacts with genetic variants in LFABP gene. To achieve this goal, we first determined the genomic structure of the human LFABP gene and then designed intronic primers to sequence the coding regions, all exon-intron splicing boundaries, and the promoter region of the gene in 24 patients showing divergent plasma lipoprotein/lipid response to fenofibrate. Sequence analysis revealed the presence of a T94A missense mutation in exon 3. Interspecies comparison revealed that threonine 94 is

  3. A novel polymorphism in the chicken adipocyte fatty acid-binding protein gene (FABP4) that alters ligand-binding and correlates with fatness.

    PubMed

    Wang, Qigui; Guan, Tianzhu; Li, Hui; Bernlohr, David A

    2009-11-01

    Similar to the mammalian FABP4 gene, the chicken (Gallus gallus) FABP4 gene consists of four exons separated by three introns and encodes a 132 amino acid protein termed the adipocyte fatty acid-binding protein (AFABP). In the current study, a novel G/A polymorphism in exon 3 of the chicken FABP4 gene was identified associated with different chicken breeds that leads to either Ser or Asn at amino acid 89 of the AFABP protein. The Baier chicken averages 0.89+/-0.12% abdominal fat and expresses the G allele (Ser 89 isoform) while the Broiler chicken typically has 3.74+/-0.23% abdominal fat and expresses the A allele (Asn 89 isoforms). cDNAs corresponding to the two AFABP isoforms were cloned and expressed in Escherichia coli as GST fusions, purified by using glutathione sepharose 4B chromatography and evaluated for lipid binding using the fluorescent surrogate ligand 1-anilinonaphthalene 8-sulphonic acid (1,8-ANS). The results showed that AFABP Ser89 exhibited a lower ligand-binding affinity with apparent dissociation constants (Kd) of 7.31+/-3.75 microM, while the AFABP Asn89 isoform bound 1,8-ANS with an apparent dissociation constant of 2.99+/-1.00 microM (P=0.02). These results suggest that the Ser89Asn polymorphism may influence chicken AFABP function and ultimately lipid deposition through changing the ligand-binding activity of AFABP. PMID:19595785

  4. The bovine fatty acid binding protein 4 gene is significantly associated with marbling and subcutaneous fat depth in Wagyu x Limousin F2 crosses.

    PubMed

    Michal, J J; Zhang, Z W; Gaskins, C T; Jiang, Z

    2006-08-01

    Fatty acid binding protein 4 (FABP4), which is expressed in adipose tissue, interacts with peroxisome proliferator-activated receptors and binds to hormone-sensitive lipase and therefore, plays an important role in lipid metabolism and homeostasis in adipocytes. The objective of this study was to investigate associations of the bovine FABP4 gene with fat deposition. Both cDNA and genomic DNA sequences of the bovine gene were retrieved from the public databases and aligned to determine its genomic organization. Primers targeting two regions of the FABP4 gene were designed: from nucleotides 5433-6106 and from nucleotides 7417-7868 (AAFC01136716). Direct sequencing of polymerase chain reaction (PCR) products on two DNA pools from high- and low-marbling animals revealed two single nucleotide polymorphisms (SNPs): AAFC01136716.1:g.7516G>C and g.7713G>C. The former SNP, detected by PCR-restriction fragment length polymorphism using restriction enzyme MspA1I, was genotyped on 246 F2 animals in a Waygu x Limousin F2 reference population. Statistical analysis showed that the FABP4 genotype significantly affected marbling score (P = 0.0398) and subcutaneous fat depth (P = 0.0246). The FABP4 gene falls into a suggestive/significant quantitative trait loci interval for beef marbling that was previously reported on bovine chromosome 14 in three other populations. PMID:16879357

  5. Fatty acid binding protein 4 regulates VEGF-induced airway angiogenesis and inflammation in a transgenic mouse model: implications for asthma.

    PubMed

    Ghelfi, Elisa; Yu, Chen-Wei; Elmasri, Harun; Terwelp, Matthew; Lee, Chun G; Bhandari, Vineet; Comhair, Suzy A; Erzurum, Serpil C; Hotamisligil, Gökhan S; Elias, Jack A; Cataltepe, Sule

    2013-04-01

    Neovascularization of the airways occurs in several inflammatory lung diseases, including asthma. Vascular endothelial growth factor (VEGF) plays an important role in vascular remodeling in the asthmatic airways. Fatty acid binding protein 4 (FABP4 or aP2) is an intracellular lipid chaperone that is induced by VEGF in endothelial cells. FABP4 exhibits a proangiogenic function in vitro, but whether it plays a role in modulation of angiogenesis in vivo is not known. We hypothesized that FABP4 promotes VEGF-induced airway angiogenesis and investigated this hypothesis with the use of a transgenic mouse model with inducible overexpression of VEGF165 under a CC10 promoter [VEGF-TG (transgenic) mice]. We found a significant increase in FABP4 mRNA levels and density of FABP4-expressing vascular endothelial cells in mouse airways with VEGF overexpression. FABP4(-/-) mouse airways showed a significant decrease in neovessel formation and endothelial cell proliferation in response to VEGF overexpression. These alterations in airway vasculature were accompanied by attenuated expression of proinflammatory mediators. Furthermore, VEGF-TG/FABP4(-/-) mice showed markedly decreased expression of endothelial nitric oxide synthase, a well-known mediator of VEGF-induced responses, compared with VEGF-TG mice. Finally, the density of FABP4-immunoreactive vessels in endobronchial biopsy specimens was significantly higher in patients with asthma than in control subjects. Taken together, these data unravel FABP4 as a potential target of pathologic airway remodeling in asthma. PMID:23391391

  6. Identification of polymorphism in fatty acid binding protein 3 (FABP3) gene and its association with milk fat traits in riverine buffalo (Bubalus bubalis).

    PubMed

    Dubey, Praveen Kumar; Goyal, Shubham; Mishra, Shailendra Kumar; Arora, Reena; Mukesh, Manishi; Niranjan, Saket Kumar; Kathiravan, Periasamy; Kataria, Ranjit Singh

    2016-04-01

    The fatty acid binding protein 3 (FABP3) gene, known to be associated with fat percentage of milk and meat in bovines, was screened among swamp and riverine buffaloes for polymorphism detection and further association with milk fat contents. An SNP g.307C > T was identified in the intron 2 (+53 exon 2) region of FABP3 gene of Indian buffaloes. The SNP identified was genotyped in 692 animals belonging to 15 riverine, swamp and hybrid (riverine × swamp) buffalo populations of diverse phenotypes and utilities, by PCR-RFLP. A marked contrast was observed between the C and T allele frequencies in three types of buffaloes. The frequency of C allele ranged from 0.67 to 0.96 in pure swamp buffalo populations, with the highest in Mizoram (0.96). Whereas the frequency of T allele was high across all the Indian riverine buffalo breeds, ranging from 0.57 to 0.96. None of the genotypes at FABP3 g.307C > T locus was found to have significant association with milk fat and other production traits in Mehsana dairy buffalo breed. Our study revealed marked differences in the allele frequencies between riverine and swamp buffaloes at FABP3 g.307C > T locus, without any significant association with different milk traits in riverine buffaloes. PMID:26894500

  7. Lack of association between the fatty acid binding protein 2 (FABP2) polymorphism with obesity and insulin resistance in two aboriginal populations from Chile.

    PubMed

    Pérez-Bravo, F; Fuentes, M; Angel, B; Sanchez, H; Carrasco, E; Santos, J L; Lera, L; Albala, C

    2006-12-01

    The aim of this study was to assess the frequency of fatty acid binding protein 2 (FABP2) Ala54Thr genetic polymorphism and to evaluate its association with obesity and insulin resistance in Chilean aboriginal populations. A sample of 96 urban Aymara and 111 urban Mapuche subjects aged 20-80 years were recruited for this cross-sectional study. Glucose, insulin and lipid profile were measured in fasting plasma samples. Insulin resistance was estimated through the HOMA-IR model. FABP2 Ala54Thr genotypes were determined by PCR followed by RFLP analysis. The allele frequency of Thr54 variant was estimated as 18.2% in Aymara subjects, which is one of the lowest reported to date. The corresponding frequency in Mapuche subjects was 31.9% (p<0.002). Regarding genotype-phenotype associations, no significant differences were found in any of the anthropometric or metabolic variables according to Ala54Thr genotypes. After adjustment by BMI and metabolic variables through a logistic regression analysis, the association of the FABP2 polymorphism with ethnic group persisted (Mapuche group: OR=2.37, 95% CI 1.319-4.277, p=0.004) It is unlikely that Ala54Thr polymorphism of the FABP2 gene plays a relevant role in obesity and insulin resistance in Chilean ethnic groups. PMID:17211557

  8. Impact of clinical context on acute kidney injury biomarker performances: differences between neutrophil gelatinase-associated lipocalin and L-type fatty acid-binding protein

    PubMed Central

    Asada, Toshifumi; Isshiki, Rei; Hayase, Naoki; Sumida, Maki; Inokuchi, Ryota; Noiri, Eisei; Nangaku, Masaomi; Yahagi, Naoki; Doi, Kent

    2016-01-01

    Application of acute kidney injury (AKI) biomarkers with consideration of nonrenal conditions and systemic severity has not been sufficiently determined. Herein, urinary neutrophil gelatinase-associated lipocalin (NGAL), L-type fatty acid-binding protein (L-FABP) and nonrenal disorders, including inflammation, hypoperfusion and liver dysfunction, were evaluated in 249 critically ill patients treated at our intensive care unit. Distinct characteristics of NGAL and L-FABP were revealed using principal component analysis: NGAL showed linear correlations with inflammatory markers (white blood cell count and C-reactive protein), whereas L-FABP showed linear correlations with hypoperfusion and hepatic injury markers (lactate, liver transaminases and bilirubin). We thus developed a new algorithm by combining urinary NGAL and L-FABP with stratification by the Acute Physiology and Chronic Health Evaluation score, presence of sepsis and blood lactate levels to improve their AKI predictive performance, which showed a significantly better area under the receiver operating characteristic curve [AUC-ROC 0.940; 95% confidential interval (CI) 0.793–0.985] than that under NGAL alone (AUC-ROC 0.858, 95% CI 0.741–0.927, P = 0.03) or L-FABP alone (AUC-ROC 0.837, 95% CI 0.697–0.920, P = 0.007) and indicated that nonrenal conditions and systemic severity should be considered for improved AKI prediction by NGAL and L-FABP as biomarkers. PMID:27605390

  9. Impact of clinical context on acute kidney injury biomarker performances: differences between neutrophil gelatinase-associated lipocalin and L-type fatty acid-binding protein.

    PubMed

    Asada, Toshifumi; Isshiki, Rei; Hayase, Naoki; Sumida, Maki; Inokuchi, Ryota; Noiri, Eisei; Nangaku, Masaomi; Yahagi, Naoki; Doi, Kent

    2016-01-01

    Application of acute kidney injury (AKI) biomarkers with consideration of nonrenal conditions and systemic severity has not been sufficiently determined. Herein, urinary neutrophil gelatinase-associated lipocalin (NGAL), L-type fatty acid-binding protein (L-FABP) and nonrenal disorders, including inflammation, hypoperfusion and liver dysfunction, were evaluated in 249 critically ill patients treated at our intensive care unit. Distinct characteristics of NGAL and L-FABP were revealed using principal component analysis: NGAL showed linear correlations with inflammatory markers (white blood cell count and C-reactive protein), whereas L-FABP showed linear correlations with hypoperfusion and hepatic injury markers (lactate, liver transaminases and bilirubin). We thus developed a new algorithm by combining urinary NGAL and L-FABP with stratification by the Acute Physiology and Chronic Health Evaluation score, presence of sepsis and blood lactate levels to improve their AKI predictive performance, which showed a significantly better area under the receiver operating characteristic curve [AUC-ROC 0.940; 95% confidential interval (CI) 0.793-0.985] than that under NGAL alone (AUC-ROC 0.858, 95% CI 0.741-0.927, P = 0.03) or L-FABP alone (AUC-ROC 0.837, 95% CI 0.697-0.920, P = 0.007) and indicated that nonrenal conditions and systemic severity should be considered for improved AKI prediction by NGAL and L-FABP as biomarkers. PMID:27605390

  10. Heart-type fatty acid-binding protein and its relation with morphological changes in rat myocardial damage model induced by isoproterenol.

    PubMed

    Hasić, Sabaheta; Jadrić, Radivoj; Cosović, Esad; Kiseljaković, Emina; Mornjaković, Zakira; Winterhalter-Jadrić, Mira

    2011-11-01

    We have investigated heart type fatty acid binding protein (H-FABP) rat serum values at different time point following subcutaneous (s.c) isoproterenol (ISO) administration and their correlation with severity of myocardial lesion. Thirty adult, male, Wistar rats were used for this study. Six rats per group were treated with a single dose of either ISO (ISO groups, dose 100 mg/kg, s.c.) at different time point (30', 60', 120', 240') or with saline (control group). Serum H-FABP was determined by enzyme-linked immunosorbent assay (ELISA) and histological analysis was performed by hematoxylin-eosin (HE) method of staining. The first serum H-FABP increase was obtained 30' following ISO administration, but maximal value was reached after 240'. Myocardial histological changes were time-dependent and correlated with serum H-FABP values (p<0.001). The results of the study suggest that H-FABP is sensitive marker for acute rat myocardial injury and its possible inclusion in myocardial injury screening studies in rats. PMID:22117831

  11. Locating high-affinity fatty acid-binding sites on albumin by x-ray crystallography and NMR spectroscopy

    PubMed Central

    Simard, J. R.; Zunszain, P. A.; Ha, C.-E.; Yang, J. S.; Bhagavan, N. V.; Petitpas, I.; Curry, S.; Hamilton, J. A.

    2005-01-01

    Human serum albumin (HSA) is a versatile transport protein for endogenous compounds and drugs. To evaluate physiologically relevant interactions between ligands for the protein, it is necessary to determine the locations and relative affinities of different ligands for their binding site(s). We present a site-specific investigation of the relative affinities of binding sites on HSA for fatty acids (FA), the primary physiological ligand for the protein. Titration of HSA with [13C]carboxyl-labeled FA was used initially to identify three NMR chemical shifts that are associated with high-affinity binding pockets on the protein. To correlate these peaks with FA-binding sites identified from the crystal structures of FA–HSA complexes, HSA mutants were engineered with substitutions of amino acids involved in coordination of the bound FA carboxyl. Titration of [13C]palmitate into solutions of HSA mutants for either FA site four (R410A/Y411A) or site five (K525A) within domain III of HSA each revealed loss of a specific NMR peak that was present in spectra of wild-type protein. Because these peaks are among the first three to be observed on titration of HSA with palmitate, sites four and five represent two of the three high-affinity long-chain FA-binding sites on HSA. These assignments were confirmed by titration of [13C]palmitate into recombinant domain III of HSA, which contains only sites four and five. These results establish a protocol for direct probing of the relative affinities of FA-binding sites, one that may be extended to examine competition between FA and other ligands for specific binding sites. PMID:16330771

  12. Locating high-affinity fatty acid-binding sites on albumin by x-ray crystallography and NMR spectroscopy.

    PubMed

    Simard, J R; Zunszain, P A; Ha, C-E; Yang, J S; Bhagavan, N V; Petitpas, I; Curry, S; Hamilton, J A

    2005-12-13

    Human serum albumin (HSA) is a versatile transport protein for endogenous compounds and drugs. To evaluate physiologically relevant interactions between ligands for the protein, it is necessary to determine the locations and relative affinities of different ligands for their binding site(s). We present a site-specific investigation of the relative affinities of binding sites on HSA for fatty acids (FA), the primary physiological ligand for the protein. Titration of HSA with [(13)C]carboxyl-labeled FA was used initially to identify three NMR chemical shifts that are associated with high-affinity binding pockets on the protein. To correlate these peaks with FA-binding sites identified from the crystal structures of FA-HSA complexes, HSA mutants were engineered with substitutions of amino acids involved in coordination of the bound FA carboxyl. Titration of [(13)C]palmitate into solutions of HSA mutants for either FA site four (R410A/Y411A) or site five (K525A) within domain III of HSA each revealed loss of a specific NMR peak that was present in spectra of wild-type protein. Because these peaks are among the first three to be observed on titration of HSA with palmitate, sites four and five represent two of the three high-affinity long-chain FA-binding sites on HSA. These assignments were confirmed by titration of [(13)C]palmitate into recombinant domain III of HSA, which contains only sites four and five. These results establish a protocol for direct probing of the relative affinities of FA-binding sites, one that may be extended to examine competition between FA and other ligands for specific binding sites. PMID:16330771

  13. Influence of liposomes rich in unsaturated or saturated fatty acids on the growth of human xenotransplanted mammary carcinomas and on the levels of heart type fatty acid binding protein.

    PubMed

    Naundorf, H; Zschiesche, W; Reszka, R; Fichtner, I

    1995-01-01

    A panel of 4 human mammary carcinomas passaged in nude mice were subjected to intraperitoneal application of cholesterol-free liposomes enriched with linoleic (unsaturated fatty acid) or stearic acid (saturated fatty acid). The liposomes were examined with regard to their influence on the tumor growth and level of heart type fatty acid binding protein (FABP). Liposomes with different fatty acid composition influenced the growth of mammary carcinomas 3366, BO, 4000 and 4151 in distinct ways. Liposomes with a high content of stearic acid significantly inhibited the growth of mammary carcinomas 3366 and BO, whereas mammary carcinomas 4000 and 4151 were not affected. The growth of mammary carcinoma 3366 was moderately increased after supplementation of liposomes rich in linoleic acid, the tumor BO was significantly inhibited and the growth of MaCa 4000 and 4151 was unchanged. Liposome treatment led to a significant increase in heart type FABP in mammary carcinomas 3366 and BO regardless of whether the animals were treated with liposomes rich in stearic or linoleic acid. Such significant changes of FABP level could not be observed in mammary carcinomas 4000 or 4151. We suggest that the lipid-mediated growth modulation seems to be dependent on an increase of heart type FABPs in these tumor models. PMID:8562891

  14. Serum Level of Heart-Type Fatty Acid Binding Protein (H-FABP) Before and After Treatment of Congestive Heart Failure in Children.

    PubMed

    Zoair, Amr; Mawlana, Wegdan; Abo-Elenin, Amany; Korrat, Mostafa

    2015-12-01

    Remodeling of the heart following injury affects the morbidity and mortality in children presented with heart failure (HF). Heart-type fatty acid binding protein (H-FABP) is a novel biomarker that could be of help to predict the prognosis and risk stratification in those children. We aimed to evaluate the diagnostic and prognostic value of H-FABP in children with heart failure before and after treatment. The study was conducted as a prospective cohort study. It included 30 children with HF as a patient group and 20 healthy children matched for age and sex as a control group. Echocardiographic assessment of the heart was done using conventional Doppler echocardiography. Serum levels of (H-FABP) were measured using enzyme-linked immunosorbent assay before and after treatment of HF. All patients were observed during follow-up period of 3 months. There was a significant difference in the serum level of H-FABP in our patients before treatment (5.278 ± 3.253 ng/ml) compared with after treatment (2.089 ± 0.160 ng/ml) with significant difference compared with the control group. There was a significant increase in the serum level of H-FABP with increase in the severity of heart failure according to Ross classification. Significant increase in the H-FABP was associated with adverse outcome. Serum levels of H-FABP strongly correlated with clinical and echocardiographic assessment of LV performance of children with HF, and its levels significantly increased in children with adverse outcome suggesting its value as a useful diagnostic and prognostic predictor (with high sensitivity and specificity). PMID:26123812

  15. Fatty acid-binding protein 5 regulates diet-induced obesity via GIP secretion from enteroendocrine K cells in response to fat ingestion.

    PubMed

    Shibue, Kimitaka; Yamane, Shunsuke; Harada, Norio; Hamasaki, Akihiro; Suzuki, Kazuyo; Joo, Erina; Iwasaki, Kanako; Nasteska, Daniela; Harada, Takanari; Hayashi, Yoshitaka; Adachi, Yasuhiro; Owada, Yuji; Takayanagi, Ryoichi; Inagaki, Nobuya

    2015-04-01

    Gastric inhibitory polypeptide (GIP) is an incretin released from enteroendocrine K cells in response to nutrient intake, especially fat. GIP is one of the contributing factors inducing fat accumulation that results in obesity. A recent study shows that fatty acid-binding protein 5 (FABP5) is expressed in murine K cells and is involved in fat-induced GIP secretion. We investigated the mechanism of fat-induced GIP secretion and the impact of FABP5-related GIP response on diet-induced obesity (DIO). Single oral administration of glucose and fat resulted in a 40% reduction of GIP response to fat but not to glucose in whole body FABP5-knockout (FABP5(-/-)) mice, with no change in K cell count or GIP content in K cells. In an ex vivo experiment using isolated upper small intestine, oleic acid induced only a slight increase in GIP release, which was markedly enhanced by coadministration of bile and oleic acid together with attenuated GIP response in the FABP5(-/-) sample. FABP5(-/-) mice exhibited a 24% reduction in body weight gain and body fat mass under a high-fat diet compared with wild-type (FABP5(+/+)) mice; the difference was not observed between GIP-GFP homozygous knock-in (GIP(gfp/gfp))-FABP5(+/+) mice and GIP(gfp/gfp)-FABP5(-/-) mice, in which GIP is genetically deleted. These results demonstrate that bile efficiently amplifies fat-induced GIP secretion and that FABP5 contributes to the development of DIO in a GIP-dependent manner. PMID:25628425

  16. Cytoprotective role of the fatty acid binding protein 4 against oxidative and endoplasmic reticulum stress in 3T3-L1 adipocytes.

    PubMed

    Kajimoto, Kazuaki; Minami, Yoshitaka; Harashima, Hideyoshi

    2014-01-01

    The fatty acid binding protein 4 (FABP4), one of the most abundant proteins in adipocytes, has been reported to have a proinflammatory function in macrophages. However, the physiological role of FABP4, which is constitutively expressed in adipocytes, has not been fully elucidated. Previously, we demonstrated that FABP4 was involved in the regulation of interleukin-6 (IL-6) and vascular endothelial growth factor (VEGF) production in 3T3-L1 adipocytes. In this study, we examined the effects of FABP4 silencing on the oxidative and endoplasmic reticulum (ER) stress in 3T3-L1 adipocytes. We found that the cellular reactive oxygen species (ROS) and 8-nitro-cyclic GMP levels were significantly elevated in the differentiated 3T3-L1 adipocytes transfected with a small interfering RNA (siRNA) against Fabp4, although the intracellular levels or enzyme activities of antioxidants including reduced glutathione (GSH), superoxide dismutase (SOD) and glutathione S-transferase A4 (GSTA4) were not altered. An in vitro evaluation using the recombinant protein revealed that FABP4 itself functions as a scavenger protein against hydrogen peroxide (H2O2). FABP4-knockdown resulted in a significant lowering of cell viability of 3T3-L1 adipocytes against H2O2 treatment. Moreover, four kinds of markers related to the ER stress response including the endoplasmic reticulum to nucleus signaling 1 (Ern1), the signal sequence receptor α (Ssr1), the ORM1-like 3 (Ormdl3), and the spliced X-box binding protein 1 (Xbp1s), were all elevated as the result of the knockdown of FABP4. Consequently, FABP4 might have a new role as an antioxidant protein against H2O2 and contribute to cytoprotection against oxidative and ER stress in adipocytes. PMID:25161868

  17. Correlation between Heart-type Fatty Acid-binding Protein Gene Polymorphism and mRNA Expression with Intramuscular Fat in Baicheng-oil Chicken.

    PubMed

    Wang, Yong; He, Jianzhong; Yang, Wenxuan; Muhantay, Gemenggul; Chen, Ying; Xing, Jinming; Liu, Jianzhu

    2015-10-01

    This study aims to determine the polymorphism and mRNA expression pattern of the heart-type fatty acid-binding protein (H-FABP) gene and their association with intramuscular fat (IMF) content in the breast and leg muscles of Baicheng oil chicken (BOC). A total of 720 chickens, including 240 black Baicheng oil chicken (BBOC), 240 silky Baicheng oil chicken (SBOC), and 240 white Baicheng oil chicken (WBOC) were raised. Three genotypes of H-FABP gene second extron following AA, AB, and BB were detected by polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP) strategy. The G939A site created AA genotype and G956A site created BB genotype. The content of IMF in AA genotype in breast muscle of BBOC was significantly higher than that of AB (p = 0.0176) and the genotype in leg muscle of WBOC was significantly higher than that of AB (p = 0.0145). The G939A site could be taken as genetic marker for higher IMF content selecting for breast muscle of BBOC and leg muscle of WBOC. The relative mRNA expression of H-FABP was measured by real-time PCR at 30, 60, 90, and 120 d. The IMF content significantly increased with age in both muscles. The mRNA expression level of H-FABP significantly decreased with age in both muscles of the three types of chickens. Moreover, a significant negative correlation between H-FABP abundance and IMF content in the leg muscles of WBOC (p = 0.035) was observed. The mRNA expression of H-FABP negatively correlated with the IMF content in both breast and leg muscles of BOC sat slaughter time. PMID:26323394

  18. Heart-type Fatty acid-binding protein in Acute Myocardial infarction Evaluation (FAME): Background and design of a diagnostic study in primary care

    PubMed Central

    Bruins Slot, Madeleine HE; van der Heijden, Geert JMG; Rutten, Frans H; van der Spoel, Onno P; Mast, E Gijs; Bredero, Ad C; Doevendans, Pieter A; Glatz, Jan FC; Hoes, Arno W

    2008-01-01

    Background Currently used biomarkers for cardiac ischemia are elevated in blood plasma after a delay of several hours and therefore unable to detect acute coronary syndrome (ACS) in a very early stage. General practitioners (GPs), however, are often confronted with patients suspected of ACS within hours after onset of complaints. This ongoing study aims to evaluate the added diagnostic value beyond clinical assessment for a rapid bedside test for heart-type fatty-acid binding protein (H-FABP), a biomarker that is detectable as soon as one hour after onset of ischemia. Methods Participating GPs perform a blinded H-FABP rapid bedside test (Cardiodetect®) in patients with symptoms suggestive of ACS such as chest pain or discomfort at rest. All patients, whether referred to hospital or not, undergo electrocardiography (ECG) and venapunction for a plasma troponin test within 12–36 hours after onset of complaints. A final diagnosis will be established by an expert panel consisting of two cardiologists and one general practitioner (blinded to the H-FABP test result), using all available patient information, also including signs and symptoms. The added diagnostic value of the H-FABP test beyond history taking and physical examination will be determined with receiver operating characteristic curves derived from multivariate regression analysis. Conclusion Reasons for presenting the design of our study include the prevention of publication bias and unacknowledged alterations in the study aim, design or data-analysis. To our knowledge this study is the first to assess the diagnostic value of H-FABP outside a hospital-setting. Several previous hospital-based studies showed the potential value of H-FABP in diagnosing ACS. Up to now however it is unclear whether these results are equally promising when the test is used in primary care. The first results are expected in the end of 2008. PMID:18412949

  19. Cytoprotective role of the fatty acid binding protein 4 against oxidative and endoplasmic reticulum stress in 3T3-L1 adipocytes

    PubMed Central

    Kajimoto, Kazuaki; Minami, Yoshitaka; Harashima, Hideyoshi

    2014-01-01

    The fatty acid binding protein 4 (FABP4), one of the most abundant proteins in adipocytes, has been reported to have a proinflammatory function in macrophages. However, the physiological role of FABP4, which is constitutively expressed in adipocytes, has not been fully elucidated. Previously, we demonstrated that FABP4 was involved in the regulation of interleukin-6 (IL-6) and vascular endothelial growth factor (VEGF) production in 3T3-L1 adipocytes. In this study, we examined the effects of FABP4 silencing on the oxidative and endoplasmic reticulum (ER) stress in 3T3-L1 adipocytes. We found that the cellular reactive oxygen species (ROS) and 8-nitro-cyclic GMP levels were significantly elevated in the differentiated 3T3-L1 adipocytes transfected with a small interfering RNA (siRNA) against Fabp4, although the intracellular levels or enzyme activities of antioxidants including reduced glutathione (GSH), superoxide dismutase (SOD) and glutathione S-transferase A4 (GSTA4) were not altered. An in vitro evaluation using the recombinant protein revealed that FABP4 itself functions as a scavenger protein against hydrogen peroxide (H2O2). FABP4-knockdown resulted in a significant lowering of cell viability of 3T3-L1 adipocytes against H2O2 treatment. Moreover, four kinds of markers related to the ER stress response including the endoplasmic reticulum to nucleus signaling 1 (Ern1), the signal sequence receptor α (Ssr1), the ORM1-like 3 (Ormdl3), and the spliced X-box binding protein 1 (Xbp1s), were all elevated as the result of the knockdown of FABP4. Consequently, FABP4 might have a new role as an antioxidant protein against H2O2 and contribute to cytoprotection against oxidative and ER stress in adipocytes. PMID:25161868

  20. Fatty acid binding proteins 4 and 5 in overweight prepubertal boys: effect of nutritional counselling and supplementation with an encapsulated fruit and vegetable juice concentrate.

    PubMed

    Canas, Jose A; Damaso, L; Hossain, J; Balagopal, P Babu

    2015-01-01

    Elevated fatty acid binding proteins (FABP) may play a role in obesity and co-morbidities. The role of nutritional interventions in modulating these levels remains unclear. The aim of this post hoc study was to determine the effect of overweight (OW) on FABP4 and FABP5 in boys in relation to indices of adiposity, insulin resistance and inflammation, and to investigate the effects of a 6-month supplementation with an encapsulated fruit and vegetable juice concentrate (FVJC) plus nutritional counselling (NC) on FABP levels. A post hoc analysis of a double-blind, randomised, placebo-controlled study of children recruited from the general paediatric population was performed. A total of thirty age-matched prepubertal boys (nine lean and twenty-one OW; aged 6-10 years) were studied. Patients received NC by a registered dietitian and were randomised to FVJC or placebo capsules for 6 months. FABP4, FABP5, glucose, insulin, homeostasis model assessment-insulin resistance (HOMA-IR), glucose-induced acute insulin response (AIR), lipid-corrected β-carotene (LCβC), adiponectin, leptin, high-sensitivity C-reactive protein (hs-CRP), IL-6 and body composition by dual-energy X-ray absorptiometry were determined before and after the intervention. FABP were higher (P < 0·01) in the OW v. lean boys and correlated directly with HOMA-IR, abdominal fat mass (AFM), hs-CRP, IL-6, and LCβC (P < 0·05 for all). FABP4 was associated with adiponectin and AIR (P < 0·05). FVJC plus NC reduced FABP4, HOMA-IR and AFM (P < 0·05 for all) but not FABP5. OW boys showed elevated FABP4 and FABP5, but only FABP4 was lowered by the FVJC supplement. PMID:26688725

  1. Fatty Acid Binding Protein 7 Is a Molecular Marker in Adenoid Cystic Carcinoma of the Salivary Glands: Implications for Clinical Significance12

    PubMed Central

    Phuchareon, Janyaporn; Overdevest, Jonathan B.; McCormick, Frank; Eisele, David W.; van Zante, Annemieke; Tetsu, Osamu

    2014-01-01

    Adenoid cystic carcinoma (ACC) is an aggressive malignant neoplasm of the salivary glands. Its diagnosis is difficult due to overlapping features with other salivary tumors. Gene expression analysis may complement traditional diagnostic methods. We searched gene expression patterns in the Gene Expression Omnibus (GEO) database and in our tumor and normal samples. The biologic and prognostic potential of the identified genes was analyzed. The GEO data set of primary xenografted ACCs revealed that expression of five genes, engrailed homeobox 1 (EN1), fatty acid binding protein 7 (FABP7), hemoglobin epsilon 1, MYB, and versican (VCAN), was dramatically increased. mRNA expression of EN1, FABP7, MYB, and VCAN distinguished our sporadic ACCs from normal tissues and benign tumors. FABP7 expression appeared to be regulated differently from EN1 and MYB and was crossly correlated with poor prognosis in our ACC cohort. Immunohistochemistry showed that FABP7 protein was predominantly expressed in the nucleus of myoepithelial cells of both tubular and cribriform subtypes. In contrast, in the solid subtype, which is often associated with a lower survival rate, FABP7 protein was uniformly expressed in cancerous cells. One case with cribriform architecture and the highest level of FABP7 mRNA showed strong FABP7 staining in both duct-type epithelial and myoepithelial cells, suggesting that diffuse expression of FABP7 protein might be related to aggressive tumor behavior and poor prognosis. We propose FABP7 as a novel biomarker in ACC. The molecule may be useful in diagnosis and for identifying more effective therapies targeting this protein or upstream molecules that regulate it. PMID:25500088

  2. Crystal Structure of the Mp1p Ligand Binding Domain 2 Reveals Its Function as a Fatty Acid-binding Protein*

    PubMed Central

    Liao, Shuang; Tung, Edward T. K.; Zheng, Wei; Chong, Ken; Xu, Yuanyuan; Dai, Peng; Guo, Yingying; Bartlam, Mark; Yuen, Kwok-Yung; Rao, Zihe

    2010-01-01

    Penicillium marneffei is a dimorphic, pathogenic fungus in Southeast Asia that mostly afflicts immunocompromised individuals. As the only dimorphic member of the genus, it goes through a phase transition from a mold to yeast form, which is believed to be a requisite for its pathogenicity. Mp1p, a cell wall antigenic mannoprotein existing widely in yeast, hyphae, and conidia of the fungus, plays a vital role in host immune response during infection. To understand the function of Mp1p, we have determined the x-ray crystal structure of its ligand binding domain 2 (LBD2) to 1.3 Å. The structure reveals a dimer between the two molecules. The dimer interface forms a ligand binding cavity, in which electron density was observed for a palmitic acid molecule interacting with LBD2 indirectly through hydrogen bonding networks via two structural water molecules. Isothermal titration calorimetry experiments measured the ligand binding affinity (Kd) of Mp1p at the micromolar level. Mutations of ligand-binding residues, namely S313A and S332A, resulted in a 9-fold suppression of ligand binding affinity. Analytical ultracentrifugation assays demonstrated that both LBD2 and Mp1p are mostly monomeric in vitro, no matter with or without ligand, and our dimeric crystal structure of LBD2 might be the result of crystal packing. Based on the conformation of the ligand-binding pocket in the dimer structure, a model for the closed, monomeric form of LBD2 is proposed. Further structural analysis indicated the biological importance of fatty acid binding of Mp1p for the survival and pathogenicity of the conditional pathogen. PMID:20053994

  3. Fatty Acid-binding Proteins 1 and 2 Differentially Modulate the Activation of Peroxisome Proliferator-activated Receptor α in a Ligand-selective Manner.

    PubMed

    Hughes, Maria L R; Liu, Bonan; Halls, Michelle L; Wagstaff, Kylie M; Patil, Rahul; Velkov, Tony; Jans, David A; Bunnett, Nigel W; Scanlon, Martin J; Porter, Christopher J H

    2015-05-29

    Nuclear hormone receptors (NHRs) regulate the expression of proteins that control aspects of reproduction, development and metabolism, and are major therapeutic targets. However, NHRs are ubiquitous and participate in multiple physiological processes. Drugs that act at NHRs are therefore commonly restricted by toxicity, often at nontarget organs. For endogenous NHR ligands, intracellular lipid-binding proteins, including the fatty acid-binding proteins (FABPs), can chaperone ligands to the nucleus and promote NHR activation. Drugs also bind FABPs, raising the possibility that FABPs similarly regulate drug activity at the NHRs. Here, we investigate the ability of FABP1 and FABP2 (intracellular lipid-binding proteins that are highly expressed in tissues involved in lipid metabolism, including the liver and intestine) to influence drug-mediated activation of the lipid regulator peroxisome proliferator-activated receptor (PPAR) α. We show by quantitative fluorescence imaging and gene reporter assays that drug binding to FABP1 and FABP2 promotes nuclear localization and PPARα activation in a drug- and FABP-dependent manner. We further show that nuclear accumulation of FABP1 and FABP2 is dependent on the presence of PPARα. Nuclear accumulation of FABP on drug binding is driven largely by reduced nuclear egress rather than an increased rate of nuclear entry. Importin binding assays indicate that nuclear access occurs via an importin-independent mechanism. Together, the data suggest that specific drug-FABP complexes can interact with PPARα to effect nuclear accumulation of FABP and NHR activation. Because FABPs are expressed in a regionally selective manner, this may provide a means to tailor the patterns of NHR drug activation in a tissue-specific manner. PMID:25847235

  4. Fasciola hepatica fatty acid binding protein inhibits TLR4 activation and suppresses the inflammatory cytokines induced by LPS in vitro and in vivo

    PubMed Central

    Martin, Ivelisse; Cabán-Hernández, Kimberly; Figueroa-Santiago, Olgary; Espino, Ana M.

    2015-01-01

    Toll-like receptor 4 (TLR4), the innate immunity receptor for bacterial endotoxins, plays a pivotal role in the induction of inflammatory responses. There is a need to develop molecules that block either activation through TLR4 or the downstream signaling pathways to inhibit the storm of inflammation typically elicited by bacterial lipopolysaccharide (LPS), which is a major cause of the high mortality associated with bacterial sepsis. We report here that a single intraperitoneal injection of 15μg Fasciola hepatica fatty acid binding protein (Fh12) 1 hour before exposure to LPS suppressed significantly the expression of serum inflammatory cytokines in a model of septic shock using C57BL/6 mice. Because macrophages are good source of IL12p70 and TNFα, and critical in driving adaptive immunity, we investigated the effect of Fh12 on the function of mouse bone marrow derived macrophages (bmMΦs). Whereas Fh12 alone did not induce cytokine expression, it significantly suppressed the expression of IL12, TNFα, IL6 and IL1β cytokines as well as iNOS2 in bmMΦs, and also impaired the phagocytic capacity of bmMΦs. Fh12 had a limited effect on the expression of inflammatory cytokines induced in response to other TLR-ligands. One mechanism used by Fh12 to exert its anti-inflammatory effect is binding to the CD14 co-receptor. Moreover, it suppresses phosphorylation of ERK, p38 and JNK. The potent anti-inflammatory properties of Fh12 demonstrated here open doors to further studies directed at exploring the potential of this molecule as a new class of drug against septic shock or other inflammatory diseases. PMID:25780044

  5. Heart-Type Fatty Acid-Binding Protein, in Early Detection of Acute Myocardial Infarction: Comparison with CK-MB, Troponin I and Myoglobin.

    PubMed

    Pyati, Anand K; Devaranavadagi, Basavaraj B; Sajjannar, Sanjeev L; Nikam, Shashikant V; Shannawaz, Mohd; Patil, Satish

    2016-10-01

    The study aimed to investigate whether heart-type fatty acid binding protein (H-FABP) measurement provides additional diagnostic value to that of conventional cardiac markers in acute myocardial infarction (AMI) within first 6 h after the onset of symptoms. The study included 120 subjects: 60 AMI cases and 60 age and sex matched controls. The cases and controls were further divided into 2 subgroups depending on the time since onset of chest pain as (1) subjects within 3 h and (2) between 3 and 6 h of onset of chest pain. In all the cases and controls, serum H-FABP concentration was measured by Immunoturbidimetric method, serum Troponin I and myoglobin concentrations by Chemiluminescence immunoassay and serum CK-MB concentration by Immuno-inhibition method. The sensitivity, specificity, positive and negative predictive values of H-FABP were significantly greater than CK-MB and myoglobin but were lesser than Troponin I in patients with suspected AMI in both within 3 h and 3-6 h groups. Receiver operating characteristic curves demonstrated greatest diagnostic ability for Troponin I (AUC = 0.99, p < 0.001) followed by H-FABP (AUC = 0.906, p < 0.001) within 3 h and 3-6 h after the onset of chest pain. In conclusion, the diagnostic value of H-FABP is greater than CK-MB and myoglobin but slightly lesser than troponin I for the early diagnosis of AMI within first 6 h of chest pain. H-FABP can be used as an additional diagnostic tool for the early diagnosis of AMI along with troponin I. PMID:27605741

  6. Urine Liver-Type Fatty Acid-Binding Protein and Kidney Injury Molecule-1 in HIV-Infected Patients Receiving Combined Antiretroviral Treatment Based on Tenofovir

    PubMed Central

    Wójcik, Kamila; Piekarska, Anna

    2014-01-01

    Abstract The aim of this study was to determine the presence of kidney tubular damage in the absence of overt evidence of glomerular dysfunction (GFR>60 ml/min without proteinuria) in HIV-infected patients receiving antiretroviral therapy. Urine kidney injury molecule-1 (KIM-1) and liver-type fatty acid-binding protein (L-FABP) levels were measured by ELISA and expressed as a ratio to creatinine. Sixty-six patients (median age 38 years) and 10 healthy controls (median age 35.5 years) were included in the study. Patients with chronic diseases such as diabetes, hypertension, heart disease, or kidney disease were excluded from the study. All patients received tenofovir/emtricitabine combined with one of three other components, namely efavirenz, atazanavir/norvir, or lopinavir/norvir. A lower concentration of L-FABP/creatinine was observed in HIV-infected as compared to healthy individuals (p=0.0353); KIM-1/creatinine was also lower in comparison with healthy controls but not statistically significantly. Patients receiving efavirenz had higher levels of L-FABP/creatinine in comparison to healthy controls (p=0.0039). Patients with anti-HCV had higher concentrations of L-FABP/creatinine as compared to the HIV-monoinfected individuals (not statistically significant) and to healthy subjects (p=0.0356). All four patients with L-FABP>17.5 μg/g creatinine were HIV/HCV coinfected. On multivariate logistic regression urine L-FABP above 5.5 μg/g creatinine was independently associated with body weight (OR=0.93 p=0.039). This study suggests that HIV/HCV-coinfected patients with lower body weight treated with tenofovir may be at an increased risk of tubular dysfunction and should be monitored more closely. The use of protease inhibitors was not associated with an increased risk of tubular disorders. PMID:24164392

  7. Development of an ultra-rapid diagnostic method based on heart-type fatty acid binding protein levels in the CSF of CJD patients.

    PubMed

    Matsui, Yuki; Satoh, Katsuya; Mutsukura, Kazuo; Watanabe, Takuya; Nishida, Noriyuki; Matsuda, Hideo; Sugino, Masaichi; Shirabe, Susumu; Eguchi, Katsumi; Kataoka, Yasufumi

    2010-10-01

    Creutzfeldt-Jakob disease (CJD) is a transmissible, fatal, neurodegenerative disease in humans. Recently, various drugs have been reported to be useful in the treatment of CJD; however, for such treatments to be useful it is essential to rapidly and accurately diagnose CJD. 124 CJD patients and 87 with other diseases causing rapid progressive dementia were examined. Cerebral spinal fluid (CSF) from CJD patients was analyzed by 2D-PAGE and the protein expression pattern was compared with that from healthy subjects. One of three CJD-specific spots was found to be fatty acid binding protein (FABP), and heart-type FABP (H-FABP) was analyzed as a new biochemical marker for CJD. H-FABP ELISA results were compared between CJD patients and patients with other diseases (n = 211). Visual readout accuracy of the Rapicheck(®) H-FABP test panel for CSF was analyzed using an independent measure of CSF H-FABP concentration. The distribution of H-FABP in the brains of CJD patients was examined by immunohistochemistry. ELISA sensitivity and specificity were 90.3% and 92.9%, respectively, and Rapicheck(®) H-FABP sensitivity and specificity were 87.9% and 96.0%, respectively. ELISA and Rapicheck(®) H-FABP assays provided comparable results for 14-3-3 protein and total tau protein. Elevated H-FABP levels were associated with an accumulation of abnormal prion protein, astrocytic gliosis, and neuronal loss in the cerebral cortices of CJD patients. In conclusion, Rapicheck(®) H-FABP of CSF specimens enabled quick and frequent diagnosis of CJD. H-FABP represents a new biomarker for CJD distinct from 14-3-3 protein and total tau protein. PMID:20499272

  8. Association of Heart-Type Fatty Acid-Binding Protein with Cardiovascular Risk Factors and All-Cause Mortality in the General Population: The Takahata Study

    PubMed Central

    Otaki, Yoichiro; Watanabe, Tetsu; Takahashi, Hiroki; Hirayama, Atushi; Narumi, Taro; Kadowaki, Shinpei; Honda, Yuki; Arimoto, Takanori; Shishido, Tetsuro; Miyamoto, Takuya; Konta, Tsuneo; Shibata, Yoko; Fukao, Akira; Daimon, Makoto; Ueno, Yoshiyuki; Kato, Takeo; Kayama, Takamasa; Kubota, Isao

    2014-01-01

    Background Despite many recent advances in medicine, preventing the development of cardiovascular diseases remains a challenge. Heart-type fatty acid-binding protein (H-FABP) is a marker of ongoing myocardial damage and has been reported to be a useful indicator for future cardiovascular events. However, it remains to be determined whether H-FABP can predict all-cause and cardiovascular deaths in the general population. Methods and Results This longitudinal cohort study included 3,503 subjects who participated in a community-based health checkup with a 7-year follow-up. Serum H-FABP was measured in registered subjects. The results demonstrated that higher H-FABP levels were associated with increasing numbers of cardiovascular risk factors, including hypertension, diabetes mellitus, obesity, and metabolic syndrome. There were 158 deaths during the follow-up period, including 50 cardiovascular deaths. Deceased subjects had higher H-FABP levels compared to surviving subjects. Multivariate Cox proportional hazard regression analysis revealed that H-FABP is an independent predictor of all-cause and cardiovascular deaths after adjustments for confounding factors. Subjects were divided into four quartiles according to H-FABP level, and Kaplan-Meier analysis demonstrated that the highest H-FABP quartile was associated with the greatest risks for all-cause and cardiovascular deaths. Net reclassification index and integrated discrimination index were significantly increased by addition of H-FABP to cardiovascular risk factors. Conclusions H-FABP level was increased in association with greater numbers of cardiovascular risk factors and was an independent risk factor for all-cause and cardiovascular deaths. H-FABP could be a useful indicator for the early identification of high-risk subjects in the general population. PMID:24847804

  9. Correlation between Heart-type Fatty Acid-binding Protein Gene Polymorphism and mRNA Expression with Intramuscular Fat in Baicheng-oil Chicken

    PubMed Central

    Wang, Yong; He, Jianzhong; Yang, Wenxuan; Muhantay, Gemenggul; Chen, Ying; Xing, Jinming; Liu, Jianzhu

    2015-01-01

    This study aims to determine the polymorphism and mRNA expression pattern of the heart-type fatty acid-binding protein (H-FABP) gene and their association with intramuscular fat (IMF) content in the breast and leg muscles of Baicheng oil chicken (BOC). A total of 720 chickens, including 240 black Baicheng oil chicken (BBOC), 240 silky Baicheng oil chicken (SBOC), and 240 white Baicheng oil chicken (WBOC) were raised. Three genotypes of H-FABP gene second extron following AA, AB, and BB were detected by polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP) strategy. The G939A site created AA genotype and G956A site created BB genotype. The content of IMF in AA genotype in breast muscle of BBOC was significantly higher than that of AB (p = 0.0176) and the genotype in leg muscle of WBOC was significantly higher than that of AB (p = 0.0145). The G939A site could be taken as genetic marker for higher IMF content selecting for breast muscle of BBOC and leg muscle of WBOC. The relative mRNA expression of H-FABP was measured by real-time PCR at 30, 60, 90, and 120 d. The IMF content significantly increased with age in both muscles. The mRNA expression level of H-FABP significantly decreased with age in both muscles of the three types of chickens. Moreover, a significant negative correlation between H-FABP abundance and IMF content in the leg muscles of WBOC (p = 0.035) was observed. The mRNA expression of H-FABP negatively correlated with the IMF content in both breast and leg muscles of BOC sat slaughter time. PMID:26323394

  10. Fatty acid binding protein 10 in the orange-spotted grouper (Epinephelus coioides): characterization and regulation under pH and temperature stress.

    PubMed

    Qi, Zeng-hua; Liu, Yu-feng; Wang, Wei-Na; Xin, Yu; Xie, Fu-xing; Wang, An-Li

    2012-04-01

    We have isolated and characterized a fatty acid binding protein from the liver of the orange-spotted grouper (Epinephelus coioides). Amino acid sequence similarity of the Ec-FABP (E. coioides-FABP) was highest to FABP10s isolated from the livers of catfish, chicken, salamander and iguana. The open-reading frame of the Ec-FABP codes for a protein of 14.0 kDa with a calculated isoelectric point of 8.5. We first expressed a FABP10 protein from orange-spotted grouper (E. coioides) in Pichia pastoris, and then characterized the antioxidative potential of our recombinant Ec-FABP by DCF fluorescence assay. RT-PCR assays showed that endogenous Ec-FABP mRNA is most strongly expressed in liver with the most abundance and intestine. Change in the groupers' blood cells respiratory burst activity was examined during and after exposure to the pH and temperature stress using flow cytometry. Further analysis of Ec-FABP gene expression in liver tissue by quantitative real-time PCR demonstrated that Ec-FABP transcript levels increased when the fish were exposed to both pH and temperature stress, but the time when its mRNA expression level peaked differed under these stresses. Western blot analyses confirmed that the Ec-FABP protein was strongly expressed in the liver after exposure to the pH and temperature stress. These results suggest that Ec-FABP expression is stimulated by pH and temperature stress and that it may play important roles in general adaptive and antioxidant responses. PMID:22182678

  11. Heart-Type Fatty Acid Binding Protein: A Better Cardiac Biomarker than CK-MB and Myoglobin in the Early Diagnosis of Acute Myocardial Infarction

    PubMed Central

    Devaranavadagi, Basavaraj B; Sajjannar, Sanjeev L; Nikam, Shashikant V; Shannawaz, Mohd; Sudharani

    2015-01-01

    Background Early diagnosis and therapeutic intervention can improve the outcome of acute myocardial infarction (AMI). However, there are no satisfactory cardiac biomarkers for the diagnosis of AMI within 6 hours of onset of symptoms. Among novel biochemical markers of AMI, heart-type fatty acid binding protein (H-FABP) is of particular interest. Aim To compare the diagnostic value of H-FABP with that of CK-MB and myoglobin in suspected AMI patients within first 6 hours after the onset of symptoms. Settings and Design The study includes 40 AMI cases and 40 non-cardiac chest pain otherwise healthy controls. The cases and controls were further divided into 2 groups depending on the time since chest pain as those subjects within 3 hours and those between 3-6 hours of onset of chest pain. Materials and Methods In all the cases and controls, serum H-FABP, CK-MB and myoglobin concentrations were measured by Immunoturbidimetric method, immuno-inhibition method and Chemiluminescence immunoassay respectively. Statistical Analysis Data is presented as mean ± SD values. Differences between means of two groups were assessed by Student t-test. Sensitivity, Specificity, Positive predictive value, Negative predictive values were calculated and ROC curve analysis was done to assess the diagnostic validity of each study parameter. Results The sensitivity, specificity, PPV, NPV of H-FABP were greater than CK-MB and myoglobin and ROC curve analysis demonstrated highest area under curve for H-FABP followed by myoglobin and CK-MB in patients with suspected AMI both within 3 hours and 3-6 hours after the onset of chest pain. Conclusion The diagnostic efficiency of H-FABP is greater than CK-MB and myoglobin for the early diagnosis of AMI within first 6 hours of chest pain. H-FABP can be used as an additional diagnostic tool for the early diagnosis of AMI. PMID:26557510

  12. The Ala54Thr Polymorphism of the Fatty Acid Binding Protein 2 Gene Modulates HDL Cholesterol in Mexican-Americans with Type 2 Diabetes

    PubMed Central

    Salto, Lorena M.; Bu, Liming; Beeson, W. Lawrence; Firek, Anthony; Cordero-MacIntyre, Zaida; De Leon, Marino

    2015-01-01

    The alanine to threonine amino acid substitution at codon 54 (Ala54Thr) of the intestinal fatty acid binding protein (FABP2) has been associated with elevated levels of insulin and blood glucose as well as with dyslipidemia. The aim of this study was to characterize the effect of this FABP2 polymorphism in Mexican-Americans with type 2 diabetes (T2D) in the context of a three-month intervention to determine if the polymorphism differentially modulates selected clinical outcomes. For this study, we genotyped 43 participant samples and performed post-hoc outcome analysis of the profile changes in fasting blood glucose, HbA1c, insulin, lipid panel and body composition, stratified by the Ala54Thr polymorphism. Our results show that the Thr54 allele carriers (those who were heterozygous or homozygous for the threonine-encoding allele) had lower HDL cholesterol and higher triglyceride levels at baseline compared to the Ala54 homozygotes (those who were homozygous for the alanine-encoding allele). Both groups made clinically important improvements in lipid profiles and glycemic control as a response to the intervention. Whereas the Ala54 homozygotes decreased HDL cholesterol in the context of an overall total cholesterol decrease, Thr54 allele carriers increased HDL cholesterol as part of an overall total cholesterol decrease. We conclude that the Ala54Thr polymorphism of FABP2 modulates HDL cholesterol in Mexican-Americans with T2D and that Thr54 allele carriers may be responsive in interventions that include dietary changes. PMID:26703680

  13. Fatty acids and hypolipidemic drugs regulate peroxisome proliferator-activated receptors α- and γ-mediated gene expression via liver fatty acid binding protein: A signaling path to the nucleus

    PubMed Central

    Wolfrum, Christian; Borrmann, Carola M.; Börchers, Torsten; Spener, Friedrich

    2001-01-01

    Peroxisome proliferator-activated receptor α (PPARα) is a key regulator of lipid homeostasis in hepatocytes and target for fatty acids and hypolipidemic drugs. How these signaling molecules reach the nuclear receptor is not known; however, similarities in ligand specificity suggest the liver fatty acid binding protein (L-FABP) as a possible candidate. In localization studies using laser-scanning microscopy, we show that L-FABP and PPARα colocalize in the nucleus of mouse primary hepatocytes. Furthermore, we demonstrate by pull-down assay and immunocoprecipitation that L-FABP interacts directly with PPARα. In a cell biological approach with the aid of a mammalian two-hybrid system, we provide evidence that L-FABP interacts with PPARα and PPARγ but not with PPARβ and retinoid X receptor-α by protein–protein contacts. In addition, we demonstrate that the observed interaction of both proteins is independent of ligand binding. Final and quantitative proof for L-FABP mediation was obtained in transactivation assays upon incubation of transiently and stably transfected HepG2 cells with saturated, monounsaturated, and polyunsaturated fatty acids as well as with hypolipidemic drugs. With all ligands applied, we observed strict correlation of PPARα and PPARγ transactivation with intracellular concentrations of L-FABP. This correlation constitutes a nucleus-directed signaling by fatty acids and hypolipidemic drugs where L-FABP acts as a cytosolic gateway for these PPARα and PPARγ agonists. Thus, L-FABP and the respective PPARs could serve as targets for nutrients and drugs to affect expression of PPAR-sensitive genes. PMID:11226238

  14. The Non-native Helical Intermediate State May Accumulate at Low pH in the Folding and Aggregation Landscape of the Intestinal Fatty Acid Binding Protein.

    PubMed

    Sarkar-Banerjee, Suparna; Chowdhury, Sourav; Paul, Simanta Sarani; Dutta, Debashis; Ghosh, Anisa; Chattopadhyay, Krishnananda

    2016-08-16

    There has been widespread interest in studying early intermediate states and their roles in protein folding. The interest in intermediate states has been further emphasized in the recent literature because of their implications for protein aggregation. Unfortunately, direct kinetic characterization of intermediates has been difficult because of the limited time resolutions offered by the kinetic techniques and the heterogeneity of the folding and aggregation landscape. Even in equilibrium experiments, the characterization of intermediate states could be difficult because (a) their populations in equilibrium could be low and/or (b) they lack any specific biochemical or biophysical signatures for their identification. In this paper, we have used fluorescence correlation spectroscopy to study the nature of a low-pH intermediate state of the intestinal fatty acid binding protein, a small protein with predominantly β-sheet structure. Our results have shown that the pH 3 intermediate diffuses faster than the folded protein and has strong helix forming propensity. These behaviors support Lim's hypothesis according to which even an entirely β-sheet protein would form helical bundles at the early stage. Using dynamic light scattering and thioflavin T binding measurements, we have observed that the pH 3 intermediate is prone to aggregation. We believe that early helix formation is the result of a local effect, which originates from the interaction of the neighboring amino acids around the hydrophobic core residues. This early intermediate reorganizes subsequently, and this structural reorganization is initiated by the destabilizing interactions induced by the distant residues, unfavorable entropic costs, and steric constraints of the hydrophobic side chains. Mutational analyses show further that the increase in the hydrophobicity in the hydrophobic core region increases the population of the α-helical intermediate, enhancing the aggregation propensity of the protein

  15. Fatty acid binding protein 3 (fabp3) is associated with insulin, lipids and cardiovascular phenotypes of the metabolic syndrome through epigenetic modifications in a northern european family population

    PubMed Central

    2013-01-01

    Background Fatty acid-binding proteins (FABPs) play regulatory roles at the nexus of lipid metabolism and signaling. Dyslipidemia in clinical manifestation frequently co-occurs with obesity, insulin resistance and hypertension in the Metabolic Syndrome (MetS). Animal studies have suggested FABPs play regulatory roles in expressing MetS phenotypes. In our family cohort of Northern European descent, transcript levels in peripheral white blood cells (PWBCs) of a key FABPs, FABP3, is correlated with the MetS leading components. However, evidence supporting the functions of FABPs in humans using genetic approaches has been scarce, suggesting FABPs may be under epigenetic regulation. The objective of this study was to test the hypothesis that CpG methylation status of a key regulator of lipid homeostasis, FABP3, is a quantitative trait associated with status of MetS phenotypes in humans. Methods We used a mass-spec based quantitative method, EpiTYPER®, to profile a CpG island that extends from the promoter to the first exon of the FABP3 gene in our family-based cohort of Northern European descent (n=517). We then conducted statistical analysis of the quantitative relationship of CpG methylation and MetS measures following the variance-component association model. Heritability of each methylation and the effect of age and sex on CpG methylation were also assessed in our families. Results We find that methylation levels of individual CpG units and the regional average are heritable and significantly influenced by age and sex. Regional methylation was strongly associated with plasma total cholesterol (p=0.00028) and suggestively associated with LDL-cholesterol (p=0.00495). Methylation at individual units was significantly associated with insulin sensitivity, lipid particle sizing and diastolic blood pressure (p<0.0028, corrected for multiple testing for each trait). Peripheral white blood cell (PWBC) expression of FABP3 in a separate group of subjects (n=128) negatively

  16. Inhibition of gene expression of carnitine palmitoyltransferase I and heart fatty acid binding protein in cyclophosphamide and ifosfamide-induced acute cardiotoxic rat models.

    PubMed

    Sayed-Ahmed, Mohamed M; Aldelemy, Meshan L; Al-Shabanah, Othman A; Hafez, Mohamed M; Al-Hosaini, Khaled A; Al-Harbi, Naif O; Al-Sharary, Shakir D; Al-Harbi, Mohamed M

    2014-09-01

    This study investigated whether cyclophosphamide (CP) and ifosfamide (IFO) therapy alters the expression of the key genes engaged in long-chain fatty acid (LCFA) oxidation outside rat heart mitochondria, and if so, whether these alterations should be viewed as a mechanism during CP- and IFO-induced cardiotoxicity. Adult male Wistar albino rats were assigned to one of the six treatment groups: Rats in group 1 (control) and group 2 (L-carnitine) were injected intraperitoneal (i.p.) with normal saline and L-carnitine (200 mg/kg/day), respectively, for 10 successive days. Animals in group 3 (CP group) were injected i.p. with normal saline for 5 days before and 5 days after a single dose of CP (200 mg/kg, i.p.). Rats in group 4 (IFO group) received normal saline for 5 successive days followed by IFO (50 mg/kg/day, i.p.) for 5 successive days. Rats in group 5 (CP-carnitine supplemented) were given the same doses of L-carnitine as group 2 for 5 days before and 5 days after a single dose of CP as group 3. Rats in group 6 (IFO-carnitine supplemented) were given the same doses of L-carnitine as group 2 for 5 days before and 5 days concomitant with IFO as group 4. Immediately, after the last dose of the treatment protocol, blood samples were withdrawn and animals were killed for biochemical, histopathological and gene expression studies. Treatment with CP and IFO significantly decreased expression of heart fatty acid binding protein (H-FABP) and carnitine palmitoyltransferase I (CPT I) genes in cardiac tissues. Moreover, CP but not IFO significantly increased acetyl-CoA carboxylase2 mRNA expression. Conversely, IFO but not CP significantly decreased mRNA expression of malonyl-CoA decarboxylase. Both CP and IFO significantly increased serum lactate dehydrogenase, creatine kinase isoenzyme MB and malonyl-CoA content and histopathological lesions in cardiac tissues. Interestingly, carnitine supplementation completely reversed all the biochemical, histopathological and

  17. Label-Free LC-MS Profiling of Skeletal Muscle Reveals Heart-Type Fatty Acid Binding Protein as a Candidate Biomarker of Aerobic Capacity.

    PubMed

    Malik, Zulezwan Ab; Cobley, James N; Morton, James P; Close, Graeme L; Edwards, Ben J; Koch, Lauren G; Britton, Steven L; Burniston, Jatin G

    2013-12-01

    Two-dimensional gel electrophoresis provides robust comparative analysis of skeletal muscle, but this technique is laborious and limited by its inability to resolve all proteins. In contrast, orthogonal separation by SDS-PAGE and reverse-phase liquid chromatography (RPLC) coupled to mass spectrometry (MS) affords deep mining of the muscle proteome, but differential analysis between samples is challenging due to the greater level of fractionation and the complexities of quantifying proteins based on the abundances of their tryptic peptides. Here we report simple, semi-automated and time efficient (i.e., 3 h per sample) proteome profiling of skeletal muscle by 1-dimensional RPLC electrospray ionisation tandem MS. Solei were analysed from rats (n = 5, in each group) bred as either high- or low-capacity runners (HCR and LCR, respectively) that exhibited a 6.4-fold difference (1,625 ± 112 m vs. 252 ± 43 m, p < 0.0001) in running capacity during a standardized treadmill test. Soluble muscle proteins were extracted, digested with trypsin and individual biological replicates (50 ng of tryptic peptides) subjected to LC-MS profiling. Proteins were identified by triplicate LC-MS/MS analysis of a pooled sample of each biological replicate. Differential expression profiling was performed on relative abundances (RA) of parent ions, which spanned three orders of magnitude. In total, 207 proteins were analysed, which encompassed almost all enzymes of the major metabolic pathways in skeletal muscle. The most abundant protein detected was type I myosin heavy chain (RA = 5,843 ± 897) and the least abundant protein detected was heat shock 70 kDa protein (RA = 2 ± 0.5). Sixteen proteins were significantly (p < 0.05) more abundant in HCR muscle and hierarchal clustering of the profiling data highlighted two protein subgroups, which encompassed proteins associated with either the respiratory chain or fatty acid oxidation. Heart-type fatty acid binding protein (FABPH) was 1.54-fold (p

  18. Metabolism of fatty acids in rat brain in microsomal membranes

    SciTech Connect

    Aeberhard, E.E.; Gan-Elepano, M.; Mead, J.F.

    1980-01-01

    Using a technique in which substrate fatty acids are incorporated into microsomal membranes followd by comparison of their rates of desaturation or elongation with those of exogenous added fatty acids it has been found that the desaturation rate is more rapid for the membrane-bound substrate than for the added fatty acid. Moreover, the product of the membrane-bound substrate is incorporated into membrane phospholipid whereas the product of the exogenous substrate is found in di- and triacyl glycerols and in free fatty acids as well. These and other findings point to a normal sequence of reaction of membrane liqids with membrane-bound substrates involving transfer of fatty acid from phospholipid to the coupled enzyme systems without ready equilibration with the free fatty acid pool.

  19. Identification, Purification, and Molecular Cloning of N-1-Naphthylphthalmic Acid-Binding Plasma Membrane-Associated Aminopeptidases from Arabidopsis1

    PubMed Central

    Murphy, Angus S.; Hoogner, Karen R.; Peer, Wendy Ann; Taiz, Lincoln

    2002-01-01

    Polar transport of the plant hormone auxin is regulated at the cellular level by inhibition of efflux from a plasma membrane (PM) carrier. Binding of the auxin transport inhibitor N-1-naphthylphthalamic acid (NPA) to a regulatory site associated with the carrier has been characterized, but the NPA-binding protein(s) have not been identified. Experimental disparities between levels of high-affinity NPA binding and auxin transport inhibition can be explained by the presence of a low-affinity binding site and in vivo hydrolysis of NPA. In Arabidopsis, colocalization of NPA amidase and aminopeptidase (AP) activities, inhibition of auxin transport by artificial β-naphthylamide substrates, and saturable displacement of NPA by the AP inhibitor bestatin suggest that PM APs may be involved in both low-affinity NPA binding and hydrolysis. We report the purification and molecular cloning of NPA-binding PM APs and associated proteins from Arabidopsis. This is the first report of PM APs in plants. PM proteins were purified by gel permeation, anion exchange, and NPA affinity chromatography monitored for tyrosine-AP activity. Lower affinity fractions contained two orthologs of mammalian APs involved in signal transduction and cell surface-extracellular matrix interactions. AtAPM1 and ATAPP1 have substrate specificities and inhibitor sensitivities similar to their mammalian orthologs, and have temporal and spatial expression patterns consistent with previous in planta histochemical data. Copurifying proteins suggest that the APs interact with secreted cell surface and cell wall proline-rich proteins. AtAPM1 and AtAPP1 are encoded by single genes. In vitro translation products of ATAPM1 and AtAPP1 have enzymatic activities similar to those of native proteins. PMID:11891249

  20. Sequence-specific {sup 1}H, {sup 13}C, and {sup 15}N resonance assignments for intestinal fatty-acid-binding protein complexed with palmitate (15.4 kDA)

    SciTech Connect

    Hodsdon, M.E.; Toner, J.J.; Cistola, D.P.

    1994-12-01

    Intestinal fatty-acid-binding protein (I-FABP) belongs to a family of soluble, cytoplasmic proteins that are thought to function in the intracellular transport and trafficking of polar lipids. Individual members of this protein family have distinct specificities and affinities for fatty acids, cholesterol, bile salts, and retinoids. We are comparing several retinol- and fatty-acid-binding proteins from intestine in order to define the factors that control molecular recognition in this family of proteins. We have established sequential resonance assignments for uniformly {sup 13}C/{sup 15}N-enriched I-FABP complexed with perdeuterated palmitate at pH7.2 and 37{degrees}C. The assignment strategy was similar to that introduced for calmodulin. We employed seven three-dimensional NMR experiments to establish scalar couplings between backbone and sidechain atoms. Backbone atoms were correlated using triple-resonance HNCO, HNCA, TOCSY-HMQC, HCACO, and HCA(CO)N experiments. Sidechain atoms were correlated using CC-TOCSY, HCCH-TOCSY, and TOCSY-HMQC. The correlations of peaks between three-dimensional spectra were established in a computer-assisted manner using NMR COMPASS (Molecular Simulations, Inc.) Using this approach, {sup 1}H, {sup 13}C, and {sup 15}N resonance assignments have been established for 120 of the 131 residues of I-FABP. For 18 residues, amide {sup 1}H and {sup 15}N resonances were unobservable, apparently because of the rapid exchange of amide protons with bulk water at pH 7.2. The missing amide protons correspond to distinct amino acid patterns in the protein sequence, which will be discussed. During the assignment process, several sources of ambiguity in spin correlations were observed. To overcome this ambiguity, the additional inter-residue correlations often observed in the HNCA experiment were used as cross-checks for the sequential backbone assignments.

  1. Proteomic analysis of human papillomavirus-related oral squamous cell carcinoma: identification of thioredoxin and epidermal-fatty acid binding protein as upregulated protein markers in microdissected tumor tissue.

    PubMed

    Melle, Christian; Ernst, Günther; Winkler, Robert; Schimmel, Bettina; Klussmann, Jens Peter; Wittekindt, Claus; Guntinas-Lichius, Orlando; von Eggeling, Ferdinand

    2009-04-01

    Human papillomavirus (HPV) infection has been identified as an etiologic agent for a subset of oral squamous cell carcinoma (OSCC) with increasing incidence. HPV DNA-positivity may confer better prognosis but the related oncogenic mechanisms are unknown. For the identification of HPV relevant proteins, we analyzed microdissected cells from HPV DNA-positive (n = 17) and HPV DNA-negative (n = 7) OSCC tissue samples. We identified 18 proteins from tumor tissues by peptide fingerprint mapping and SELDI MS that were separated using 2-DE. Among a number of signals that were detected as significantly different in the protein profiling analysis, we identified thioredoxin (TRX) and epidermal-fatty acid binding protein as upregulated in HPV related tumor tissue. This study, investigating for the first time proteomic changes in microdissected HPV infected tumor tissue, provides an indication on the oncogenic potential of viruses. PMID:19337991

  2. Plasma membrane phospholipase A2 controls hepatocellular fatty acid uptake and is responsive to pharmacological modulation: implications for nonalcoholic steatohepatitis.

    PubMed

    Stremmel, Wolfgang; Staffer, Simone; Wannhoff, Andreas; Pathil, Anita; Chamulitrat, Walee

    2014-07-01

    Excess hepatic fat accumulation leads to nonalcoholic steatohepatitis (NASH), a serious threat to health for which no effective treatment is available. However, the mechanism responsible for fatty acid uptake by hepatocytes remains unclear. Using the human hepatocyte-derived tumor cell line HepG2, we found that fatty acid influx is mediated by a heterotetrameric plasma membrane protein complex consisting of plasma membrane fatty acid-binding protein, caveolin-1, CD36, and calcium-independent membrane phospholipase A2 (iPLA2β). Blocking iPLA2β with the bile acid-phospholipid conjugate ursodeoxycholate-lysophosphatidylethanolamide (UDCA-LPE) caused the dissociation of the complex, thereby inhibiting fatty acid influx (IC50 47 μM), and suppressed the synthesis of all subunits through a reduction in lysophosphatidylcholine from 8.0 to 3.5 μmol/mg of protein and corresponding depletion of phosphorylated c-Jun N-terminal kinase. These findings were substantiated by an observed 56.5% decrease in fatty acid influx in isolated hepatocytes derived from iPLA2β-knockout mice. Moreover, steatosis and inflammation were abrogated by UDCA-LPE treatment in a cellular model of NASH. Thus, iPLA2β acts as an upstream checkpoint for mechanisms that regulate fatty acid uptake, and its inhibition by UDCA-LPE qualifies this nontoxic compound as a therapeutic candidate for the treatment of NASH.-Stremmel, W., Staffer, S., Wannhoff, A., Pathil, A., Chamulitrat, W. Plasma membrane phospholipase A2 controls hepatocellular fatty acid uptake and is responsive to pharmacological modulation: implications for nonalcoholic steatohepatitis. PMID:24719358

  3. Fatty acids, membrane viscosity, serotonin and ischemic heart disease

    PubMed Central

    2010-01-01

    Novel markers for ischemic heart disease are under investigation by the scientific community at international level. This work focuses on a specific platelet membrane fatty acid condition of viscosity which is linked to molecular aspects such as serotonin and G proteins, factors involved in vascular biology. A suggestive hypothesis is considered about the possibility to use platelet membrane viscosity, in relation to serotonin or, indirectly, the fatty acid profile, as indicator of ischemic risk. PMID:20825633

  4. Effects of the gut microbiota on host adiposity are modulated by the short-chain fatty-acid binding G protein-coupled receptor, Gpr41

    PubMed Central

    Samuel, Buck S.; Shaito, Abdullah; Motoike, Toshiyuki; Rey, Federico E.; Backhed, Fredrik; Manchester, Jill K.; Hammer, Robert E.; Williams, S. Clay; Crowley, Jan; Yanagisawa, Masashi; Gordon, Jeffrey I.

    2008-01-01

    The distal human intestine harbors trillions of microbes that allow us to extract calories from otherwise indigestible dietary polysaccharides. The products of polysaccharide fermentation include short-chain fatty acids that are ligands for Gpr41, a G protein-coupled receptor expressed by a subset of enteroendocrine cells in the gut epithelium. To examine the contribution of Gpr41 to energy balance, we compared Gpr41−/− and Gpr41+/+ mice that were either conventionally-raised with a complete gut microbiota or were reared germ-free and then cocolonized as young adults with two prominent members of the human distal gut microbial community: the saccharolytic bacterium, Bacteroides thetaiotaomicron and the methanogenic archaeon, Methanobrevibacter smithii. Both conventionally-raised and gnotobiotic Gpr41−/− mice colonized with the model fermentative community are significantly leaner and weigh less than their WT (+/+) littermates, despite similar levels of chow consumption. These differences are not evident when germ-free WT and germ-free Gpr41 knockout animals are compared. Functional genomic, biochemical, and physiologic studies of germ-free and cocolonized Gpr41−/− and +/+ littermates disclosed that Gpr41-deficiency is associated with reduced expression of PYY, an enteroendocrine cell-derived hormone that normally inhibits gut motility, increased intestinal transit rate, and reduced harvest of energy (short-chain fatty acids) from the diet. These results reveal that Gpr41 is a regulator of host energy balance through effects that are dependent upon the gut microbiota. PMID:18931303

  5. Fatty Acids Inhibit Apical Membrane Chloride Channels in Airway Epithelia

    NASA Astrophysics Data System (ADS)

    Anderson, Matthew P.; Welsh, Michael J.

    1990-09-01

    Apical membrane Cl^- channels control the rate of transepithelial Cl^- secretion in airway epithelia. cAMP-dependent protein kinase and protein kinase C regulate Cl^- channels by phosphorylation; in cystic fibrosis cells, phosphorylation-dependent activation of Cl^- channels is defective. Another important signaling system involves arachidonic acid, which is released from cell membranes during receptor-mediated stimulation. Here we report that arachidonic acid reversibly inhibited apical membrane Cl^- channels in cell-free patches of membrane. Arachidonic acid itself inhibited the channel and not a cyclooxygenase or lipoxygenase metabolite because (i) inhibitors of these enzymes did not block the response, (ii) fatty acids that are not substrates for the enzymes had the same effect as arachidonic acid, and (iii) metabolites of arachidonic acid did not inhibit the channel. Inhibition occurred only when fatty acids were added to the cytosolic surface of the membrane patch. Unsaturated fatty acids were more potent than saturated fatty acids. Arachidonic acid inhibited Cl^- channels from both normal and cystic fibrosis cells. These results suggest that fatty acids directly inhibit apical membrane Cl^- channels in airway epithelial cells.

  6. Glycated albumin with loss of fatty acid binding capacity contributes to enhanced arachidonate oxygenation and platelet hyperactivity: relevance in patients with type 2 diabetes.

    PubMed

    Blache, Denis; Bourdon, Emmanuel; Salloignon, Pauline; Lucchi, Géraldine; Ducoroy, Patrick; Petit, Jean-Michel; Verges, Bruno; Lagrost, Laurent

    2015-03-01

    High plasma concentrations of nonesterified fatty acids (NEFAs), transported bound to serum albumin, are associated with type 2 diabetes (T2D). The effects of albumin on platelet function were investigated in vitro. Modifications of albumin, such as those due to glycoxidation, were found in patients with T2D, and the consequences of these modifications on biological mechanisms related to NEFA handling were investigated. Mass spectrometry profiles of albumin from patients with T2D differed from those from healthy control subjects. Diabetic albumin showed impaired NEFA binding capacity, and both structural and functional alterations could be reproduced in vitro by incubating native albumin with glucose and methylglyoxal. Platelets incubated with albumin isolated from patients with T2D aggregated approximately twice as much as platelets incubated with albumin isolated from healthy control subjects. Accordingly, platelets incubated with modified albumin produced significantly higher amounts of arachidonate metabolites than did platelets incubated with control albumin. We concluded that higher amounts of free arachidonate are made available for the generation of active metabolites in platelets when the NEFA binding capacity of albumin is blunted by glycoxidation. This newly described mechanism, in addition to hypoalbuminemia, may contribute to platelet hyperactivity and increased thrombosis, known to occur in patients with T2D. PMID:25157094

  7. Fatty acids binding to human serum albumin: Changes of reactivity and glycation level of Cysteine-34 free thiol group with methylglyoxal.

    PubMed

    Pavićević, Ivan D; Jovanović, Vesna B; Takić, Marija M; Penezić, Ana Z; Aćimović, Jelena M; Mandić, Ljuba M

    2014-10-17

    Fatty acids (FAs) binding to human serum albumin (HSA) could lead to the changes of Cys-34 thiol group accessibility and reactivity, i.e. its scavenger capacity and antioxidant property. The influence of saturated, mono and poly unsaturated, and fish oil FAs binding to HSA on the carbonylation level and the reactivity of HSA-SH and HSA modified with methylglyoxal (MG-HSA-SH) was investigated. Changes of thiol group reactivity were followed by determination of pseudo first order rate constant (k') for thiols reaction with 5,5'-dithiobis(2-nitrobenzoic acid). HSA changes were monitored using native PAG electrophoresis and fluorescence spectroscopy. For FA/HSA molar ratios screening, qTLC and GC were used. FAs increase thiol group carbonylation levels from 8% to 20%. The k' values obtained for FAs-free HSA-SH and FAs-free MG-HSA-SH are almost equal (7.5×10(-3) and 7.7×10(-3)s(-1), resp.). Binding of all FAs amplify the reactivity (k' values from 14.6×10(-3) to 26.0×10(-3)s(-1)) of HSA-SH group for 2-3.5times in the order: palmitic, docosahexaenoic, fish oil extract, stearic, oleic, myristic and eicosapentaenoic acid, due to HSA conformational changes. FAs-bound MG-HSA-SH samples follow that pattern, but their k' values (from 9.8×10(-3) to 14.3×10(-3)s(-1)) were lower compared to unmodified HSA due to additional conformation changes of HSA molecules during carbonylation. Carbonylation level and reactivity of Cys34 thiol group of unmodified and carbonylated HSA depend on type of FAs bound to HSA, which implies the possibility for modulation of -SH reactivity (scavenger capacity and antioxidant property) by FAs as a supplement. PMID:25451573

  8. Fatty acid binding protein 4 and 5 play a crucial role in thermogenesis under the conditions of fasting and cold stress.

    PubMed

    Syamsunarno, Mas Rizky A A; Iso, Tatsuya; Yamaguchi, Aiko; Hanaoka, Hirofumi; Putri, Mirasari; Obokata, Masaru; Sunaga, Hiroaki; Koitabashi, Norimichi; Matsui, Hiroki; Maeda, Kazuhisa; Endo, Keigo; Tsushima, Yoshito; Yokoyama, Tomoyuki; Kurabayashi, Masahiko

    2014-01-01

    Hypothermia is rapidly induced during cold exposure when thermoregulatory mechanisms, including fatty acid (FA) utilization, are disturbed. FA binding protein 4 (FABP4) and FABP5, which are abundantly expressed in adipose tissues and macrophages, have been identified as key molecules in the pathogenesis of overnutrition-related diseases, such as insulin resistance and atherosclerosis. We have recently shown that FABP4/5 are prominently expressed in capillary endothelial cells in the heart and skeletal muscle and play a crucial role in FA utilization in these tissues. However, the role of FABP4/5 in thermogenesis remains to be determined. In this study, we showed that thermogenesis is severely impaired in mice lacking both FABP4 and FABP5 (DKO mice), as manifested shortly after cold exposure during fasting. In DKO mice, the storage of both triacylglycerol in brown adipose tissue (BAT) and glycogen in skeletal muscle (SkM) was nearly depleted after fasting, and a biodistribution analysis using 125I-BMIPP revealed that non-esterified FAs (NEFAs) are not efficiently taken up by BAT despite the robustly elevated levels of serum NEFAs. In addition to the severe hypoglycemia observed in DKO mice during fasting, cold exposure did not induce the uptake of glucose analogue 18F-FDG by BAT. These findings strongly suggest that DKO mice exhibit pronounced hypothermia after fasting due to the depletion of energy storage in BAT and SkM and the reduced supply of energy substrates to these tissues. In conclusion, FABP4/5 play an indispensable role in thermogenesis in BAT and SkM. Our study underscores the importance of FABP4/5 for overcoming life-threatening environments, such as cold and starvation. PMID:24603714

  9. Fatty acid uptake in normal human myocardium

    SciTech Connect

    Vyska, K.; Meyer, W.; Stremmel, W.; Notohamiprodjo, G.; Minami, K.; Machulla, H.J.; Gleichmann, U.; Meyer, H.; Koerfer, R. )

    1991-09-01

    Fatty acid binding protein has been found in rat aortic endothelial cell membrane. It has been identified to be a 40-kDa protein that corresponds to a 40-kDa fatty acid binding protein with high affinity for a variety of long chain fatty acids isolated from rat heart myocytes. It is proposed that this endothelial membrane fatty acid binding protein might mediate the myocardial uptake of fatty acids. For evaluation of this hypothesis in vivo, influx kinetics of tracer-labeled fatty acids was examined in 15 normal subjects by scintigraphic techniques. Variation of the plasma fatty acid concentration and plasma perfusion rate has been achieved by modulation of nutrition state and exercise conditions. The clinical results suggest that the myocardial fatty acid influx rate is saturable by increasing fatty acid plasma concentration as well as by increasing plasma flow. For analysis of these data, functional relations describing fatty acid transport from plasma into myocardial tissue in the presence and absence of an unstirred layer were developed. The fitting of these relations to experimental data indicate that the free fatty acid influx into myocardial tissue reveals the criteria of a reaction on a capillary surface in the vicinity of flowing plasma but not of a reaction in extravascular space or in an unstirred layer and that the fatty acid influx into normal myocardium is a saturable process that is characterized by the quantity corresponding to the Michaelis-Menten constant, Km, and the maximal velocity, Vmax, 0.24 {plus minus} 0.024 mumol/g and 0.37 {plus minus} 0.013 mumol/g(g.min), respectively. These data are compatible with a nondiffusional uptake process mediated by the initial interaction of fatty acids with the 40-kDa membrane fatty acid binding protein of cardiac endothelial cells.

  10. A single-nucleotide polymorphism in the 3′-UTR region of the adipocyte fatty acid binding protein 4 gene is associated with prognosis of triple-negative breast cancer

    PubMed Central

    Wang, Wenmiao; Yuan, Peng; Yu, Dianke; Du, Feng; Zhu, Anjie; Li, Qing; Zhang, Pin; Lin, Dongxin; Xu, Binghe

    2016-01-01

    Triple-negative breast cancer (TNBC) is a subtype of breast cancer with poor prognosis and high heterogeneity. The aim of this study was to screen patients for single-nucleotide polymorphisms (SNPs) associated with the prognosis of TNBC. Database-derived SNPs (NextBio, Ensembl, NCBI and MirSNP) located in the 3′-untranslated regions (3′-UTRs) of genes that are differentially expressed in breast cancer were selected. The possible associations between 111 SNPs and progression risk among 323 TNBC patients were investigated using a two-step case-control study with a discovery cohort (n=162) and a validation cohort (n=161). We identified the rs1054135 SNP in the adipocyte fatty acid binding protein 4 (FABP4) gene as a predictor of TNBC recurrence. The G allele of rs1054135 was associated with a reduced risk of disease progression as well as a prolonged disease-free survival time (DFS), with a hazard ratio (HR) for recurrence in the combined sample of 0.269 [95%CI: 0.098−0.735;P=0.001]. Notably, for individuals having the rs1054135 SNP with the AA/AG genotype, the magnitude of increased tumour recurrence risk for overweight patients (BMI≥25kg/m2) was significantly elevated (HR2.53; 95%CI: 1.06–6.03). Immunohistochemical staining of adipocytes adjacent to TNBC tissues showed that the expression level of FABP4 was statistically significantly lower in patients with the rs1054135-GG genotype and those in the disease-free group (P=0.0004 and P=0.0091, respectively). These results suggested that the expression of a lipid metabolism-related gene and an important SNP in the 3′-UTR of FABP4 are associated with TNBC prognosis, which may aid in the screening of high-risk patients with TNBC recurrence and the development of novel chemotherapeutic agents. PMID:26959740

  11. Heart-type fatty acid binding protein and high-sensitivity troponin T are myocardial damage markers that could predict adverse clinical outcomes in patients with peripheral artery disease

    PubMed Central

    Otaki, Yoichiro; Takahashi, Hiroki; Watanabe, Tetsu; Yamaura, Gensai; Funayama, Akira; Arimoto, Takanori; Shishido, Tetsuro; Miyamoto, Takuya; Kubota, Isao

    2015-01-01

    Background Despite many recent advances in endovascular therapy (EVT), peripheral artery disease (PAD) is an increasing health problem with high mortality. Heart-type fatty acid-binding protein (H-FABP) and high-sensitivity troponin T (hsTnT) are markers of ongoing myocardial damage and have been reported to be useful indicators of future cardiovascular events. However, it remains to be determined whether H-FABP and hsTnT can predict adverse clinical outcomes in patients with PAD. Methods and results We enrolled 208 de novo PAD patients who underwent EVT. Serum H-FABP and hsTnT were measured in all patients before EVT. During the median follow-up period of 694 days, there were 40 major adverse cardiovascular and cerebrovascular events (MACCEs) including all-cause deaths, and re-hospitalizations due to cardiovascular and cerebrovascular diseases and amputations. H-FABP and hsTnT were found to be higher in patients with critical limb ischemia (CLI) compared to those without this condition. Multivariate Cox proportional hazard regression analysis revealed that both H-FABP and hsTnT were independent predictors of MACCEs after adjustment for confounding factors. Kaplan–Meier analysis demonstrated that patients in the highest tertile according to H-FABP levels, as well as those in the highest hsTnT tertile, were at greatest risk for MACCEs. The net reclassification index was significantly improved by the addition of H-FABP as well as the addition of hsTnT to traditional risk factors. Conclusion The myocardial damage markers H-FABP and hsTnT were increased in PAD patients with CLI and could predict MACCEs in PAD patients. PMID:26673681

  12. Comparison of a qualitative measurement of heart-type fatty acid-binding protein with other cardiac markers as an early diagnostic marker in the diagnosis of non-ST - segment elevation myocardial infarction

    PubMed Central

    Gerede, Demet Menekşe; Güleç, Sadi; Kılıçkap, Mustafa; Kaya, Cansın Tulunay; Vurgun, Veysel Kutay; Özcan, Özgür Ulaş; Göksülük, Hüseyin; Erol, Çetin

    2015-01-01

    Summary Objective: Heart-type fatty acid-binding protein (H-FABP) is a novel cardiac marker used in the early diagnosis of acute myocardial infarction (AMI), which shows myocyte injury. Our study aimed to compare bedside H-FABP measurements with routine creatine kinase-MB (CK-MB) and troponin I (TnI) tests for the early diagnosis of non-ST-elevation MI (NSTEMI), as well as for determining its exclusion capacity. Methods A total of 48 patients admitted to the emergency room within the first 12 hours of onset of ischaemic-type chest pain lasting more than 30 minutes and who did not have ST-segment elevation on electrocardiography (ECG) were included in the study. Definite diagnoses of NSTEMI were made in 24 patients as a result of 24-hour follow up, and the remaining 24 patients did not develop MI. Results When various subgroups were analysed according to admission times, H-FABP was found to be a better diagnostic marker compared to CK-MB and TnI (accuracy index 85%), with a high sensitivity (79%) and specificity (93%) for early diagnosis (≤ six hours). The respective sensitivities of bedside H-FABP and TnI tests were 89 vs 33% (p < 0.05) for patients presenting within three hours of onset of symptoms. Conclusion Bedside H-FABP measurements may contribute to correct early diagnoses, as its levels are elevated soon following MI, and measurement is easy, with a rapid result. PMID:26212703

  13. A newly developed kit for the measurement of urinary liver-type fatty acid-binding protein as a biomarker for acute kidney injury in patients with critical care.

    PubMed

    Sato, Ryo; Suzuki, Yasushi; Takahashi, Gaku; Kojika, Masahiro; Inoue, Yoshihiro; Endo, Shigeatsu

    2015-03-01

    In recent years, it has been reported that the urinary level of Liver-type fatty acid-binding protein (L-FABP) serves as a useful biomarker for diagnosing acute kidney injury (AKI) or sepsis complicated by AKI. However, because the urinary level of L-FABP is currently measured by enzyme-linked immunosorbent assay (ELISA), several days may elapse before the results of the measurement become available. We have newly developed a simplified kit, the Dip-test, for measuring the urinary level of L-FABP. The Dip-test was measured at 80 measurement points (22 points in noninfectious disease, 13 points in SIRS, 20 points in infectious disease, and 25 points in sepsis) in 20 patients. The urinary L-FABP levels as determined by ELISA in relation to the results of the Dip-test were as follows: 10.10 ± 12.85 ng/ml in patients with a negative Dip-test ([-] group), 41.93 ± 50.51 ng/ml in patients with a ± test ([±] group), 70.36 ± 73.70 ng/ml in patients with a positive test ([+] group), 1048.96 ± 2117.68 ng/ml in patients with a 2 + test ([2+] group), and 23,571.55 ± 21,737.45 ng/ml in patients with a 3 + test ([3+] group). The following tendency was noted: the stronger the positive Dip-test reaction, the higher the urinary L-FABP level. Multigroup comparison revealed a significant differences in the urinary L-FABP levels between the Dip-test (-) group and each of the other groups. In this study, the usefulness of the Dip-test, our newly developed simplified kit for measuring the urinary L-FABP level, is suggested. PMID:25499195

  14. Rosuvastatin Decreases Intestinal Fatty Acid Binding Protein (I-FABP), but Does Not Alter Zonulin or Lipopolysaccharide Binding Protein (LBP) Levels, in HIV-Infected Subjects on Antiretroviral Therapy

    PubMed Central

    Funderburg, Nicholas T.; Boucher, Morgan; Sattar, Abdus; Kulkarni, Manjusha; Labbato, Danielle; Kinley, Bruce I.; McComsey, Grace A.

    2016-01-01

    Introduction Altered gastrointestinal (GI) barrier integrity and subsequent microbial translocation may contribute to immune activation in HIV infection. We have reported that rosuvastatin improved several markers of immune activation in HIV+ participants, but the effect of statin treatment on markers of GI barrier dysfunction is unknown. Methods SATURN-HIV is a randomized, double-blind, placebo-controlled trial assessing the effect of rosuvastatin (10mg/daily) on markers of cardiovascular disease, inflammation, and immune activation in ART-treated patients. Gut-barrier integrity was assessed by the surrogate markers intestinal fatty acid binding protein (I-FABP), a marker of enterocyte death, and zonulin-1, a marker of gut epithelial cell function. Levels of lipopolysaccharide binding protein (LBP) were measured as a marker of microbial translocation. Results Rosuvastatin significantly reduced levels of I-FABP during the treatment period compared to the placebo. There was no effect of rosuvastatin treatment on levels of zonulin or LBP. Baseline levels of LBP were directly related to several markers of immune activation in samples from all participants, including soluble CD163, IP-10, VCAM-1, TNFR-II, and the proportion of CD4+ and CD8+ T cells expressing CD38 and HLA-DR. Many of these relationships, however, were not seen in the statin arm alone at baseline or over time, as inflammatory markers often decreased and LBP levels were unchanged. Conclusions Forty-eight weeks of rosuvastatin treatment reduced levels of I-FABP, but did not affect levels of zonulin or LBP. The reduction in levels of inflammatory markers that we have reported with rosuvastatin treatment is likely mediated through other mechanisms not related to gut integrity or microbial translocation.

  15. Level of urinary liver-type fatty acid-binding protein is associated with cardiac markers and electrocardiographic abnormalities in type-2 diabetes with chronic kidney disease stage G1 and G2.

    PubMed

    Maeda, Yoshiteru; Suzuki, Atsushi; Ishii, Junnichi; Sekiguchi-Ueda, Sahoko; Shibata, Megumi; Yoshino, Yasumasa; Asano, Shogo; Hayakawa, Nobuki; Nakamura, Kazuhiro; Akiyama, Yasukazu; Kitagawa, Fumihiko; Sakuishi, Toshiaki; Fujita, Takashi; Hashimoto, Shuji; Ozaki, Yukio; Itoh, Mitsuyasu

    2015-05-01

    Urinary liver-type fatty acid-binding protein (L-FABP) reflects the degree of stress in proximal tubules of the kidney. We examined the level of L-FABP in type-2 diabetes mellitus (T2DM) patients with chronic kidney disease (CKD) stage G1 and G2, and its relationship with cardiac markers and electrocardiographic (ECG) abnormalities. T2DM patients whose estimated glomerular filtration rate (eGFR) was ≥60 mL/min/1.73 m(2) were recruited [n = 276 (165 males), mean age 64 years]. The median level of urinary L-FABP was 6.6 μg/gCr. Urinary L-FABP showed significant correlation with urinary albumin-to-creatinine ratio (ACR) (r = 0.51, p < 0.0001). Median (25th-75th percentile) eGFR was 82 (72-95) mL/min/1.73 m2. We divided patients into four subgroups (group 1, L-FABP ≤8.4 μg/gCr and ACR ≤30 mg/gCr; group 2, L-FABP ≤8.4 μg/gCr and ACR >30 mg/gCr; group 3, L-FABP >8.4 μg/gCr and ACR ≤30 mg/gCr; group 4, L-FABP >8.4 μg/gCr and ACR >30 mg/gCr). Compared with group 1, group 4 was significantly higher in systolic blood pressure, and eGFR using standardized serum cystatin C, high-sensitivity troponin T, and N-terminal pro-brain natriuretic peptide (NT-proBNP). Group 4 had significantly higher level of NT-proBNP than group 3. Groups 2, 3 and 4 showed more ECG abnormalities than group 1. These findings suggest that simultaneous measurement of urinary L-FABP and ACR should be useful to assess cardiovascular damage reflecting on the elevation of cardiac markers and ECG abnormalities in T2DM with CKD G1 and G2. PMID:24626813

  16. Evaluation of New Diagnostic Biomarkers in Pediatric Sepsis: Matrix Metalloproteinase-9, Tissue Inhibitor of Metalloproteinase-1, Mid-Regional Pro-Atrial Natriuretic Peptide, and Adipocyte Fatty-Acid Binding Protein

    PubMed Central

    Alqahtani, Mashael F.; Smith, Craig M.; Weiss, Scott L.; Dawson, Susan; Ralay Ranaivo, Hantamalala; Wainwright, Mark S.

    2016-01-01

    Elevated plasma concentrations of matrix metalloproteinase-9 (MMP-9), tissue inhibitor of metalloproteinase-1 (TIMP-1), mid-regional pro-atrial natriuretic peptide (mrProANP), and adipocyte fatty-acid-binding proteins (A-FaBPs) have been investigated as biomarkers for sepsis or detection of acute neurological injuries in adults, but not children. We carried out a single-center, prospective observational study to determine if these measures could serve as biomarkers to identify children with sepsis. A secondary aim was to determine if these biomarkers could identify children with neurologic complications of sepsis. A total of 90 patients ≤ 18 years-old were included in this study. 30 with severe sepsis or septic shock were compared to 30 age-matched febrile and 30 age-matched healthy controls. Serial measurements of each biomarker were obtained, beginning on day 1 of ICU admission. In septic patients, MMP9-/TIMP-1 ratios (Median, IQR, n) were reduced on day 1 (0.024, 0.004–0.174, 13), day 2 (0.020, 0.002–0.109, 10), and day 3 (0.018, 0.003–0.058, 23) compared with febrile (0.705, 0.187–1.778, 22) and healthy (0.7, 0.4–1.2, 29) (p< 0.05) controls. A-FaBP and mrProANP (Median, IQR ng/mL, n) were elevated in septic patients compared to control groups on first 2 days after admission to the PICU (p <0.05). The area under the curve (AUC) for MMP-9/TIMP-1 ratio, mrProANP, and A-FaBP to distinguish septic patients from healthy controls were 0.96, 0.99, and 0.76, respectively. MMP-9/TIMP-1 ratio was inversely and mrProANP was directly related to PIM-2, PELOD, and ICU and hospital LOS (p<0.05). A-FaBP level was associated with PELOD, hospital and ICU length of stay (p<0.05). MMP-9/TIMP-1 ratio associated with poor Glasgow Outcome Score (p<0.05). A-FaBP levels in septic patients with neurological dysfunction (29.3, 17.2–54.6, 7) were significantly increased compared to septic patients without neurological dysfunction (14.6, 13.3–20.6, 11). MMP-9/TIMP-1 ratios

  17. NMR unfolding studies on a liver bile acid binding protein reveal a global two-state unfolding and localized singular behaviors.

    PubMed

    D'Onofrio, Mariapina; Ragona, Laura; Fessas, Dimitrios; Signorelli, Marco; Ugolini, Raffaella; Pedò, Massimo; Assfalg, Michael; Molinari, Henriette

    2009-01-01

    The folding properties of a bile acid binding protein, belonging to a subfamily of the fatty acid binding proteins, have been here investigated both by hydrogen exchange measurements, using the SOFAST NMR approach, and urea denaturation experiments. The urea unfolding profiles of individual residues, acting as single probes, were simultaneously analyzed through a global fit, according to a two-state unfolding model. The resulting conformational stability DeltaG(U)(H(2)O)=7.2+/-0.25kcal mol(-1) is in good agreement with hydrogen exchange stability DeltaG(op). While the majority of protein residues satisfy this model, few amino-acids display a singular behavior, not directly amenable to the presence of a folding intermediate, as reported for other fatty acid binding proteins. These residues are part of a protein patch characterized by enhanced plasticity. To explain this singular behavior a tentative model has been proposed which takes into account the interplay between the dynamic features and the formation of transient aggregates. A functional role for this plasticity, related to translocation across the nuclear membrane, is discussed. PMID:18977333

  18. Identification and Characterization of Linoleic Acid as an Endogenous Modulator of in Vitro N-1-Naphthylphthalamic Acid Binding.

    PubMed Central

    Suttle, J. C.

    1997-01-01

    An endogenous inhibitor of the in vitro binding of the phytotropin N-1-naphthylphthalamic acid to microsomal membranes was detected in extracts prepared from etiolated pea (Pisum sativum L.) epicotyls. Following extensive purification, the inhibitor was identified as linoleic acid. Authentic linoleic acid inhibited N-1-naphthylphthalamic acid binding noncompetitively in a dose-dependent manner, exhibiting a 50% inhibitory concentration of approximately 24 ([mu]M. Using a variety of fatty acids and their derivatives, this inhibition was found to exhibit strict structural requirements, with both linoleic and linolenic acids being the most inhibitory. A variety of membrane-solubilizing detergents elicited no such inhibitory activity when tested at equivalent concentrations. The possible physiological significance of this interaction is discussed and it is proposed that linoleic acid serves as an intracellular modulator of phytotropin binding and therefore polar auxin transport. PMID:12223622

  19. Copper toxicity towards Saccharomyces cerevisiae: dependence on plasma membrane fatty acid composition.

    PubMed Central

    Avery, S V; Howlett, N G; Radice, S

    1996-01-01

    One major mechanism of copper toxicity towards microorganisms is disruption of plasma membrane integrity. In this study, the influence of plasma membrane fatty acid composition on the susceptibility of Saccharomyces cerevisiae to Cu2+ toxicity was investigated. Microbial fatty acid composition is highly variable, depending on both intrinsic and environmental factors. Manipulation was achieved in this study by growth in fatty acid-supplemented medium. Whereas cells grown under standard conditions contained only saturated and monounsaturated fatty acids, considerable incorporation of the diunsaturated fatty acid linoleate (18:2) (to more than 65% of the total fatty acids) was observed in both whole-cell homogenates and plasma membrane-enriched fractions from cells grown in linoleate-supplemented medium. Linoleate enrichment had no discernible effect on the growth of S. cerevisiae. However, linoleate-enriched cells were markedly more susceptible to copper-induced plasma membrane permeabilization. Thus, after addition of Cu(NO3)2, rates of cellular K+ release (loss of membrane integrity) were at least twofold higher from linoleate-supplemented cells than from unsupplemented cells; this difference increased with reductions in the Cu2+ concentration supplied. Levels of cellular Cu accumulation were also higher in linoleate-supplemented cells. These results were correlated with a very marked dependence of whole-cell Cu2+ toxicity on cellular fatty acid unsaturation. For example, within 10 min of exposure to 5 microM Cu2+, only 3% of linoleate-enriched cells remained viable (capable of colony formation). In contrast, 100% viability was maintained in cells previously grown in the absence of a fatty acid supplement. Cells displaying intermediate levels of linoleate incorporation showed intermediate Cu2+ sensitivity, while cells enriched with the triunsaturated fatty acid linolenate (18:3) were most sensitive to Cu2+. These results demonstrate for the first time that changes

  20. The effect of hydroxylated fatty acid-containing phospholipids in the remodeling of lipid membranes.

    PubMed

    Piotto, Stefano; Trapani, Alfonso; Bianchino, Erminia; Ibarguren, Maitane; López, David J; Busquets, Xavier; Concilio, Simona

    2014-06-01

    The synthetic fatty acid 2-hydroxyoleic acid (2OHOA) is an antitumor drug that regulates membrane lipid composition and structure. An important effect of this drug is the restoration of sphingomyelin (SM) levels in cancer cell membranes, where the SM concentration is lower than in non-tumor cells. It is well known that free fatty acid concentration in cell membranes is lower than 5%, and that fatty acid excess is rapidly incorporated into phospholipids. In a recent work, we have considered the effect of free 2OHOA in model membranes in liquid ordered (Lo) and liquid disordered (Ld) phases, by using all-atom molecular dynamics. This study concerns membranes that are modified upon incorporation of 2OHOA into different phospholipids. 2OHOA-containing phospholipids have a permanent effect on lipid membranes, making a Ld membrane surface more compact and less hydrated, whereas the opposite effect is observed in Lo domains. Moreover, the hydroxyl group of fatty acid chains increases the propensity of Ld model membranes to form hexagonal or other non-lamellar structures. This article is part of a Special Issue entitled: Membrane Structure and Function: Relevance in the Cell's Physiology, Pathology and Therapy. PMID:24463068

  1. Erythrocyte Membrane Fatty Acid Composition in Premenopausal Patients with Iron Deficiency Anemia.

    PubMed

    Aktas, Mehmet; Elmastas, Mahfuz; Ozcicek, Fatih; Yilmaz, Necmettin

    2016-03-01

    Iron deficiency anemia (IDA) is one of the most common nutritional disorders in the world. In the present study, we evaluated erythrocyte membrane fatty acid composition in premenopausal patients with IDA. Blood samples of 102 premenopausal women and 88 healthy control subjects were collected. After the erythrocytes were separated from the blood samples, the membrane lipids were carefully extracted, and the various membrane fatty acids were measured by gas chromatography (GC). Statistical analyses were performed with the SPSS software program. We used blood ferritin concentration <15 ng/mL as cut-off for the diagnosis of IDA. The five most abundant individual fatty acids obtained were palmitic acid (16:0), oleic acid (18:1, n-9c), linoleic acid (18:2, n-6c), stearic acid (18:0), and erucic acid (C22:1, n-9c). These compounds constituted about 87% of the total membrane fatty acids in patients with IDA, and 79% of the total membrane fatty acids in the control group. Compared with control subjects, case patients had higher percentages of palmitic acid (29.9% case versus 25.3% control), oleic acid (16.8% case versus 15.1% control), and stearic acid (13.5% case versus 10.5% control), and lower percentages of erucic acid (11.5% case versus 13.6% control) and linoleic acid (15.2% case versus 15.4% control) in their erythrocyte membranes. In conclusion, the total-erythrocyte-membrane saturated fatty acid (SFA) composition in premenopausal women with IDA was found to be higher than that in the control group; however, the total-erythrocyte-membrane unsaturated fatty acid (UFA) composition in premenopausal women with IDA was found to be lower than that in the control group. The differences in these values were statistically significant. PMID:26876679

  2. Specific effect of polyunsaturated fatty acids on the cholesterol-poor membrane domain in a model membrane.

    PubMed

    Onuki, Yoshinori; Hagiwara, Chihiro; Sugibayashi, Ko; Takayama, Kozo

    2008-08-01

    To understand more fully the effect of polyunsaturated fatty acids (PUFAs) on lipid bilayers, we investigated the effects of treatment with fatty acids on the properties of a model membrane. Three kinds of liposomes comprising dipalmitoylphosphatidylcholine (DPPC), dioleylphosphatidylcholine (DOPC), and cholesterol (Ch) were used as the model membrane, and the fluorescence anisotropy of 1,6-diphenyl-1,3,5-hexatriene (DPH) and detergent insolubility were determined. Characterization of the liposomes clarified that DPPC, DPPC/Ch, and DPPC/DOPC/Ch existed as solid-ordered phase (L beta), liquid-ordered phase (l o), and a mixture of l o and liquid-disordered phase (L alpha) membranes at room temperature. Treatment with unsaturated fatty acids such as oleic acid (OA), eicosapentaenoic acid (EPA), and docosahexaenoic acid (DHA) markedly decreased the fluorescence anisotropy value and detergent insolubility. PUFAs and OA had different effects on the model membranes. In DPPC liposomes, the most prominent change was induced by PUFAs, whereas, in DPPC/Ch and DPPC/DOPC/Ch liposomes, OA had a stronger effect than PUFAs. The effect of PUFAs was strongly affected by the amount of Ch in the membrane, which confirmed a specific effect of PUFAs on the Ch-poor membrane domain. We further explored the effect of fatty acids dispersed in a water-in-oil-in-water multiple emulsion and found that unsaturated fatty acids acted on the membranes even when incorporated in emulsion form. These findings suggest that treatment with PUFAs increases the segregation of ordered and disordered phase domains in membranes. PMID:18670110

  3. Photoaffinity labeling of retinoic acid-binding proteins.

    PubMed Central

    Bernstein, P S; Choi, S Y; Ho, Y C; Rando, R R

    1995-01-01

    Retinoid-binding proteins are essential mediators of vitamin A function in vertebrate organisms. They solubilize and stabilize retinoids, and they direct the intercellular and intracellular trafficking, transport, and metabolic function of vitamin A compounds in vision and in growth and development. Although many soluble retinoid-binding proteins and receptors have been purified and extensively characterized, relatively few membrane-associated enzymes and other proteins that interact with retinoids have been isolated and studied, due primarily to their inherent instabilities during purification. In an effort to identify and purify previously uncharacterized retinoid-binding proteins, it is shown that radioactively labeled all-trans-retinoic acid can be used as a photoaffinity labeling reagent to specifically tag two known retinoic acid-binding proteins, cellular retinoic acid-binding protein and albumin, in complex mixtures of cytosolic proteins. Additionally, a number of other soluble and membrane-associated proteins that bind all-trans-[11,12-3H]retinoic acid with high specificity are labeled utilizing the same photoaffinity techniques. Most of these labeled proteins have molecular weights that do not correspond to any known retinoid-binding proteins. Thus, photoaffinity labeling with all-trans-retinoic acid and related photoactivatable retinoids is a method that should prove extremely useful in the identification and purification of novel soluble and membrane-associated retinoid-binding proteins from ocular and nonocular tissues. Images Fig. 1 Fig. 2 Fig. 3 Fig. 4 PMID:7846032

  4. Nature and nurture in atherosclerosis: The roles of acylcarnitine and cell membrane-fatty acid intermediates.

    PubMed

    Blair, Harry C; Sepulveda, Jorge; Papachristou, Dionysios J

    2016-03-01

    Macrophages recycle components of dead cells, including cell membranes. When quantities of lipids from cell membranes of dead cells exceed processing capacity, phospholipid and cholesterol debris accumulate as atheromas. Plasma lipid profiles, particularly HDL and LDL cholesterol, are important tools to monitor atherosclerosis risk. Membrane lipids are exported, as triglycerides or phospholipids, or as cholesterol or cholesterol esters, via lipoproteins for disposal, for re-use in cell membranes, or for fat storage. Alternative assays evaluate other aspects of lipid pathology. A key process underlying atherosclerosis is backup of macrophage fatty acid catabolism. This can be quantified by accumulation of acylcarnitine intermediates in extracellular fluid, a direct assay of adequacy of β-oxidation to deal with membrane fatty acid recycling. Further, membranes of somatic cells, such as red blood cells (RBC), incorporate fatty acids that reflect dietary intake. Changes in RBC lipid composition occur within days of ingesting modified fats. Since diets with high saturated fat content or artificial trans-fatty acids promote atherosclerosis, RBC lipid content shifts occur with atherosclerosis, and can show cellular adaptation to pathologically stiff membranes by increased long-chain doubly unsaturated fatty acid production. Additional metabolic changes with atherosclerosis of potential utility include inflammatory cytokine production, modified macrophage signaling pathways, and altered lipid-handling enzymes. Even after atherosclerotic lesions appear, approaches to minimize macrophage overload by reducing rate of fat metabolism are promising. These include preventive measures, and drugs including statins and the newer PCSK9 inhibitors. New cell-based biochemical and cytokine assays provide data to prevent or monitor atherosclerosis progression. PMID:26133667

  5. Fatty acid stimulation of membrane phosphatidylinositol hydrolysis by brain phosphatidylinositol phosphodiesterase.

    PubMed Central

    Irvine, R F; Letcher, A J; Dawson, R M

    1979-01-01

    The hydrolysis of membrane-bound phosphatidylinositol in rat liver microsomal fraction by the soluble phosphatidylinositol phosphodiesterase from rat brain was markedly stimulated by oleic acid or arachidonic acid. The stimulation did not require added calcium, although it was abolished by EDTA. Lysophosphatidylcholine also totally suppressed the stimulation. A possible role for the fatty acid content of a membrane in controlling phosphatidylinositol turnover is suggested. PMID:220968

  6. FAX1, a Novel Membrane Protein Mediating Plastid Fatty Acid Export

    PubMed Central

    Li, Nannan; Gügel, Irene Luise; Giavalisco, Patrick; Zeisler, Viktoria; Schreiber, Lukas; Soll, Jürgen; Philippar, Katrin

    2015-01-01

    Fatty acid synthesis in plants occurs in plastids, and thus, export for subsequent acyl editing and lipid assembly in the cytosol and endoplasmatic reticulum is required. Yet, the transport mechanism for plastid fatty acids still remains enigmatic. We isolated FAX1 (fatty acid export 1), a novel protein, which inserts into the chloroplast inner envelope by α-helical membrane-spanning domains. Detailed phenotypic and ultrastructural analyses of FAX1 mutants in Arabidopsis thaliana showed that FAX1 function is crucial for biomass production, male fertility and synthesis of fatty acid-derived compounds such as lipids, ketone waxes, or pollen cell wall material. Determination of lipid, fatty acid, and wax contents by mass spectrometry revealed that endoplasmatic reticulum (ER)-derived lipids decreased when FAX1 was missing, but levels of several plastid-produced species increased. FAX1 over-expressing lines showed the opposite behavior, including a pronounced increase of triacyglycerol oils in flowers and leaves. Furthermore, the cuticular layer of stems from fax1 knockout lines was specifically reduced in C29 ketone wax compounds. Differential gene expression in FAX1 mutants as determined by DNA microarray analysis confirmed phenotypes and metabolic imbalances. Since in yeast FAX1 could complement for fatty acid transport, we concluded that FAX1 mediates fatty acid export from plastids. In vertebrates, FAX1 relatives are structurally related, mitochondrial membrane proteins of so-far unknown function. Therefore, this protein family might represent a powerful tool not only to increase lipid/biofuel production in plants but also to explore novel transport systems involved in vertebrate fatty acid and lipid metabolism. PMID:25646734

  7. FAX1, a novel membrane protein mediating plastid fatty acid export.

    PubMed

    Li, Nannan; Gügel, Irene Luise; Giavalisco, Patrick; Zeisler, Viktoria; Schreiber, Lukas; Soll, Jürgen; Philippar, Katrin

    2015-02-01

    Fatty acid synthesis in plants occurs in plastids, and thus, export for subsequent acyl editing and lipid assembly in the cytosol and endoplasmatic reticulum is required. Yet, the transport mechanism for plastid fatty acids still remains enigmatic. We isolated FAX1 (fatty acid export 1), a novel protein, which inserts into the chloroplast inner envelope by α-helical membrane-spanning domains. Detailed phenotypic and ultrastructural analyses of FAX1 mutants in Arabidopsis thaliana showed that FAX1 function is crucial for biomass production, male fertility and synthesis of fatty acid-derived compounds such as lipids, ketone waxes, or pollen cell wall material. Determination of lipid, fatty acid, and wax contents by mass spectrometry revealed that endoplasmatic reticulum (ER)-derived lipids decreased when FAX1 was missing, but levels of several plastid-produced species increased. FAX1 over-expressing lines showed the opposite behavior, including a pronounced increase of triacyglycerol oils in flowers and leaves. Furthermore, the cuticular layer of stems from fax1 knockout lines was specifically reduced in C29 ketone wax compounds. Differential gene expression in FAX1 mutants as determined by DNA microarray analysis confirmed phenotypes and metabolic imbalances. Since in yeast FAX1 could complement for fatty acid transport, we concluded that FAX1 mediates fatty acid export from plastids. In vertebrates, FAX1 relatives are structurally related, mitochondrial membrane proteins of so-far unknown function. Therefore, this protein family might represent a powerful tool not only to increase lipid/biofuel production in plants but also to explore novel transport systems involved in vertebrate fatty acid and lipid metabolism. PMID:25646734

  8. Fatty acid composition of plasma lipids and erythrocyte membranes during simulated extravehicular activity

    NASA Astrophysics Data System (ADS)

    Skedina, M. A.; Katuntsev, V. P.; Buravkova, L. B.; Naidina, V. P.

    Ten subjects (from 27 to 41 years) have been participated in 32 experiments. They were decompressed from ground level to 40-35 kPa in altitude chamber when breathed 100% oxygen by mask and performed repeated cycles of exercises (3.0 Kcal/min). The intervals between decompressions were 3-5 days. Plasma lipid and erythrocyte membrane fatty acid composition was evaluated in the fasting venous blood before and immediately after hypobaric exposure. There were 7 cases decompression sickness (DCS). Venous gas bubbles (GB) were detected in 27 cases (84.4%). Any significant changes in the fatty acid composition of erythrocyte membranes and plasma didn't practically induce after the first decompression. However, by the beginning of the second decompression the total lipid level in erythrocyte membranes decreased from 54.6 mg% to 40.4 mg% in group with DCS symptoms and from 51.2 mg% to 35.2 mg% (p < 0.05) without DCS symptoms. In group with DCS symptoms a tendency to increased level of saturated fatty acids in erythrocyte membranes (16:0, 18:0), the level of the polyunsaturated linoleic fatty acid (18:2) and arachidonic acid (20:4) tended to be decreased by the beginning of the second decompression. Insignificant changes in blood plasma fatty acid composition was observed in both groups. The obtained biochemical data that indicated the simulated extravehicular activity (EVA) condition is accompanied by the certain changes in the blood lipid metabolism, structural and functional state of erythrocyte membranes, which are reversible. The most pronounced changes are found in subjects with DCS symptoms.

  9. Altered erythrocyte membrane fatty acid profile in typical Rett syndrome: effects of omega-3 polyunsaturated fatty acid supplementation.

    PubMed

    Signorini, Cinzia; De Felice, Claudio; Leoncini, Silvia; Durand, Thierry; Galano, Jean-Marie; Cortelazzo, Alessio; Zollo, Gloria; Guerranti, Roberto; Gonnelli, Stefano; Caffarelli, Carla; Rossi, Marcello; Pecorelli, Alessandra; Valacchi, Giuseppe; Ciccoli, Lucia; Hayek, Joussef

    2014-11-01

    This study mainly aims at examining the erythrocyte membrane fatty acid (FAs) profile in Rett syndrome (RTT), a genetically determined neurodevelopmental disease. Early reports suggest a beneficial effects of omega-3 polyunsaturated fatty acids (ω-3 PUFAs) on disease severity in RTT. A total of 24 RTT patients were assigned to ω-3 PUFAs-containing fish oil for 12 months in a randomized controlled study (average DHA and EPA doses of 72.9, and 117.1mg/kgb.w./day, respectively). A distinctly altered FAs profile was detectable in RTT, with deficient ω-6 PUFAs, increased saturated FAs and reduced trans 20:4 FAs. FAs changes were found to be related to redox imbalance, subclinical inflammation, and decreased bone density. Supplementation with ω-3 PUFAs led to improved ω-6/ω-3 ratio and serum plasma lipid profile, decreased PUFAs peroxidation end-products, normalization of biochemical markers of inflammation, and reduction of bone hypodensity as compared to the untreated RTT group. Our data indicate that a significant FAs abnormality is detectable in the RTT erythrocyte membranes and is partially rescued by ω-3 PUFAs. PMID:25240461

  10. Effects of bleomycin and antioxidants on the fatty acid profile of testicular cancer cell membranes.

    PubMed

    Cort, A; Ozben, T; Melchiorre, M; Chatgilialoglu, C; Ferreri, C; Sansone, A

    2016-02-01

    Bleomycin is used in chemotherapy regimens for the treatment of patients having testicular germ-cell tumor (TGCT). There is no study in the literature investigating the effects of bleomycin on membrane lipid profile in testicular cancer cells. We investigated membrane fatty acid (FA) profiles isolated, derivatized and analyzed by gas chromatography of NTera-2 testicular cancer cells incubated with bleomycin (Bleo) for 24 h in the absence and presence of N-Acetyl-L-Cysteine (NAC) and curcumin (Cur) as commonly used antioxidant adjuvants. At the same time the MAPK pathway and EGFR levels were followed up. Bleomycin treatment increased significantly saturated fatty acids (SFA) of phospholipids at the expense of monounsaturated (MUFA) and polyunsaturated fatty acids (PUFA). Bleomycin also led to a significant increase in the trans lipid isomers of oleic and arachidonic acids due to its free radical producing effect. Incubation with bleomycin increased the p38 MAPK and JNK levels and downregulated EGFR pathway. Coincubation of bleomycin with NAC reversed effects caused by bleomycin. Our results highlight the important role of membrane fatty acid remodeling occurring during the use of bleomycin and its concurrent use with antioxidants which can adjuvate the cytotoxic effects of the chemotherapeutic agents. PMID:26656160

  11. Does membrane fatty acid composition modulate mitochondrial functions and their thermal sensitivities?

    PubMed

    Lemieux, H; Blier, P U; Tardif, J-C

    2008-01-01

    We investigated the effect of modifying fatty acid modification of heart mitochondrial membranes by dietary intervention on the functions and thermal sensitivity of electron transport system complexes embedded in the inner mitochondrial membrane. Four groups of rats were fed diets differing in their fat (coconut, olive or fish oil) and antioxidant (fish oil with or without probucol) contents. After 16 weeks of feeding, the coconut and olive oil groups had lower long-chain n-3 polyunsaturated fatty acids contents and a lower unsaturation index compared to both fish oil groups. These differences in fatty acid composition were not related to any differences in the mitochondrial respiration rate induced at Complexes I, II or IV, or to differences in their thermal sensitivity. The coconut oil group showed a lower mitochondrial affinity for pyruvate at 5 degrees C (k(mapp)=6.4+/-1.8) compared to any other groups (k(mapp)=3.8+/-0.5; 4.7+/-0.8; 3.6+/-1.1, for olive, fish oil and fish oil and probucol groups, respectively). At least in rat heart, our results do not support a major impact of the fatty acid composition of the mitochondrial membrane on the function of mitochondrial enzymatic complexes or on their temperature sensitivity. PMID:17993286

  12. Fatty acid remodeling by LPCAT3 enriches arachidonate in phospholipid membranes and regulates triglyceride transport

    PubMed Central

    Hashidate-Yoshida, Tomomi; Harayama, Takeshi; Hishikawa, Daisuke; Morimoto, Ryo; Hamano, Fumie; Tokuoka, Suzumi M; Eto, Miki; Tamura-Nakano, Miwa; Yanobu-Takanashi, Rieko; Mukumoto, Yoshiko; Kiyonari, Hiroshi; Okamura, Tadashi; Kita, Yoshihiro; Shindou, Hideo; Shimizu, Takao

    2015-01-01

    Polyunsaturated fatty acids (PUFAs) in phospholipids affect the physical properties of membranes, but it is unclear which biological processes are influenced by their regulation. For example, the functions of membrane arachidonate that are independent of a precursor role for eicosanoid synthesis remain largely unknown. Here, we show that the lack of lysophosphatidylcholine acyltransferase 3 (LPCAT3) leads to drastic reductions in membrane arachidonate levels, and that LPCAT3-deficient mice are neonatally lethal due to an extensive triacylglycerol (TG) accumulation and dysfunction in enterocytes. We found that high levels of PUFAs in membranes enable TGs to locally cluster in high density, and that this clustering promotes efficient TG transfer. We propose a model of local arachidonate enrichment by LPCAT3 to generate a distinct pool of TG in membranes, which is required for normal directionality of TG transfer and lipoprotein assembly in the liver and enterocytes. DOI: http://dx.doi.org/10.7554/eLife.06328.001 PMID:25898003

  13. Fatty acid remodeling by LPCAT3 enriches arachidonate in phospholipid membranes and regulates triglyceride transport.

    PubMed

    Hashidate-Yoshida, Tomomi; Harayama, Takeshi; Hishikawa, Daisuke; Morimoto, Ryo; Hamano, Fumie; Tokuoka, Suzumi M; Eto, Miki; Tamura-Nakano, Miwa; Yanobu-Takanashi, Rieko; Mukumoto, Yoshiko; Kiyonari, Hiroshi; Okamura, Tadashi; Kita, Yoshihiro; Shindou, Hideo; Shimizu, Takao

    2015-01-01

    Polyunsaturated fatty acids (PUFAs) in phospholipids affect the physical properties of membranes, but it is unclear which biological processes are influenced by their regulation. For example, the functions of membrane arachidonate that are independent of a precursor role for eicosanoid synthesis remain largely unknown. Here, we show that the lack of lysophosphatidylcholine acyltransferase 3 (LPCAT3) leads to drastic reductions in membrane arachidonate levels, and that LPCAT3-deficient mice are neonatally lethal due to an extensive triacylglycerol (TG) accumulation and dysfunction in enterocytes. We found that high levels of PUFAs in membranes enable TGs to locally cluster in high density, and that this clustering promotes efficient TG transfer. We propose a model of local arachidonate enrichment by LPCAT3 to generate a distinct pool of TG in membranes, which is required for normal directionality of TG transfer and lipoprotein assembly in the liver and enterocytes. PMID:25898003

  14. Dehydration of multilamellar fatty acid membranes: Towards a computational model of the stratum corneum

    NASA Astrophysics Data System (ADS)

    MacDermaid, Christopher M.; DeVane, Russell H.; Klein, Michael L.; Fiorin, Giacomo

    2014-12-01

    The level of hydration controls the cohesion between apposed lamellae of saturated free fatty acids found in the lipid matrix of stratum corneum, the outermost layer of mammalian skin. This multilamellar lipid matrix is highly impermeable to water and ions, so that the local hydration shell of its fatty acids may not always be in equilibrium with the acidity and relative humidity, which significantly change over a course of days during skin growth. The homeostasis of the stratum corneum at each moment of its growth likely requires a balance between two factors, which affect in opposite ways the diffusion of hydrophilic species through the stratum corneum: (i) an increase in water order as the lipid lamellae come in closer contact, and (ii) a decrease in water order as the fraction of charged fatty acids is lowered by pH. Herein molecular dynamics simulations are employed to estimate the impact of both effects on water molecules confined between lamellae of fatty acids. Under conditions where membrane undulations are energetically favorable, the charged fatty acids are able to sequester cations around points of contact between lamellae that are fully dehydrated, while essentially maintaining a multilamellar structure for the entire system. This observation suggests that the undulations of the fatty acid lamellae control the diffusion of hydrophilic species through the water phase by altering the positional and rotational order of water molecules in the embedded/occluded "droplets."

  15. Fatty acid membrane assembly on coacervate microdroplets as a step towards a hybrid protocell model

    NASA Astrophysics Data System (ADS)

    Dora Tang, T.-Y.; Rohaida Che Hak, C.; Thompson, Alexander J.; Kuimova, Marina K.; Williams, D. S.; Perriman, Adam W.; Mann, Stephen

    2014-06-01

    Mechanisms of prebiotic compartmentalization are central to providing insights into how protocellular systems emerged on the early Earth. Protocell models are based predominantly on the membrane self-assembly of fatty-acid vesicles, although membrane-free scenarios that involve liquid-liquid microphase separation (coacervation) have also been considered. Here we integrate these alternative models of prebiotic compartmentalization and develop a hybrid protocell model based on the spontaneous self-assembly of a continuous fatty-acid membrane at the surface of preformed coacervate microdroplets prepared from cationic peptides/polyelectrolytes and adenosine triphosphate or oligo/polyribonucleotides. We show that the coacervate-supported membrane is multilamellar, and mediates the selective uptake or exclusion of small and large molecules. The coacervate interior can be disassembled without loss of membrane integrity, and fusion and growth of the hybrid protocells can be induced under conditions of high ionic strength. Our results highlight how notions of membrane-mediated compartmentalization, chemical enrichment and internalized structuration can be integrated in protocell models via simple chemical and physical processes.

  16. Fatty acid membrane assembly on coacervate microdroplets as a step towards a hybrid protocell model.

    PubMed

    Dora Tang, T-Y; Rohaida Che Hak, C; Thompson, Alexander J; Kuimova, Marina K; Williams, D S; Perriman, Adam W; Mann, Stephen

    2014-06-01

    Mechanisms of prebiotic compartmentalization are central to providing insights into how protocellular systems emerged on the early Earth. Protocell models are based predominantly on the membrane self-assembly of fatty-acid vesicles, although membrane-free scenarios that involve liquid-liquid microphase separation (coacervation) have also been considered. Here we integrate these alternative models of prebiotic compartmentalization and develop a hybrid protocell model based on the spontaneous self-assembly of a continuous fatty-acid membrane at the surface of preformed coacervate microdroplets prepared from cationic peptides/polyelectrolytes and adenosine triphosphate or oligo/polyribonucleotides. We show that the coacervate-supported membrane is multilamellar, and mediates the selective uptake or exclusion of small and large molecules. The coacervate interior can be disassembled without loss of membrane integrity, and fusion and growth of the hybrid protocells can be induced under conditions of high ionic strength. Our results highlight how notions of membrane-mediated compartmentalization, chemical enrichment and internalized structuration can be integrated in protocell models via simple chemical and physical processes. PMID:24848239

  17. Membrane Stresses Induced by Overproduction of Free Fatty Acids in Escherichia coli.

    SciTech Connect

    Lennen, Rebecca M.; Kruziki, Max A.; Kumar, Kritika; Zinkel, Robert A.; Burnum, Kristin E.; Lipton, Mary S.; Hoover, Spencer W.; Ranatunga, Don Ruwan; Wittkopp, Tyler M.; Marner II, Wesley D.; Pfleger, Brian F.

    2011-11-01

    Microbially produced fatty acids are potential precursors to high energy density biofuels, including alkanes and alkyl ethyl esters by either catalytic conversion of free fatty acids (FFAs) or enzymatic conversions of acyl-acyl carrier protein or acyl-coenzyme A intermediates. Metabolic engineering efforts aimed at overproducing FFAs in Escherichia coli have achieved less than 30% of the maximum theoretical yield on the supplied carbon source. In this work, the viability, morphology, transcript levels, and protein levels of a strain of E. coli that overproduces medium chain length FFAs was compared to an engineered control strain. By early stationary phase, an 85% reduction in viable cell counts and exacerbated loss of inner membrane integrity were observed in the FFA overproducing strain. These effects were enhanced in strains endogenously producing FFAs compared to strains exposed to exogenously fed FFAs. Under two sets of cultivation conditions, long chain unsaturated fatty acid content greatly increased and the expression of genes and proteins required for unsaturated fatty acid biosynthesis were significantly decreased. Membrane stresses were further implicated by increased expression of genes and proteins of the phage shock response, the MarA/Rob/SoxS regulon, and the nuo and cyo operons of aerobic respiration. Gene deletion studies confirmed the importance of the phage shock proteins and Rob for maintaining cell viability, however little to no change in FFA titers was observed after 24 h cultivation. The results of this study serve as a baseline for future targeted attempts to improve FFA yields and titers in E. coli.

  18. Spectrum of Membrane Morphological Responses to Antibacterial Fatty Acids and Related Surfactants.

    PubMed

    Yoon, Bo Kyeong; Jackman, Joshua A; Kim, Min Chul; Cho, Nam-Joon

    2015-09-22

    Medium-chain saturated fatty acids and related compounds (e.g., monoglycerides) represent one class of membrane-active surfactants with antimicrobial properties. Most related studies have been in vitro evaluations of bacterial growth inhibition, and there is limited knowledge about how the compounds in this class destabilize lipid bilayers, which are the purported target within the bacterial cell membrane. Herein, the interaction between three representative compounds in this class and a supported lipid bilayer platform was investigated using quartz crystal microbalance-dissipation and fluorescence microscopy in order to examine membrane destabilization. The three tested compounds were lauric acid, sodium dodecyl sulfate, and glycerol monolaurate. For each compound, we discovered striking differences in the resulting morphological changes of supported lipid bilayers. The experimental trends indicate that the compounds have membrane-disruptive behavior against supported lipid bilayers principally above the respective critical micelle concentration values. The growth inhibition properties of the compounds against standard and methicillin-resistant Staphylococcus aureus bacterial strains were also tested. Taken together, the findings in this work improve our knowledge about how saturated fatty acids and related compounds destabilize lipid bilayers, offering insight into the corresponding molecular mechanisms that lead to membrane morphological responses. PMID:26325618

  19. Envelope Membranes from Spinach Chloroplasts Are a Site of Metabolism of Fatty Acid Hydroperoxides.

    PubMed Central

    Blee, E.; Joyard, J.

    1996-01-01

    Enzymes in envelope membranes from spinach (Spinacia oleracea L.) chloroplasts were found to catalyze the rapid breakdown of fatty acid hydroperoxides. In contrast, no such activities were detected in the stroma or in thylakoids. In preparations of envelope membranes, 9S-hydroperoxy-10(E),12(Z)-octadecadienoic acid, 13S-hydroperoxy-9(Z),11(E)-octadecadienoic acid, or 13S-hydroperoxy-9(Z),11(E),15(Z)-octadecatrienoic acid were transformed at almost the same rates (1-2 [mu]mol min-1 mg-1 protein). The products formed were separated by reversed-phase high-pressure liquid chromatography and further characterized by gas chromatography-mass spectrometry. Fatty acid hydroperoxides were cleaved (a) into aldehydes and oxoacid fragments, corresponding to the functioning of a hydroperoxide lyase, (b) into ketols that were spontaneously formed from allene oxide synthesized by a hydroperoxide dehydratase, (c) into hydroxy compounds synthesized enzymatically by a system that has not yet been characterized, and (d) into oxoenes resulting from the hydroperoxidase activity of a lipoxygenase. Chloroplast envelope membranes therefore contain a whole set of enzymes that catalyze the synthesis of a variety of fatty acid derivatives, some of which may act as regulatory molecules. The results presented demonstrate a new role for the plastid envelope within the plant cell. PMID:12226196

  20. The effect of natural and synthetic fatty acids on membrane structure, microdomain organization, cellular functions and human health.

    PubMed

    Ibarguren, Maitane; López, David J; Escribá, Pablo V

    2014-06-01

    This review deals with the effects of synthetic and natural fatty acids on the biophysical properties of membranes, and on their implication on cell function. Natural fatty acids are constituents of more complex lipids, like triacylglycerides or phospholipids, which are used by cells to store and obtain energy, as well as for structural purposes. Accordingly, natural and synthetic fatty acids may modify the structure of the lipid membrane, altering its microdomain organization and other physical properties, and provoking changes in cell signaling. Therefore, by modulating fatty acids it is possible to regulate the structure of the membrane, influencing the cell processes that are reliant on this structure and potentially reverting pathological cell dysfunctions that may provoke cancer, diabetes, hypertension, Alzheimer's and Parkinson's disease. The so-called Membrane Lipid Therapy offers a strategy to regulate the membrane composition through drug administration, potentially reverting pathological processes by re-adapting cell membrane structure. Certain fatty acids and their synthetic derivatives are described here that may potentially be used in such therapies, where the cell membrane itself can be considered as a target to combat disease. This article is part of a Special Issue entitled: Membrane Structure and Function: Relevance in the Cell's Physiology, Pathology and Therapy. PMID:24388951

  1. Interactions of a very long chain fatty acid with model membranes and serum albumin. Implications for the pathogenesis of adrenoleukodystrophy.

    PubMed Central

    Ho, J K; Moser, H; Kishimoto, Y; Hamilton, J A

    1995-01-01

    Adrenoleukodystrophy (ALD) is an inherited disorder of fatty acid metabolism marked by accumulation of very long chain saturated fatty acids (VLCFA), especially the 26-carbon acid, hexacosanoic acid (HA), in membranes and tissues. We have studied interactions of 13C-enriched HA with model membranes (phospholipid bilayer vesicles) and bovine serum albumin (BSA) by 13C NMR spectroscopy to compare properties of HA with those of typical dietary fatty acids. In phospholipid bilayers the carboxyl group of HA is localized in the aqueous interface, with an apparent pKa (7.4) similar to other fatty acids; the acyl chain must then penetrate very deeply into the membrane. Desorption of HA from vesicles (t1+2 = 3 h) is orders of magnitude slower than shorter chain fatty acids. In mixtures of vesicles and BSA, HA partitions much more favorably to phospholipid bilayers than typical fatty acids. BSA binds a maximum of only 1 mole of HA at one binding site. Calorimetric experiments show strong perturbations of acyl chains of phospholipids by HA. We predict that disruptive effects of VLCFA on cell membrane structure and function may explain the neurological manifestations of ALD patients. These effects will be further amplified by slow desorption of VLCFA from membranes and by the ineffective binding to serum albumin. PMID:7657817

  2. Free fatty acids as a major component of the chlorosulfolipid membrane of Ochromonas danica

    SciTech Connect

    Winicov, I.

    1985-01-01

    This work is an attempt to determine whether or not free fatty acids are components of the natural membrane of Ochromonas danica. If the FFAs were artifacts, they would most likely have been produced during solvent extraction or during the procedure for flagellar detachment. Attempts to denature putative solvent-activated lipase(s) through exposure to boiling isopropanol or by crosslinking the flagella with glutaraldehyde prior to extraction failed to eliminate the free fatty acid fraction, nor to significantly alter its composition. Exposure of flagella to albumin resulted in the net transfer of FFAs to the supernatant phase, showing their presence is not caused by solvent activated lipolysis. Finally levels of labelled free fatty acids failed to rise as a function of time after deflagellation in cells grown in the presence of (10-/sup 14/C)-oleic acid. Acid hydrolysis of the total labelled lipid at elevated temperature increased the percentage of counts occurring as unesterified fatty acids (from 2.6% to 64.8%). This, taken together with a corresponding loss of the more polar labelled material (66.8% to 8.2%) indicates that some esterified lipids were present, but probably not broken down during the isolation procedure.

  3. Temperature-mediated variations in cellular membrane fatty acid composition of Staphylococcus aureus in resistance to pulsed electric fields.

    PubMed

    Wang, Lang-Hong; Wang, Man-Sheng; Zeng, Xin-An; Liu, Zhi-Wei

    2016-08-01

    Effects of growth temperature on cell membrane fatty acid composition, fluidity and lethal and sublethal injury by pulsed electric fields (PEF) in Staphylococcus aureus ATCC 43300 (S. aureus) in the stationary phase were investigated. Analysis of the membrane fatty acids by gas chromatography-mass spectrometry (GC-MS) revealed that branched chain fatty acids (iso C14:0, iso C15:0, anteiso C15:0 and anteiso C17:0) and straight chain fatty acids (C12:0, C14:0, C16:0, C17:0 and C18:0) were primary constituents in the membrane. The S. aureus changed its membrane fatty acid composition and its overall fluidity when exposed to different temperatures. The PEF lethal and sublethal effects were assessed, and results suggested that the degree of inactivation depended on the cell membrane structure, electric field strength and treatment time. The PEF inactivation kinetics including lethal and sublethal injury fractions were fitted with non-linear Weibull distribution, suggesting that inactivation of the first log cycle of S. aureus population was significantly affected by growth temperature, and the membrane of cells became more fluid, and easier to induce electroportion in low temperatures. Moreover, the morphology of S. aureus cells were investigated by electron microscopy, showing that various temperature-modified cells were distorted to differing extents and some even collapsed due to deep irreversible electroporation after PEF treatment. PMID:27155566

  4. Differential stability of photosynthetic membranes and fatty acid composition at elevated temperature in Symbiodinium

    NASA Astrophysics Data System (ADS)

    Díaz-Almeyda, E.; Thomé, P. E.; El Hafidi, M.; Iglesias-Prieto, R.

    2011-03-01

    Coral reefs are threatened by increasing surface seawater temperatures resulting from climate change. Reef-building corals symbiotic with dinoflagellates in the genus Symbiodinium experience dramatic reductions in algal densities when exposed to temperatures above the long-term local summer average, leading to a phenomenon called coral bleaching. Although the temperature-dependent loss in photosynthetic function of the algal symbionts has been widely recognized as one of the early events leading to coral bleaching, there is considerable debate regarding the actual damage site. We have tested the relative thermal stability and composition of membranes in Symbiodinium exposed to high temperature. Our results show that melting curves of photosynthetic membranes from different symbiotic dinoflagellates substantiate a species-specific sensitivity to high temperature, while variations in fatty acid composition under high temperature rather suggest a complex process in which various modifications in lipid composition may be involved. Our results do not support the role of unsaturation of fatty acids of the thylakoid membrane as being mechanistically involved in bleaching nor as being a dependable tool for the diagnosis of thermal susceptibility of symbiotic reef corals.

  5. Membrane Stresses Induced by Overproduction of Free Fatty Acids in Escherichia coli▿†

    PubMed Central

    Lennen, Rebecca M.; Kruziki, Max A.; Kumar, Kritika; Zinkel, Robert A.; Burnum, Kristin E.; Lipton, Mary S.; Hoover, Spencer W.; Ranatunga, Don R.; Wittkopp, Tyler M.; Marner, Wesley D.; Pfleger, Brian F.

    2011-01-01

    Microbially produced fatty acids are potential precursors to high-energy-density biofuels, including alkanes and alkyl ethyl esters, by either catalytic conversion of free fatty acids (FFAs) or enzymatic conversion of acyl-acyl carrier protein or acyl-coenzyme A intermediates. Metabolic engineering efforts aimed at overproducing FFAs in Escherichia coli have achieved less than 30% of the maximum theoretical yield on the supplied carbon source. In this work, the viability, morphology, transcript levels, and protein levels of a strain of E. coli that overproduces medium-chain-length FFAs was compared to an engineered control strain. By early stationary phase, an 85% reduction in viable cell counts and exacerbated loss of inner membrane integrity were observed in the FFA-overproducing strain. These effects were enhanced in strains endogenously producing FFAs compared to strains exposed to exogenously fed FFAs. Under two sets of cultivation conditions, long-chain unsaturated fatty acid content greatly increased, and the expression of genes and proteins required for unsaturated fatty acid biosynthesis were significantly decreased. Membrane stresses were further implicated by increased expression of genes and proteins of the phage shock response, the MarA/Rob/SoxS regulon, and the nuo and cyo operons of aerobic respiration. Gene deletion studies confirmed the importance of the phage shock proteins and Rob for maintaining cell viability; however, little to no change in FFA titer was observed after 24 h of cultivation. The results of this study serve as a baseline for future targeted attempts to improve FFA yields and titers in E. coli. PMID:21948837

  6. Study and comparison of two enzyme membrane reactors for fatty acids and glycerol production

    SciTech Connect

    Molinari, R.; Santoro, M.E.; Drioli, E. . Dept. of Chemical Engineering and Materials Inst. on Membranes and Chemical Reactors-CNR, Arcavacata di Rende )

    1994-11-01

    Two enzyme membrane reactors (EMR), (1) with one substrate (olive oil) in an oil-in-water emulsion (E-EMR) and (2) with two separated liquid phases (oil and water) (TSLP-EMR), have been studied for the conversion of the triglycerides to fatty acids and glycerol. The enzyme was Candida cylindracea lipase confined on the pressurized face or entrapped in the sponge side of capillary ultrafiltration membranes. Two methods for immobilizing the enzyme in the TSLP-EMR were used: ultrafiltration on a virgin membrane and ultrafiltration on glutaraldehyde pretreated membranes. A multiple use of the reactor was obtained immobilizing the enzyme on the membrane preactivated with glutaraldehyde. The TSLP-EMR showed a specific activity of 0.529 mmol/(mg[center dot]h) versus a specific activity of 0.170 mmol/(mg[center dot]h) of the E-EMR. The rate of fatty acid production in the TSLP-EMR was linear with time showing no enzyme deactivation in an operating time of 80 h. The kinetics observed in the two reactors was different: an equilibrium reaction product-inhibited for the E-EMR and an apparent irreversible reaction of zero order for the TSLP-EMR. Taking into account that in the TSLP-EMR, compared to the E-EMR, (1) the specific activity was higher, (2) the specific rate was constant with the time, and (3) the two products were already separated after the reaction, the TSLP-EMR configuration seems the more convenient.

  7. Comparison between omega-3 and omega-6 polyunsaturated fatty acid intakes as assessed by a food frequency questionnaire and erythrocyte membrane fatty acid composition in young children

    PubMed Central

    Orton, Heather D.; Szabo, Nancy J.; Clare-Salzler, Michael; Norris, Jill M.

    2010-01-01

    Objective We conducted a dietary validation study in youth aged 1 to 11 years by comparing dietary intake of omega-3 and omega-6 polyunsaturated fatty acids (PUFA) as assessed by a parent-completed semi-quantitative food frequency questionnaire (FFQ) over time to erythrocyte membrane composition of the same fatty acids. Design The study population included youth aged 1 to 11 years who were participants in the Diabetes Autoimmunity Study in the Young (DAISY), a longitudinal study in Denver, Colorado that is following a cohort of youth at risk for developing Type I diabetes. Four hundred four children who had erythrocyte membrane fatty acid data matched to an FFQ corresponding to the same time frame for a total of 917 visits (matches) were included. PUFA intake was expressed as both g/day (adjusted for total energy) and as percent of total fat intake. We used mixed models to test the association and calculate the correlation between the erythrocyte membrane estimates and PUFA intake using all records of data for each youth. Results Intakes of total omega-3 fatty acids (β=0.52, p<0.0001, ρ=0.23) and marine PUFAs (β=1.62, p<0.0001, ρ=0.42), as a percent of total fat in the diet, were associated with percent of omega-3 and marine PUFAs in the erythrocyte membrane. Intakes of omega-6 PUFAs (β=0.04, p=0.418, ρ=0.05) and arachidonic acid (β=0.31, p=0.774, ρ=0.01) were not associated. Conclusions In these young children, a FFQ using parental report provided estimates of average long-term intakes of marine PUFAs that correlated well with their erythrocyte cell membrane fatty acid status. PMID:17440518

  8. Interactions of fatty acids with phosphatidylethanolamine membranes: X-ray diffraction and molecular dynamics studies

    PubMed Central

    Cordomí, Arnau; Prades, Jesús; Frau, Juan; Vögler, Oliver; Funari, Sérgio S.; Perez, Juan J.; Escribá, Pablo V.; Barceló, Francisca

    2010-01-01

    An experimental and theoretical study on 1,2-dielaidoyl-sn-glycero-3-phosphoethanolamine (DEPE) membranes containing fatty acids (FAs) was performed by means of X-ray diffraction analysis and molecular dynamics (MD) simulations. The study was aimed at understanding the interactions of several structurally related FAs with biomembranes, which is necessary for further rational lipid drug design in membrane-lipid therapy. The main effect of FAs was to promote the formation of a HII phase, despite a stabilization of the coexisting Lα + HII phases. Derivatives of OA exhibited a specific density profile in the direction perpendicular to the bilayer that reflects differences in the relative localization of the carboxylate group within the polar region of the membrane as well as in the degree of membrane penetration of the FA acyl chain. Hydroxyl and methyl substituents at carbon-2 in the FA acyl chain were identified as effective modulators of the position of carboxylate group in the lipid bilayer. Our data highlight the specific potential of each FA in modulating the membrane structure properties. PMID:19965616

  9. Implications of modifying membrane fatty acid composition on membrane oxidation, integrity, and storage viability of freeze-dried probiotic, Lactobacillus acidophilus La-5.

    PubMed

    Hansen, Marie-Louise R W; Petersen, Mikael A; Risbo, Jens; Hümmer, Magdalena; Clausen, Anders

    2015-01-01

    The aim of this study was to investigate the effect of altering the fatty acid profile of the lipid membrane on storage survival of freeze-dried probiotic, Lactobacillus acidophilus La-5, as well as study the membrane integrity and lipid oxidation. The fatty acid composition of the lipid membrane of L. acidophilus La-5 was significantly different upon growth in MRS (containing Tween 80, an oleic acid source), or in MRS with Tween 20 (containing C12:0 and C14:0), linoleic, or linolenic acid supplemented. Bacteria grown in MRS showed the highest storage survival rates. No indications of loss of membrane integrity could be found, and membrane integrity could therefore not be connected with loss of viability. Survival of bacteria grown with linoleic or linolenic acid was more negatively affected by the presence of oxygen, than bacteria grown in MRS or with Tween 20 supplemented. A small, but significant, loss of linolenic acid during storage could be identified, and an increase of volatile secondary oxidation products during storage was found for bacteria grown in MRS, or with linoleic, or linolenic acid supplemented, but not for bacteria grown with Tween 20. Overall, the results indicate that lipid oxidation and loss of membrane integrity are not the only or most important detrimental reactions which can occur during storage. By altering the fatty acid composition, it was also found that properties of oleic acid gave rise to more robust bacteria than more saturated or unsaturated fatty acids did. PMID:25823709

  10. Change in membrane fatty acid compositions and cold-induced responses in chickpea.

    PubMed

    Kazemi Shahandashti, Seyyede Sanam; Maali Amiri, Reza; Zeinali, Hassan; Ramezanpour, Seyyede Sanaz

    2013-02-01

    Plant cells often increase cold tolerance by reprogramming their genes expression which results in adjusted metabolic alternations, a process enhanced under cold acclimation (CA) phase. In present study, we assessed the changes of membrane fatty acid compositions and defense machine (like antioxidative enzymes) along with damage indexes like electrolyte leakage index (ELI) and malondialdehyde (MDA) during CA, cold stress (CS) and recovery (R) phases in chickpea (Cicer arietinum L.). Results showed an increase in unsaturated fatty acids ratio compare to saturated ones which is a sign of cold tolerance especially after CA phase. Antioxidant enzymes had an important role during CA and R phases while CS affected their activity which can be a sign for associating other metabolites or enzymes activities to create cold tolerance in plants. To investigation of enzymes assay under experimental treatments, the expression pattern of some enzymes including superoxide dismutase (sod), catalase (cat) and lipoxygenase (lox) was studied using quantitative real time PCR. LOX activity has shown a bilateral behavior: a positive relation with membrane damage index in CA and an interesting link with double bond index (DBI) in CS indicating probably its role in secondary metabolites like jasmonic acid signaling pathway. It was suggested that increased DBI and low LOX activity under CS could be a reason for plant cold tolerance. PMID:23065233

  11. Determination of the Fatty Acid Content of Biological Membranes: A Highly Versatile GC-MS Experiment

    NASA Astrophysics Data System (ADS)

    Schultz, Emeric; Pugh, Michael Eugene

    2001-07-01

    The experiment involves the GC-MS of fatty acid methyl esters (FAMEs) obtained from bacterial membranes. It takes about 2 h, from cell harvest to injection. This experiment is done in a lab course for non-science majors and in biochemistry. For non-science majors the focus is on GC-MS as a technique for fingerprinting and on the underlying basis of that fingerprinting. In biochemistry the focus is on the composition of membranes and how this changes with temperature--specifically how the ratio of saturated to unsaturated fatty acids changes to maintain constant cell fluidity. Combined with a parallel DNA experiment, the two major types of intermolecular forces important for the structure and function of biomolecules are compared. How this versatile experiment could be adapted in other chemistry courses is presented. The experiment has obvious appeal to biology majors, can be used to develop several important chemistry concepts, involves teamwork, and employs an important instrument. It could be used in the laboratory portion of a course other than biochemistry to fulfill the new ACS biochemistry requirement.

  12. Cell membrane fatty acid changes and desaturase expression of Saccharomyces bayanus exposed to high pressure homogenization in relation to the supplementation of exogenous unsaturated fatty acids

    PubMed Central

    Serrazanetti, Diana I.; Patrignani, Francesca; Russo, Alessandra; Vannini, Lucia; Siroli, Lorenzo; Gardini, Fausto; Lanciotti, Rosalba

    2015-01-01

    Aims: The aim of this work was to study the responses of Saccharomyces bayanus cells exposed to sub-lethal high-pressure homogenization (HPH) and determine whether the plasmatic membrane can sense HPH in the presence, or absence, of exogenous unsaturated fatty acids (UFAs) in the growth medium. Methods and Results: High-pressure homogenization damaged and caused the collapse of cell walls and membranes of a portion of cells; however, HPH did not significantly affect S. bayanus cell viability (less than 0.3 Log CFU ml-1). HPH strongly affected the membrane fatty acid (FA) composition by increasing the percentage of total UFA when compared with saturated fatty acids. The gene expression showed that the transcription of OLE1, ERG3, and ERG11 increased after HPH. The presence of exogenous UFA abolished HPH-induced effects on the OLE1 and ERG3 genes, increased the percentage of membrane lipids and decreased the expression of OLE1 and ERG3 within 30 min of treatment. Conclusion: The results suggest a key role for UFA in the microbial cell response to sub-lethal stress. In addition, these data provide insight into the molecular basis of the response of S. bayanus to this innovative technology. Significance and Impact of the Study: Elucidation of the mechanism of action for sub-lethal HPH will enable the utilization of this technology to modulate the starter performance at the industrial scale. PMID:26528258

  13. Effect of Selection for High Activity-Related Metabolism on Membrane Phospholipid Fatty Acid Composition in Bank Voles.

    PubMed

    Stawski, Clare; Valencak, Teresa G; Ruf, Thomas; Sadowska, Edyta T; Dheyongera, Geoffrey; Rudolf, Agata; Maiti, Uttaran; Koteja, Paweł

    2015-01-01

    Endothermy, high basal metabolic rates (BMRs), and high locomotor-related metabolism were important steps in the evolution of mammals. It has been proposed that the composition of membrane phospholipid fatty acids plays an important role in energy metabolism and exercise muscle physiology. In particular, the membrane pacemaker theory of metabolism suggests that an increase in cell membrane fatty acid unsaturation would result in an increase in BMR. We aimed to determine whether membrane phospholipid fatty acid composition of heart, liver, and gastrocnemius muscles differed between lines of bank voles selected for high swim-induced aerobic metabolism-which also evolved an increased BMR-and unselected control lines. Proportions of fatty acids significantly differed among the organs: liver was the least unsaturated, whereas the gastrocnemius muscles were most unsaturated. However, fatty acid proportions of the heart and liver did not differ significantly between selected and control lines. In gastrocnemius muscles, significant differences between selection directions were found: compared to control lines, membranes of selected voles were richer in saturated C18:0 and unsaturated C18:2n-6 and C18:3n-3, whereas the pattern was reversed for saturated C16:0 and unsaturated C20:4n-6. Neither unsaturation index nor other combined indexes of fatty acid proportions differed between lines. Thus, our results do not support the membrane pacemaker hypothesis. However, the differences between selected and control lines in gastrocnemius muscles reflect chain lengths rather than number of double bonds and are probably related to differences in locomotor activity per se rather than to differences in the basal or routine metabolic rate. PMID:26658414

  14. Survey of Extreme Solvent Tolerance in Gram-Positive Cocci: Membrane Fatty Acid Changes in Staphylococcus haemolyticus Grown in Toluene

    PubMed Central

    Nielsen, Lindsey E.; Kadavy, Dana R.; Rajagopal, Soumitra; Drijber, Rhae; Nickerson, Kenneth W.

    2005-01-01

    We exploited the unique ecological niche of oil fly larval guts to isolate a strain of Staphylococcus haemolyticus which may be the most solvent-tolerant gram-positive bacterium yet described. This organism is able to tolerate 100% toluene, benzene, and p-xylene on plate overlays and saturating levels of these solvents in monophasic liquid cultures. A comparison of membrane fatty acids by gas chromatography after growth in liquid media with and without toluene showed that in cells continuously exposed to solvent the proportion of anteiso fatty acids increased from 25.8 to 33.7% while the proportion of 20:0 straight-chain fatty acids decreased from 19.3 to 10.1%. No changes in the membrane phospholipid composition were noted. Thus, S. haemolyticus alters its membrane fluidity via fatty acid composition to become more fluid when it is exposed to solvent. This response is opposite that commonly found in gram-negative bacteria, which change their fatty acids so that the cytoplasmic membrane is less fluid. Extreme solvent tolerance in S. haemolyticus is not accompanied by abnormal resistance to anionic or cationic detergents. Finally, six strains of Staphylococcus aureus and five strains of Staphylococcus epidermidis, which were not obtained by solvent selection, also exhibited exceptional solvent tolerance. PMID:16151101

  15. Origin of membrane dipole potential: contribution of the phospholipid fatty acid chains.

    PubMed

    Peterson, Uwe; Mannock, David A; Lewis, Ruthven N A H; Pohl, Peter; McElhaney, Ronald N; Pohl, Elena E

    2002-08-01

    The large intrinsic membrane dipole potential, phi(d), is important for protein insertion and functioning as well as for ion transport across natural and model membranes. However, the origin of phi(d) is controversial. From experiments carried out with lipid monolayers, a significant dependence on the fatty acid chain length is suggested, whereas in experiments with lipid bilayers, the contribution of additional -CH(2)-groups seems negligibly small compared with that of the phospholipid carbonyl groups and lipid-bound water molecules. To compare the impact of the -CH(2)-groups of dipalmitoylphosphatidylcholine (DPPC) near and far from the glycerol backbone, we have varied the structure of DPPC by incorporation of sulfur atoms in place of methylene groups in different positions of the fatty acid chain. The phi(d) of symmetric lipid bilayers containing one heteroatom was obtained from the charge relaxation of oppositely charged hydrophobic ions. We have found that the substitution for a S-atom of a -CH(2)-group decreases phi(d). The effect (deltaphi(d) = -22.6 mV) is most pronounced for S-atoms near the lipid head group while a S-atom substitution in the C(13)- or C(14)-position of the hydrocarbon chain does not effect the bilayer dipole potential. Most probably deltaphi(d) does not originate from an altered dipole potential of the acyl chain containing an heteroatom but is mediated by the disruption of chain packing, leading to a decreased density of lipid dipoles in the membrane. PMID:12191841

  16. New insights into the molecular mechanism of intestinal fatty acid absorption

    PubMed Central

    Wang, Tony Y.; Liu, Min; Portincasa, Piero; Wang, David Q.-H.

    2013-01-01

    Background Dietary fat is the most important energy source of all the nutrients. Fatty acids, stored as triacylglycerols in the body, are an important reservoir of stored energy and derive primarily from animal fats and vegetable oils. Design Although the molecular mechanisms for the transport of water-insoluble amphipathic fatty acids across cell membranes have been debated for many years, it is now believed that the dominant means for intestinal fatty acid uptake is via membrane-associated fatty acid-binding proteins, i.e., fatty acid transporters on the apical membrane of enterocytes. Results These findings indicate that intestinal fatty acid absorption is a multistep process that is regulated by multiple genes at the enterocyte level, and intestinal fatty acid absorption efficiency could be determined by factors influencing intraluminal fatty acid molecules across the brush border membrane of enterocytes. To facilitate research on intestinal, hepatic and plasma triacylglycerol metabolism, it is imperative to establish standard protocols for precisely and accurately measuring the efficiency of intestinal fatty acid absorption in humans and animal models. In this review, we will discuss the chemical structure and nomenclature of fatty acids and summarize recent progress in investigating the molecular mechanisms underlying the intestinal absorption of fatty acids, with a particular emphasis on the physical-chemistry of intestinal lipids and the molecular physiology of intestinal fatty acid transporters. Conclusions A better understanding of the molecular mechanism of intestinal fatty acid absorption should lead to novel approaches to the treatment and the prevention of fatty acid-related metabolic diseases that are prevalent worldwide. PMID:24102389

  17. Adaptive modification of membrane phospholipid fatty acid composition and metabolic thermosuppression of brown adipose tissue in heat-acclimated rats

    NASA Astrophysics Data System (ADS)

    Saha, S. K.; Ohno, T.; Tsuchiya, K.; Kuroshima, A.

    Thermogenesis, especially facultative thermogenesis by brown adipose tissue (BAT), is less important in high ambient temperature and the heat-acclimated animals show a lower metabolic rate. Adaptive changes in the metabolic activity of BAT are generally found to be associated with a modification of membrane phospholipid fatty acid composition. However, the effect of heat acclimation on membrane phospholipid fatty acid composition is as yet unknown. In this study, we examined the thermogenic activity and phospholipid fatty acid composition of interscapular BAT from heat-acclimated rats (control: 25+/-1°C, 50% relative humidity and heat acclimation: 32+/-0.5°C, 50% relative humidity). Basal thermogenesis and the total thermogenic capacity after noradrenaline stimulation, as estimated by in vitro oxygen consumption of BAT (measured polarographically using about 1-mm3 tissue blocks), were smaller in the heat-acclimated group than in the control group. There was no difference in the tissue content of phospholipids between the groups when expressed per microgram of DNA. The phospholipid fatty acid composition was analyzed by a capillary gas chromatograph. The state of phospholipid unsaturation, as estimated by the number of double bonds per fatty acid molecule, was similar between the groups. The saturated fatty acid level was higher in the heat-acclimated group. Among the unsaturated fatty acids, heat acclimation decreased docosahexaenoic acid and oleic acid levels, and increased the arachidonic acid level. The tissue level of docosahexaenoic acid correlated with the basal oxygen consumption of BAT (r=0.6, P<0.01) and noradrenaline-stimulated maximum values of oxygen consumption (r=0.5, P<0.05). Our results show that heat acclimation modifies the BAT phospholipid fatty acids, especially the n-3 polyunsaturated fatty acid docosahexaenoic acid, which is possibly involved in the metabolic thermosuppression.

  18. Within and between breed differences in freezing tolerance and plasma membrane fatty acid composition of boar sperm.

    PubMed

    Waterhouse, K E; Hofmo, P O; Tverdal, A; Miller, R R

    2006-05-01

    The response of sperm to cryopreservation and the fertility of frozen-thawed semen varies between species. Besides species differences in sperm physiology, structure and biochemistry, factors such as sperm transport and female reproductive tract anatomy will affect fertility of frozen-thawed semen. Therefore, studying differences in sperm cryotolerance between breeds and individuals instead of between species may reveal sources of variability in sperm cryotolerance. In the present study, the effect of cooling, re-warming and freezing and thawing on plasma membrane and acrosome integrity of sperm within and between Norwegian Landrace and Duroc breeds was studied. Furthermore, the relation between post-thaw survival rate and fatty acid composition of the sperm plasma membranes was investigated. Flow cytometry assessments of plasma membrane and acrosome integrity revealed no significant differences between breeds; however there were significant male-to-male variations within breeds in post-thaw percentages of live sperm (plasma membrane intact). The most abundant fatty acids in the plasma membranes from both breeds were palmitic acid (16:0), stearic acid (18:0), oleic acid (18:1, n-9), docosapentaenoic acid (22:5, n-6) and docosahexaenoic acid (22:6, n-3). The ratio of sigma operator 22:5, n-6 and 22:6, n-3/ sigma operator all other membrane fatty acids was significantly related to survival rate (plasma membrane integrity) of sperm for both Norwegian Landrace (correlation coefficient (r(s)) = 0.64, P < 0.05) and Duroc (r(s) = 0.67, P < 0.05) boars. In conclusion, male-to-male differences in sperm survival rate after freezing and thawing may be partly related to the amount of long-chain polyunsaturated fatty acids in the sperm plasma membranes. PMID:16672353

  19. Modifications in membrane fatty acid composition of Salmonella typhimurium in response to growth conditions and their effect on heat resistance.

    PubMed

    Alvarez-Ordóñez, Avelino; Fernández, Ana; López, Mercedes; Arenas, Ricardo; Bernardo, Ana

    2008-04-30

    The effects of growth temperature (in the range 10-45 degrees C) and acidification up to pH 4.5 of the culture medium (Brain Heart Infusion, BHI) with different organic acids (acetic, citric and lactic) and hydrochloric acid on membrane fatty acid composition and heat resistance of Salmonella typhimurium CECT 443 were studied. The heat resistance was maximal in cells grown at 45 degrees C (cells grown in non-acidified BHI showed a D58-value of 0.90 min) and decreased with decreasing growth temperature up to 10 degrees C (D58-value of 0.09 min). The growth of cells in acidified media caused an increase in their heat resistance. In general, acid adapted cells showed D-values of between 1.5 and 2 times higher than the corresponding for non-acid adapted control cells. This cross-protection response, which has important implications in food processing, was not dependent on the pH value and the acid used to acidify the growth medium. A membrane adaptation corresponding to an increase in the unsaturated to saturated fatty acids ratio (UFA/SFA) and membrane fluidity was observed at low growth temperature. Moreover, the acidification of the growth medium caused a decrease in UFA/SFA ratio and in the C18:1 relative concentration, and an increase in cyclopropane fatty acids (CFA) content mainly due to the increase in cyc19 relative concentration. Thus, acid adapted cells showed CFA levels 1.5 times higher than non-acid adapted control cells. A significant proportion of unsaturated fatty acids were converted to their cyclopropane derivatives during acid adaptation. These changes in membrane fatty acid composition result in cells with decreased membrane fluidity. A clear relation between membrane fatty acid composition and heat resistance was observed. In general, D-values were maximum for cells with low UFA/SFA ratio, and, consequently, with low membrane fluidity. Moreover, CFA formation played a major role in protecting acid adapted cells from heat inactivation. However

  20. Influence of dietary partially hydrogenated fat high in trans fatty acids on lipid composition and function of intestinal brush border membrane in rats.

    PubMed

    Ghafoorunissa, S A.I.

    2001-02-01

    The effect of dietary hydrogenated fat (Indian vanaspati) high in trans fatty acids (6 en%) on lipid composition, fluidity and function of rat intestinal brush border membrane was studied at 2 and 8 en% of linoleic acid. Three groups of weanling rats were fed rice-pulse based diet containing 10% fat over a ten week period: Group I (groundnut oil), Group II (vanaspati), Group III (vanaspati + safflower oil). The functionality of the brush border membrane was assessed by the activity of membrane bound enzymes and transport of D-glucose and L-leucine. The levels of total cholesterol and phospholipids were similar in all groups. The data on fatty acid composition of membrane phospholipids showed that, at 2 en% of linoleic acid in the diet, trans fatty acids lowered arachidonic acid and increased linoleic acid contents indicating altered polyunsaturated fatty acid metabolism. Alkaline phosphatase activity was increased while the activities of sucrase, gamma-glutamyl transpeptidase and transport of D-glucose and L-leucine were not altered by dietary trans fatty acids. However at higher intake of linoleic acid in the diet, trans fatty acids have no effect on polyunsaturated fatty acid composition and alkaline phosphatase activity of intestinal brush border membrane. These data suggest that feeding dietary fat high in trans fatty acids is associated with alteration in intestinal brush border membrane polyunsaturated fatty acid composition and alkaline phosphatase activity only when the dietary linoleic acid is low. PMID:11182555

  1. Gene characterized for membrane desaturase that produces (E)-11 isomers of mono- and diunsaturated fatty acids.

    PubMed

    Liu, Weitian; Jiao, Hongmei; Murray, Nancy C; O'Connor, Marion; Roelofs, Wendell L

    2002-01-22

    Moth species have evolved integral membrane desaturases that exhibit a wide diversity in substrate specificity, as well as in regiospecificity and stereospecificity of the unsaturated products. We report here the cloning and expression of a single desaturase from the sex pheromone gland of the light brown apple moth, Epiphyas postvittana, that makes E11 isomers of monounsaturated (E11-16 and E11-14) fatty acids and a diunsaturated (E9,E11-14) fatty acid. In the pheromone gland, the monoene precursor is made available by beta oxidation of E11-16 acid with a subsequent two-carbon loss to E9-14 acid. A functional assay using a baculovirus expression system required addition of myristic acid and E9-14 acid precursors to demonstrate the unusual regiospecificity and stereospecificity of this desaturase. The amino acid sequence of this desaturase has approximately 61% identity to that of Z11-desaturases from two other insect species, and only approximately 48% identity to the metabolic Z9-desaturases in those species. A pheromone-gland Z9-desaturase gene also was found with the light brown apple moth that differed in its deduced amino acid sequence (66% identity) with the metabolic Z9-desaturase from fat body in this species. PMID:11805319

  2. Dual Role for Phospholipid:Diacylglycerol Acyltransferase: Enhancing Fatty Acid Synthesis and Diverting Fatty Acids from Membrane Lipids to Triacylglycerol in Arabidopsis Leaves[C][W

    PubMed Central

    Fan, Jilian; Yan, Chengshi; Zhang, Xuebin; Xu, Changcheng

    2013-01-01

    There is growing interest in engineering green biomass to expand the production of plant oils as feed and biofuels. Here, we show that PHOSPHOLIPID:DIACYLGLYCEROL ACYLTRANSFERASE1 (PDAT1) is a critical enzyme involved in triacylglycerol (TAG) synthesis in leaves. Overexpression of PDAT1 increases leaf TAG accumulation, leading to oil droplet overexpansion through fusion. Ectopic expression of oleosin promotes the clustering of small oil droplets. Coexpression of PDAT1 with oleosin boosts leaf TAG content by up to 6.4% of the dry weight without affecting membrane lipid composition and plant growth. PDAT1 overexpression stimulates fatty acid synthesis (FAS) and increases fatty acid flux toward the prokaryotic glycerolipid pathway. In the trigalactosyldiacylglycerol1-1 mutant, which is defective in eukaryotic thylakoid lipid synthesis, the combined overexpression of PDAT1 with oleosin increases leaf TAG content to 8.6% of the dry weight and total leaf lipid by fourfold. In the plastidic glycerol-3-phosphate acyltransferase1 mutant, which is defective in the prokaryotic glycerolipid pathway, PDAT1 overexpression enhances TAG content at the expense of thylakoid membrane lipids, leading to defects in chloroplast division and thylakoid biogenesis. Collectively, these results reveal a dual role for PDAT1 in enhancing fatty acid and TAG synthesis in leaves and suggest that increasing FAS is the key to engineering high levels of TAG accumulation in green biomass. PMID:24076979

  3. Ozonolysis products of membrane fatty acids activate eicosanoid metabolism in human airway epithelial cells

    SciTech Connect

    Leikauf, G.D.; Zhao, Q.; Zhou, S.; Santrock, J. )

    1993-12-01

    When inhaled, ozone reacts at the airway luminal surface with unsaturated fatty acids contained in the extracellular fluid and plasma membrane to form an aldehyde and hydroxyhydroperoxide. The resulting hydroxyhydroperoxide degrades in aqueous systems to yield a second aldehyde and hydrogen peroxide (H2O2). Previously, we demonstrated that ozone can augment eicosanoid metabolism in bovine airway epithelial cells. To examine structure-activity relationships of ozone-fatty acid degradation products on eicosanoid metabolism in human airway epithelial cells, 3-, 6-, and 9-carbon saturated aldehydes and hydroxyhydroperoxides were synthesized and purified. Eicosanoid metabolism was evaluated by determination of total 3H-activity release from confluent cells previously incubated with [3H]arachidonic acid and by identification of specific metabolites with high performance liquid chromatography and radioimmunoassay. The major metabolites detected were prostaglandin E2, prostaglandin F2 alpha, and 15-hydroxyeicosatetraenoic acid. The 9-carbon aldehyde, nonanal, in contrast to 3- or 6-carbon aldehydes, stimulated release at concentrations > or = 100 microM, suggesting that the stimulatory effect increases with increasing chain length. When tested under identical conditions, the 3-, 6-, and 9-carbon hydroxyhydroperoxides were more potent than the corresponding aldehydes. Again, a greater effect was noted when the chain length was increased. One possible explanation for the increased potency of the hydroxyhydroperoxides over the aldehydes could be due to degradation of the hydroxyhydroperoxide into H2O2 and aldehyde. We consider this an unlikely explanation because responses varied with chain length (although each hydroxyhydroperoxide would produce an equivalent amount of H2O2) and because exposure to H2O2 alone or H2O2 plus hexanal produced a response dissimilar to 1-hydroxy-1-hexanehydroperoxide.

  4. Effect of unsaturations on the physical properties of a model membrane with the highly polyunsaturated docosahexaenoic fatty acid

    NASA Astrophysics Data System (ADS)

    Saiz, Leonor; Klein, Michael L.

    2001-03-01

    Polyunsaturated fatty acids are an essential component of biomembranes. The docosahexaenoic fatty acid (DHA), in particular, is found in high concentrations in retinal and neuronal tissue and in the olfactory bulb. Furthermore, it is well known the ability of DHA rich membranes to modulate membrane protein function, in some situations, by modifying the membrane physical properties. A particularly well studied situation is the DHA effect onthe activity of the visual receptor (protein) rhodopsin. Here, we study at a microscopic level this type of complex systems under physiological conditions. In this way, we can probe the molecular origin of the peculiarities that the system confers to membranes. To this purpose, the structure of a fully hydrated mixed (saturated/polyunsaturated) chain lipid bilayer in the biologically relevant liquid crystalline phase has been examined by performing molecular dynamics simulations. The model membrane, a 1-stearoyl- 2-docosahexaenoic- sn-glycero- 3-phosphatidylcholine (18:0/22:6 PC) lipid bilayer, was investigated at room temperature and ambient pressure and the results obtained in the nanosecond time scale were in good agreement with the available experimental data. Among the effects of the multiple unsaturations on the physical properties of these membranes, we focus on the enhanced permeability to water and small organic solvents, the decreased area compressibility modulus, and the domain formation and chain segregation.

  5. Identification of Novel Genetic Determinants of Erythrocyte Membrane Fatty Acid Composition among Greenlanders

    PubMed Central

    Sandholt, Camilla Helene; Grarup, Niels; Jørgensen, Marit Eika; Færgeman, Nils Joakim; Bjerregaard, Peter; Pedersen, Oluf; Moltke, Ida; Hansen, Torben; Albrechtsen, Anders

    2016-01-01

    Fatty acids (FAs) are involved in cellular processes important for normal body function, and perturbation of FA balance has been linked to metabolic disturbances, including type 2 diabetes. An individual’s level of FAs is affected by diet, lifestyle, and genetic variation. We aimed to improve the understanding of the mechanisms and pathways involved in regulation of FA tissue levels, by identifying genetic loci associated with inter-individual differences in erythrocyte membrane FA levels. We assessed the levels of 22 FAs in the phospholipid fraction of erythrocyte membranes from 2,626 Greenlanders in relation to single nucleotide polymorphisms genotyped on the MetaboChip or imputed. We identified six independent association signals. Novel loci were identified on chromosomes 5 and 11 showing strongest association with oleic acid (rs76430747 in ACSL6, beta (SE): -0.386% (0.034), p = 1.8x10-28) and docosahexaenoic acid (rs6035106 in DTD1, 0.137% (0.025), p = 6.4x10-8), respectively. For a missense variant (rs80356779) in CPT1A, we identified a number of novel FA associations, the strongest with 11-eicosenoic acid (0.473% (0.035), p = 2.6x10-38), and for variants in FADS2 (rs174570), LPCAT3 (rs2110073), and CERS4 (rs11881630) we replicated known FA associations. Moreover, we observed metabolic implications of the ACSL6 (rs76430747) and CPT1A (rs80356779) variants, which both were associated with altered HbA1c (0.051% (0.013), p = 5.6x10-6 and -0.034% (0.016), p = 3.1x10-4, respectively). The latter variant was also associated with reduced insulin resistance (HOMA-IR, -0.193 (0.050), p = 3.8x10-6), as well as measures of smaller body size, including weight (-2.676 kg (0.523), p = 2.4x10-7), lean mass (-1.200 kg (0.271), p = 1.7x10-6), height (-0.966 cm (0.230), p = 2.0x10-5), and BMI (-0.638 kg/m2 (0.181), p = 2.8x10-4). In conclusion, we have identified novel genetic determinants of FA composition in phospholipids in erythrocyte membranes, and have shown examples of

  6. Identification of Novel Genetic Determinants of Erythrocyte Membrane Fatty Acid Composition among Greenlanders.

    PubMed

    Andersen, Mette Korre; Jørsboe, Emil; Sandholt, Camilla Helene; Grarup, Niels; Jørgensen, Marit Eika; Færgeman, Nils Joakim; Bjerregaard, Peter; Pedersen, Oluf; Moltke, Ida; Hansen, Torben; Albrechtsen, Anders

    2016-06-01

    Fatty acids (FAs) are involved in cellular processes important for normal body function, and perturbation of FA balance has been linked to metabolic disturbances, including type 2 diabetes. An individual's level of FAs is affected by diet, lifestyle, and genetic variation. We aimed to improve the understanding of the mechanisms and pathways involved in regulation of FA tissue levels, by identifying genetic loci associated with inter-individual differences in erythrocyte membrane FA levels. We assessed the levels of 22 FAs in the phospholipid fraction of erythrocyte membranes from 2,626 Greenlanders in relation to single nucleotide polymorphisms genotyped on the MetaboChip or imputed. We identified six independent association signals. Novel loci were identified on chromosomes 5 and 11 showing strongest association with oleic acid (rs76430747 in ACSL6, beta (SE): -0.386% (0.034), p = 1.8x10-28) and docosahexaenoic acid (rs6035106 in DTD1, 0.137% (0.025), p = 6.4x10-8), respectively. For a missense variant (rs80356779) in CPT1A, we identified a number of novel FA associations, the strongest with 11-eicosenoic acid (0.473% (0.035), p = 2.6x10-38), and for variants in FADS2 (rs174570), LPCAT3 (rs2110073), and CERS4 (rs11881630) we replicated known FA associations. Moreover, we observed metabolic implications of the ACSL6 (rs76430747) and CPT1A (rs80356779) variants, which both were associated with altered HbA1c (0.051% (0.013), p = 5.6x10-6 and -0.034% (0.016), p = 3.1x10-4, respectively). The latter variant was also associated with reduced insulin resistance (HOMA-IR, -0.193 (0.050), p = 3.8x10-6), as well as measures of smaller body size, including weight (-2.676 kg (0.523), p = 2.4x10-7), lean mass (-1.200 kg (0.271), p = 1.7x10-6), height (-0.966 cm (0.230), p = 2.0x10-5), and BMI (-0.638 kg/m2 (0.181), p = 2.8x10-4). In conclusion, we have identified novel genetic determinants of FA composition in phospholipids in erythrocyte membranes, and have shown examples of

  7. Derivatives of the cationic plant alkaloids berberine and palmatine amplify protonophorous activity of fatty acids in model membranes and mitochondria.

    PubMed

    Pustovidko, Antonina V; Rokitskaya, Tatiana I; Severina, Inna I; Simonyan, Ruben A; Trendeleva, Tatiana A; Lyamzaev, Konstantin G; Antonenko, Yuri N; Rogov, Anton G; Zvyagilskaya, Renata A; Skulachev, Vladimir P; Chernyak, Boris V

    2013-09-01

    Previously it has been shown by our group that berberine and palmatine, penetrating cations of plant origin, when conjugated with plastoquinone (SkQBerb and SkQPalm), can accumulate in isolated mitochondria or in mitochondria of living cells and effectively protect them from oxidative damage. In the present work, we demonstrate that SkQBerb, SkQPalm, and their analogs lacking the plastoquinone moiety (C10Berb and C10Palm) operate as mitochondria-targeted compounds facilitating protonophorous effect of free fatty acids. These compounds induce proton transport mediated by small concentrations of added fatty acids both in planar and liposomal model lipid membranes. In mitochondria, such an effect can be carried out by endogenous fatty acids and the adenine nucleotide translocase. PMID:23026390

  8. Free fatty acids chain length distribution affects the permeability of skin lipid model membranes.

    PubMed

    Uchiyama, Masayuki; Oguri, Masashi; Mojumdar, Enamul H; Gooris, Gert S; Bouwstra, Joke A

    2016-09-01

    The lipid matrix in the stratum corneum (SC) plays an important role in the barrier function of the skin. The main lipid classes in this lipid matrix are ceramides (CERs), cholesterol (CHOL) and free fatty acids (FFAs). The aim of this study was to determine whether a variation in CER subclass composition and chain length distribution of FFAs affect the permeability of this matrix. To examine this, we make use of lipid model membranes, referred to as stratum corneum substitute (SCS). We prepared SCS containing i) single CER subclass with either a single FFA or a mixture of FFAs and CHOL, or ii) a mixture of various CER subclasses with either a single FFA or a mixture of FFAs and CHOL. In vitro permeation studies were performed using ethyl-p-aminobenzoic acid (E-PABA) as a model drug. The flux of E-PABA across the SCS containing the mixture of FFAs was higher than that across the SCS containing a single FA with a chain length of 24 C atoms (FA C24), while the E-PABA flux was not effected by the CER composition. To select the underlying factors for the changes in permeability, the SCSs were examined by Fourier transform infrared spectroscopy (FTIR) and Small angle X-ray scattering (SAXS). All lipid models demonstrated a similar phase behavior. However, when focusing on the conformational ordering of the individual FFA chains, the shorter chain FFA (with a chain length of 16, 18 or 20 C atoms forming only 11m/m% of the total FFA level) had a higher conformational disordering, while the conformational ordering of the chains of the CER and FA C24 and FA C22 hardly did not change irrespective of the composition of the SCS. In conclusion, the conformational mobility of the short chain FFAs present only at low levels in the model SC lipid membranes has a great impact on the permeability of E-PABA. PMID:27287726

  9. Structural consequences of genetically engineered saturation of the fatty acids of phosphatidylglycerol in tobacco thylakoid membranes. An FTIR study.

    PubMed

    Szalontai, Balázs; Kóta, Zoltán; Nonaka, Hideko; Murata, Norio

    2003-04-15

    The role of phosphatidylglycerol (PG) in protein-lipid interactions and membrane dynamics has been studied in the thylakoids of wild type and manipulated tobacco plants transformed with complementary DNAs for glycerol-3-phosphate acyltransferases (GPATs) from squash and Arabidopsis. The expression of the foreign enzymes resulted in the level of saturation of the PG molecules being higher in the squash and lower in the Arabidopsis transformants, as compared with the level in wild-type tobacco. For the analysis of fatty acyl chain dynamics in the thylakoid membranes, the nu(sym)CH(2) vibration bands of the infrared specta were decomposed into two components, corresponding to ordered and disordered fatty acyl chain segments. With this approach, it was shown that in squash GPAT-transformed tobacco thylakoids a rigid lipid domain exists below 25 degrees C. Above 25 degrees C, the dynamics of all thylakoid membranes were very similar, regardless of the manipulations. PG seems to tune the dynamics at the protein-lipid interface rather than to affect the structure of the proteins directly. Above 50 degrees C, the frequencies of the disordered nu(sym)CH(2) component bands were decreased. This lipid-related phenomenon correlated with protein denaturing. It is demonstrated that the protein aggregation appearing upon heat denaturing changes the conformational distribution of the disordered lipid population. The data also reveal that the protein stability does not depend on the fatty acid composition of the PG molecules; other lipids should provide the environment governing the protein stability in the thylakoid membrane. This is the first such detailed analysis of the infrared spectra of biological membranes that permits a differentiation between structurally different lipid populations within a membrane. PMID:12680783

  10. Emission wavelength-dependent decay of the 9-anthroyloxy-fatty acid membrane probes.

    PubMed Central

    Matayoshi, E D; Kleinfeld, A M

    1981-01-01

    Using the phase-modulation technique, we have measured the fluorescence decay of 2- and 12-(9-anthroyloxy)-stearic acid (2- and 12-AS) and 16-(9-anthroyloxy)-palmitic acid (16-AP) bound to egg phosphatidylcholine vesicles or dissolved in nonpolar solvents. Heterogeneity analysis demonstrates that the decay is generally not monoexponential and exhibits large component variations across it emission spectrum. The mean decay time increases (and in parallel, the steady-state polarization decreases) monotonically with increasing wavelength from values at the blue end. The decay at the red side of the emission spectrum contains an exponential term with a negative amplitude, indicating that emission occurs from intermediates created in the excited-state. This behavior is interpreted as arising from intramolecular fluorophore relaxation occurring on the time scale of the fluorescence lifetime. We believe this to be the first study of wavelength-dependent fluorescent emission which is dominated by an intramolecular relaxation process. Although the three probes exhibit qualitatively similar effects, the emission band variations are greatest for 2-AS and smallest for 16-AP. The differences among the probes are not entirely due to environmental factors as demonstrated, for example, by the emission polarization differences observed in the isotropic solvent paraffin oil. In summary, while these findings point out some of the complexities in the 9-anthroyloxy-fatty acids as membrane probes, they also indicate how these complexities might be used as a sensitive measure of lipid-probe interaction. PMID:7260317

  11. Extracorporeal membrane oxygenation promotes long chain fatty acid oxidation in the immature swine heart in vivo

    SciTech Connect

    Kajimoto, Masaki; O'Kelly-Priddy, Colleen M.; Ledee, Dolena R.; Xu, Chun; Isern, Nancy G.; Olson, Aaron; Portman, Michael A.

    2013-09-01

    Extracorporeal membrane oxygenation (ECMO) supports infants and children with severe cardiopulmonary compromise. Nutritional support for these children includes provision of medium- and long-chain fatty acids (FAs). However, ECMO induces a stress response, which could limit the capacity for FA oxidation. Metabolic impairment could induce new or exacerbate existing myocardial dysfunction. Using a clinically relevant piglet model, we tested the hypothesis that ECMO maintains the myocardial capacity for FA oxidation and preserves myocardial energy state. Provision of 13-Carbon labeled medium-chain FA (octanoate), longchain free FAs (LCFAs), and lactate into systemic circulation showed that ECMO promoted relative increases in myocardial LCFA oxidation while inhibiting lactate oxidation. Loading of these labeled substrates at high dose into the left coronary artery demonstrated metabolic flexibility as the heart preferentially oxidized octanoate. ECMO preserved this octanoate metabolic response, but also promoted LCFA oxidation and inhibited lactate utilization. Rapid upregulation of pyruvate dehydrogenase kinase-4 (PDK4) protein appeared to participate in this metabolic shift during ECMO. ECMO also increased relative flux from lactate to alanine further supporting the role for pyruvate dehydrogenase inhibition by PDK4. High dose substrate loading during ECMO also elevated the myocardial energy state indexed by phosphocreatine to ATP ratio. ECMO promotes LCFA oxidation in immature hearts, while maintaining myocardial energy state. These data support the appropriateness of FA provision during ECMO support for the immature heart.

  12. [Peculiarities of the phospholipid and fatty acid composition of erythrocyte plasma membranes of the Black Sea fish].

    PubMed

    Silkin, Iu A; Silkina, E N; Zabelinskiĭ, S A

    2012-01-01

    The phospholipid and the fatty acid composition of the main phospholipids families of erythrocyte plasma membranes was studied in two species of cartilaginous fish: the common thrasher (Raja clavata L.) and the common stingray (Dasyatis pastinaca) and three bony fish species: the scorpion fish (Scorpaena porcus L.), the smarida (Spicara flexuosa Raf.), and the horse mackerel (Trachurus mediterraneus ponticus Aleev). It was shown that in the studied fish, 70.0-80.0 % of all membrane phospholipids were composed of phosphatidylcholine and phosphatidylethanolamine. Phosphatidylserine, monophosphoinositide, and sphingomyelin were minor components whose content in the erythrocyte membrane fluctuated from 3.0 % to 13.0 %. The fatty acid phospholipids composition was represented by a large specter of acids. From saturated acids, basic for plasma membranes are palmitic (C16: 0) and stearic (C18: 0) acids. From unsaturated acids, the larger part belong to mono-, tetra-, penta-, and hexaenoic acids in fish phospholipids. The calculation of the double bond index and of the unsaturation coefficient showed difference in the deformation ability of erythrocyte membranes of the studied fish. PMID:22567974

  13. Trans Fatty Acid Derived Phospholipids Show Increased Membrane Cholesterol and Reduced Receptor Activation as Compared to Their Cis Analogs

    PubMed Central

    Niu, Shui-Lin; Mitchell, Drake C.; Litman, Burton J.

    2005-01-01

    The consumption of trans fatty acid (TFA) is linked to the elevation of LDL cholesterol and is considered to be a major health risk factor for coronary heart disease. Despite several decades of extensive research on this subject, the underlying mechanism of how TFA modulates serum cholesterol levels remains elusive. In this study, we examined the molecular interaction of TFA-derived phospholipid with cholesterol and the membrane receptor rhodopsin in model membranes. Rhodopsin is a prototypical member of the G-protein coupled receptor family. It has a well-characterized structure and function and serves as a model membrane receptor in this study. Phospholipid–cholesterol affinity was quantified by measuring cholesterol partition coefficients. Phospholipid–receptor interactions were probed by measuring the level of rhodopsin activation. Our study shows that phospholipid derived from TFA had a higher membrane cholesterol affinity than their cis analogues. TFA phospholipid membranes also exhibited a higher acyl chain packing order, which was indicated by the lower acyl chain packing free volume as determined by DPH fluorescence and the higher transition temperature for rhodopsin thermal denaturation. The level of rhodopsin activation was diminished in TFA phospholipids. Since membrane cholesterol level and membrane receptors are involved in the regulation of cholesterol homeostasis, the combination of higher cholesterol content and reduced receptor activation associated with the presence of TFA–phospholipid could be factors contributing to the elevation of LDL cholesterol. PMID:15766276

  14. Partitioning of liquid-ordered/liquid-disordered membrane microdomains induced by the fluidifying effect of 2-hydroxylated fatty acid derivatives.

    PubMed

    Ibarguren, Maitane; López, David J; Encinar, José A; González-Ros, José M; Busquets, Xavier; Escribá, Pablo V

    2013-11-01

    Cellular functions are usually associated with the activity of proteins and nucleic acids. Recent studies have shown that lipids modulate the localization and activity of key membrane-associated signal transduction proteins, thus regulating the cell's physiology. Membrane Lipid Therapy aims to reverse cell dysfunctions (i.e., diseases) by modulating the activity of membrane signaling proteins through regulation of the lipid bilayer structure. The present work shows the ability of a series of 2-hydroxyfatty acid (2OHFA) derivatives, varying in the acyl chain length and degree of unsaturation, to regulate the membrane lipid structure. These molecules have shown greater therapeutic potential than their natural non-hydroxylated counterparts. We demonstrated that both 2OHFA and natural FAs induced reorganization of lipid domains in model membranes of POPC:SM:PE:Cho, modulating the liquid-ordered/liquid-disordered structures ratio and the microdomain lipid composition. Fluorescence spectroscopy, confocal microscopy, Fourier transform infrared spectroscopy and differential detergent solubilization experiments showed a destabilization of the membranes upon addition of the 2OHFAs and FAs which correlated with the observed disordering effect. The changes produced by these synthetic fatty acids on the lipid structure may constitute part of their mechanism of action, leading to changes in the localization/activity of membrane proteins involved in signaling cascades, and therefore modulating cell responses. PMID:23792066

  15. The role of putrescine in the regulation of proteins and fatty acids of thylakoid membranes under salt stress.

    PubMed

    Shu, Sheng; Yuan, Yinghui; Chen, Jie; Sun, Jin; Zhang, Wenhua; Tang, Yuanyuan; Zhong, Min; Guo, Shirong

    2015-01-01

    Polyamines can alleviate the inhibitory effects of salinity on plant growth by regulating photosynthetic efficiency. However, little information is available to explain the specific mechanisms underlying the contribution of polyamines to salt tolerance of the photosynthetic apparatus. Here, we investigated the role of putrescine (Put) on the photosynthetic apparatus of cucumber seedlings under salt stress. We found that NaCl stress resulted in severe ion toxicity and oxidative stress in cucumber chloroplasts. In addition, salinity caused a significant increase in the saturated fatty acid contents of thylakoid membranes. Put altered unsaturated fatty acid content, thereby alleviating the disintegration of thylakoid grana lamellae and reducing the number of plastoglobuli in thylakoid membranes. BN-PAGE revealed Put up-regulated the expression of ATP synthase, CP47, D1, Qb, and psbA proteins and down-regulated CP24, D2, and LHCII type III in NaCl-stressed thylakoid membranes. qRT-PCR analysis of gene expression was used to compare transcript and protein accumulation among 10 candidate proteins. For five of these proteins, induced transcript accumulation was consistent with the pattern of induced protein accumulation. Our results suggest that Put regulates protein expression at transcriptional and translational levels by increasing endogenous polyamines levels in thylakoid membranes, which may stabilise photosynthetic apparatus under salt stress. PMID:26435404

  16. The role of putrescine in the regulation of proteins and fatty acids of thylakoid membranes under salt stress

    PubMed Central

    Shu, Sheng; Yuan, Yinghui; Chen, Jie; Sun, Jin; Zhang, Wenhua; Tang, Yuanyuan; Zhong, Min; Guo, Shirong

    2015-01-01

    Polyamines can alleviate the inhibitory effects of salinity on plant growth by regulating photosynthetic efficiency. However, little information is available to explain the specific mechanisms underlying the contribution of polyamines to salt tolerance of the photosynthetic apparatus. Here, we investigated the role of putrescine (Put) on the photosynthetic apparatus of cucumber seedlings under salt stress. We found that NaCl stress resulted in severe ion toxicity and oxidative stress in cucumber chloroplasts. In addition, salinity caused a significant increase in the saturated fatty acid contents of thylakoid membranes. Put altered unsaturated fatty acid content, thereby alleviating the disintegration of thylakoid grana lamellae and reducing the number of plastoglobuli in thylakoid membranes. BN-PAGE revealed Put up-regulated the expression of ATP synthase, CP47, D1, Qb, and psbA proteins and down-regulated CP24, D2, and LHCII type III in NaCl-stressed thylakoid membranes. qRT-PCR analysis of gene expression was used to compare transcript and protein accumulation among 10 candidate proteins. For five of these proteins, induced transcript accumulation was consistent with the pattern of induced protein accumulation. Our results suggest that Put regulates protein expression at transcriptional and translational levels by increasing endogenous polyamines levels in thylakoid membranes, which may stabilise photosynthetic apparatus under salt stress. PMID:26435404

  17. Membrane fatty acid composition and fluidity are involved in the resistance to freezing of Lactobacillus buchneri R1102 and Bifidobacterium longum R0175

    PubMed Central

    Louesdon, Séverine; Charlot-Rougé, Séverine; Tourdot-Maréchal, Raphaëlle; Bouix, Marielle; Béal, Catherine

    2015-01-01

    Determinations of membrane fatty acid composition and fluidity were used together with acidification activity and viability measurements to characterize the physiological state after freezing of Lactobacillus buchneri R1102 and Bifidobacterium longum R0175 cells harvested in the exponential and stationary growth phases. For both strains, lower membrane fluidity was achieved in cells harvested in the stationary growth phase. This change was linked to a lower unsaturated-to-saturated fatty acid ratio for both strains and a higher cyclic-to-saturated fatty acid ratio for L. buchneri R1102 alone. These membrane properties were linked to survival and to maintenance of acidification activity of the cells after freezing, which differed according to the strain and the growth phase. Survival of B. longum R0175 was increased by 10% in cells with low membrane fluidity and high relative saturated fatty acid contents, without any change in acidification activity. Acidification activity was more degraded (70 min) in L. buchneri R1102 cells displaying low membrane fluidity and high saturated and cyclic fatty acid levels. Finally, this study showed that membrane modifications induced by the growth phase differed among bacterial strains in terms of composition. By lowering membrane fluidity, these modifications could be beneficial for survival of B. longum R0175 during the freezing process but detrimental for maintenance of acidification activity of L. buchneri R1102. PMID:24981007

  18. Transport of endocannabinoids across the plasma membrane and within the cell.

    PubMed

    Fowler, Christopher J

    2013-05-01

    Endocannabinoids are readily accumulated from the extracellular space by cells. Although their uptake properties have the appearance of a process of facilitated diffusion, it is by no means clear as to whether there is a plasma membrane transporter dedicated to this task. Intracellular carrier proteins that shuttle the endocannabinoid anandamide from the plasma membrane to its intracellular targets such as the metabolic enzyme, fatty acid amide hydrolase, have been identified. These include proteins with other primary functions, such as fatty-acid-binding proteins and heat shock protein 70, and possibly a fatty acid amide hydrolase-like anandamide transporter protein. Thus, anandamide uptake can be adequately described as a diffusion process across the plasma membrane followed by intracellular carrier-mediated transport to effector molecules, catabolic enzymes and sequestration sites, although it is recognized that different cells are likely to utilize different mechanisms of endocannabinoid transport depending upon the utility of the endocannabinoid for the cell in question. PMID:23441874

  19. Penetration depth of surfactant peptide KL4 into membranes is determined by fatty acid saturation.

    PubMed

    Antharam, Vijay C; Elliott, Douglas W; Mills, Frank D; Farver, R Suzanne; Sternin, Edward; Long, Joanna R

    2009-05-20

    KL(4) is a 21-residue functional peptide mimic of lung surfactant protein B, an essential protein for lowering surface tension in the alveoli. Its ability to modify lipid properties and restore lung compliance was investigated with circular dichroism, differential scanning calorimetry, and solid-state NMR spectroscopy. KL(4) binds fluid lamellar phase PC/PG lipid membranes and forms an amphipathic helix that alters lipid organization and acyl chain dynamics. The binding and helicity of KL(4) is dependent on the level of monounsaturation in the fatty acid chains. At physiologic temperatures, KL(4) is more peripheral and dynamic in fluid phase POPC/POPG MLVs but is deeply inserted into fluid phase DPPC/POPG vesicles, resulting in immobilization of the peptide. Substantial increases in the acyl chain order are observed in DPPC/POPG lipid vesicles with increasing levels of KL(4), and POPC/POPG lipid vesicles show small decreases in the acyl chain order parameters on addition of KL(4). Additionally, a clear effect of KL(4) on the orientation of the fluid phase PG headgroups is observed, with similar changes in both lipid environments. Near the phase transition temperature of the DPPC/POPG lipid mixtures, which is just below the physiologic temperature of lung surfactant, KL(4) causes phase separation with the DPPC remaining in a gel phase and the POPG partitioned between gel and fluid phases. The ability of KL(4) to differentially partition into lipid lamellae containing varying levels of monounsaturation and subsequent changes in curvature strain suggest a mechanism for peptide-mediated lipid organization and trafficking within the dynamic lung environment. PMID:19450480

  20. Triiodothyronine activates lactate oxidation without impairing fatty acid oxidation and improves weaning from extracorporeal membrane oxygenation

    SciTech Connect

    Kajimoto, Masaki; Ledee, Dolena R.; Xu, Chun; Kajimoto, Hidemi; Isern, Nancy G.; Portman, Michael A.

    2014-01-01

    Background: Extracorporeal membrane oxygenation (ECMO) provides a rescue for children with severe cardiac failure. We previously showed that triiodothyronine (T3) improves cardiac function by modulating pyruvate oxidation during weaning. This study was focused on fatty acid (FA) metabolism modulated by T3 for weaning from ECMO after cardiac injury. Methods: Nineteen immature piglets (9.1-15.3 kg) were separated into 3 groups with ECMO (6.5 hours) and wean: normal circulation (Group-C);transient coronary occlusion (10 minutes) followed by ECMO (Group-IR); and IR with T3 supplementation (Group-IR-T3). 13-Carbon labeled lactate, medium-chain and long-chain FAs were infused as oxidative substrates. Substrate fractional contribution to the citric acid cycle (FC) was analyzed by 13-Carbon nuclear magnetic resonance. Results: ECMO depressed circulating T3 levels to 40% baseline at 4 hours and were restored in Group-IR-T3. Group-IR decreased cardiac power, which was not fully restorable and 2 pigs were lost because of weaning failure. Group-IR also depressed FC-lactate, while the excellent contractile function and energy efficiency in Group-IR-T3 occurred along with a marked FC-lactate increase and [ATP]/[ADP] without either decreasing FC-FAs or elevating myocardial oxygen consumption over Group-C or -IR. Conclusions: T3 releases inhibition of lactate oxidation following ischemia-reperfusion injury without impairing FA oxidation. These findings indicate that T3 depression during ECMO is maladaptive, and that restoring levels improves metabolic flux and enhances contractile function during weaning.

  1. Penetration Depth of Surfactant Peptide KL4 into Membranes Is Determined by Fatty Acid Saturation

    PubMed Central

    Antharam, Vijay C.; Elliott, Douglas W.; Mills, Frank D.; Farver, R. Suzanne; Sternin, Edward; Long, Joanna R.

    2009-01-01

    KL4 is a 21-residue functional peptide mimic of lung surfactant protein B, an essential protein for lowering surface tension in the alveoli. Its ability to modify lipid properties and restore lung compliance was investigated with circular dichroism, differential scanning calorimetry, and solid-state NMR spectroscopy. KL4 binds fluid lamellar phase PC/PG lipid membranes and forms an amphipathic helix that alters lipid organization and acyl chain dynamics. The binding and helicity of KL4 is dependent on the level of monounsaturation in the fatty acid chains. At physiologic temperatures, KL4 is more peripheral and dynamic in fluid phase POPC/POPG MLVs but is deeply inserted into fluid phase DPPC/POPG vesicles, resulting in immobilization of the peptide. Substantial increases in the acyl chain order are observed in DPPC/POPG lipid vesicles with increasing levels of KL4, and POPC/POPG lipid vesicles show small decreases in the acyl chain order parameters on addition of KL4. Additionally, a clear effect of KL4 on the orientation of the fluid phase PG headgroups is observed, with similar changes in both lipid environments. Near the phase transition temperature of the DPPC/POPG lipid mixtures, which is just below the physiologic temperature of lung surfactant, KL4 causes phase separation with the DPPC remaining in a gel phase and the POPG partitioned between gel and fluid phases. The ability of KL4 to differentially partition into lipid lamellae containing varying levels of monounsaturation and subsequent changes in curvature strain suggest a mechanism for peptide-mediated lipid organization and trafficking within the dynamic lung environment. PMID:19450480

  2. Remodeling of the Vibrio cholerae membrane by incorporation of exogenous fatty acids from host and aquatic environments

    PubMed Central

    Giles, David K.; Hankins, Jessica V.; Guan, Ziqiang; Trent, M. Stephen

    2011-01-01

    Summary The Gram-negative bacteria Vibrio cholerae poses significant public health concerns by causing an acute intestinal infection afflicting millions of people each year. V. cholerae motility, as well as virulence factor expression and outer membrane protein production, have been shown to be affected by bile (Childers & Klose, 2007). The current study examines the effects of bile on V. cholerae phospholipids. Bile exposure caused significant alterations to the phospholipid profile of V. cholerae but not of other enteric pathogens. These changes consisted of a quantitative increase and migratory difference in cardiolipin, decreases in phosphatidylglycerol and phosphatidylethanolamine, and the dramatic appearance of an unknown phospholipid determined to be lyso-phosphatidylethanolamine. Major components of bile were not responsible for the observed changes, but long chain polyunsaturated fatty acids, which are minor components of bile, were shown to be incorporated into phospholipids of V. cholerae. Although the bile-induced phospholipid profile was independent of the V. cholerae virulence cascade, we identified another relevant environment in which V. cholerae assimilates unique fatty acids into its membrane phospholipids—marine sediment. Our results suggest that Vibrio species possess unique machinery conferring the ability to take up a wider range of exogenous fatty acids than other enteric bacteria. PMID:21255114

  3. Graphene oxide induces plasma membrane damage, reactive oxygen species accumulation and fatty acid profiles change in Pichia pastoris.

    PubMed

    Zhang, Meng; Yu, Qilin; Liang, Chen; Liu, Zhe; Zhang, Biao; Li, Mingchun

    2016-10-01

    During the past couple of years, graphene nanomaterials were extremely popular among the scientists due to the promising properties in many aspects. Before the materials being well applied, we should first focus on their biosafety and toxicity. In this study, we investigated the toxicity of synthesized graphene oxide (GO) against the model industrial organism Pichia pastoris. We found that the synthesized GO showed dose-dependent toxicity to P. pastoris, through cell membrane damage and intracellular reactive oxygen species (ROS) accumulation. In response to these cell stresses, cells had normal unsaturated fatty acid (UFA) levels but increased contents of polyunsaturated fatty acid (PUFA) with up-regulation of UFA synthesis-related genes on the transcriptional level, which made it overcome the stress under GO attack. Two UFA defective strains (spt23Δ and fad12Δ) were used to demonstrate the results above. Hence, this study suggested a close connection between PUFAs and cell survival against GO. PMID:27376352

  4. Seasonal Assessment of Biomass and Fatty Acid Productivity by Tetraselmis sp. in the Ocean Using Semi-Permeable Membrane Photobioreactors.

    PubMed

    Kim, Z-Hun; Park, Hanwool; Lee, Choul-Gyun

    2016-06-28

    A green microalga, Tetraselmis sp., was cultivated in the coastal seawater of Young-Heung Island using semi-permeable membrane photobioreactors (SPM-PBRs) in different seasons. The microalgae in the SPM-PBRs were able to grow on nutrients diffused into the PBRs from the surrounding seawater through SPMs. The biomass productivity varied depending on the ion permeabilities of the SPMs and environmental conditions, whereas the quality and quantity of fatty acids were constant. The temperature of seawater had a greater influence than solar radiation did on productivity of Tetraselmis sp. in SPM-PBRs. SPM-PBRs could provide technologies for concurrent algal biomass and fatty acids production, and eutrophication reduction in the ocean. PMID:26975763

  5. Cold Shock Response of Bacillus subtilis: Isoleucine-Dependent Switch in the Fatty Acid Branching Pattern for Membrane Adaptation to Low Temperatures†

    PubMed Central

    Klein, Wolfgang; Weber, Michael H. W.; Marahiel, Mohamed A.

    1999-01-01

    Bacillus subtilis has developed sophisticated mechanisms to withstand fluctuations in temperature. Membrane fatty acids are the major determinants for a sufficiently fluid membrane state to ensure the membrane’s function at all temperatures. The fatty acid profile of B. subtilis is characterized by a high content of branched fatty acids irrespective of the growth medium. Here, we report on the importance of isoleucine for B. subtilis to survive cold shock from 37 to 15°C. Cold shock experiments with strain JH642 revealed a cold-protective function for all intermediates of anteiso-branched fatty acid biosynthesis. Metabolites related to iso-branched or straight-chain fatty acid biosynthesis were not protective. Fatty acid profiles of different B. subtilis wild-type strains proved the altered branching pattern by an increase in the anteiso-branched fatty acid content and a concomitant decrease of iso-branched species during cold shock. There were no significant changes in the fatty acid saturation or acyl chain length. The cold-sensitive phenotype of isoleucine-deficient strains in the absence of isoleucine correlated with their inability to synthesize more anteiso-branched fatty acids, as shown by the fatty acid profile. The switch to a fatty acid profile dominated by anteiso-C15:0 and C17:0 at low temperatures and the cold-sensitive phenotype of isoleucine-deficient strains in the absence of isoleucine focused our attention on the critical role of anteiso-branched fatty acids in the growth of B. subtilis in the cold. PMID:10464205

  6. High-resolution structural model of porcine P2 myelin membrane protein with associated fatty acid ligand: fact or artifact?

    PubMed

    Sedzik, Jan; Jastrzebski, Jan Pawel

    2011-06-01

    Myelin membrane is a biological complex of glial cells origin; it is composed of 25% (w/w) proteins and 75% lipids, and more than 300 proteins are associated with central nervous system myelin (for peripheral nervous system myelin, such data are lacking). Myelin plays an important role in maintaining propagation of nerve signals. To uncover the nature of propagation phenomena, it is essential to study biochemistry of myelin proteins and lipids, myelin composition, and myelin structure. Nearly all myelin proteins are like antigens, causing clinically well-defined devastating diseases; multiple sclerosis and Guillain-Barré syndrome are two of them. In this article, a high-resolution study (1.8 Å) of porcine myelin P2 protein is presented. Myelin was purified from porcine intradural spinal roots, which were stored at -80°C for 10 years before myelin and P2 protein were purified (spinal roots were a gift of Prof. Kunio Kitamura, Saitama Medical School). The three-dimensional structural analysis uncovered embedded 18-carbons-long fatty acid. Some speculative interpretation is presented, to uncover how this ligand of fatty acid may form cholesterol ester and stabilize the myelin structure or form simple raft microdomain. Protein crystallography indicates that the ligand may be 18-carbons-long fatty acid. This is unlike previous work with mass spectrometry, in which three ligands were determined. In other protein crystallography-based studies of P2 (bovine), an oleic fatty acid was suggested, but, for recombinant (human) protein, palmitic acid was found. There is no fatty acid ligand in equine P2 protein. PMID:21425316

  7. Effects of cold water swimming on blood rheological properties and composition of fatty acids in erythrocyte membranes of untrained older rats.

    PubMed

    Teległów, Aneta; Dabrowski, Zbigniew; Marchewka, Anna; Tabarowski, Zbigniew; Bilski, Jan; Jaśkiewicz, Jerzy; Gdula-Argasińska, Joanna; Głodzik, Jacek; Lizak, Dorota; Kepińska, Magdalena

    2011-01-01

    This is the first report on the effects of a single bout of swimming to exhaustion in cold water on rat erythrocyte deformability, aggregation and fatty acid composition in erythrocyte membranes. The results indicate that there was a significant decrease in body temperature of experimental rats swimming in water at 4 degrees C and 25 degrees C when compared to the control. Erythrocyte aggregation indices did not change after swimming in water at 4 degrees C whereas erythrocyte deformability increased at shear stress 1,13 [Pa] and 15,96 [Pa]. Physical effort performed in water at 4 degrees C when compared to the control group resulted in an increase in monounsaturated and polyunsaturated n-3 fatty acid content in erythrocyte membranes that influenced the increase in their fluidity and permeability even though that of polyunsaturated n-6 fatty acids decreased. Physical effort performed in 25 degrees C water resulted in an increase in saturated fatty acid content and a decrease in all polyunsaturated fatty acids and polyunsaturated n-6 fatty acids when compared to the control group. Swimming of untrained old rats in cold water affected rheological properties oferythrocytes in a negligible way while changes in the fatty acid composition of erythrocyte membranes were more pronounced. PMID:22195477

  8. Seasonal upregulation of catabolic enzymes and fatty acid transporters in the flight muscle of migrating hoary bats, Lasiurus cinereus.

    PubMed

    McGuire, Liam P; Fenton, M Brock; Guglielmo, Christopher G

    2013-06-01

    The high energy density of fat, and limited capacity for carbohydrate storage suggest that migrating bats should fuel endurance flights with fat, as observed in migrating birds. Yet, cursorial mammals are unable to support high intensity exercise with fat stores. We hypothesized that migratory bats and birds have converged on similar physiological mechanisms to fuel endurance flight with fat. We predicted bats would seasonally upregulate fatty acid transport and oxidation pathways when migration demands were high. We studied seasonal variation in mitochondrial oxidative enzyme activities and fatty acid transport protein expression in the flight muscle of hoary bats (Lasiurus cinereus). Carnitine palmitoyl transferase, 3-hydroxyacyl-CoA dehydrogenase and citrate synthase activity increased during migration. There were no changes in expression of fatty acid translocase or plasma membrane fatty acid binding protein. Heart-type fatty acid binding protein expression increased 5-fold in migrating females, but did not vary seasonally in males. An aerial insectivore lifestyle, and the coincidence of migration and pregnancy may explain differences in transporter expression compared to previously studied birds. Overall, our results are consistent with seasonal upregulation of lipid metabolism and aerobic capacity, and confirm that migration poses distinct physiological challenges for bats. PMID:23545469

  9. Permeation of aldopentoses and nucleosides through fatty acid and phospholipid membranes: implications to the origins of life.

    PubMed

    Wei, Chenyu; Pohorille, Andrew

    2013-02-01

    Permeation of aldopentoses and nucleosides through fatty acid and phospholipid membranes was investigated by way of molecular dynamics simulations. Calculated permeability coefficients of membranes to aldopentoses, which exist predominantly in the pyranose form, are in a very good agreement with experimental results. The unexpected preferential permeation of ribose, compared to its diastereomers, found by Sacerdote and Szostak, is explained in terms of inter- and intramolecular interactions involving hydroxyl groups. In aqueous solution, these groups favor the formation of intermolecular hydrogen bonds with neighboring water molecules. Inside the membrane, however, they form intramolecular hydrogen bonds, which in ribose are arranged in a chain. In its diastereomers this chain is broken, which yields higher free energy barrier to transfer through membranes. Faster permeation of ribose would lead to its preferential accumulation inside cells if sugars were converted sufficiently quickly to nonpermeable derivatives. An estimate for the rate of such reaction was derived. Preferential accumulation of ribose would increase the probability of correct monomers' incorporation during synthesis of nucleic acids inside protocells. The same mechanism does not apply to nucleosides or their activated derivatives because sugars are locked in the furanose form, which contains fewer exocyclic hydroxyl groups than does pyranose. The results of this study underscore concerted early evolution of membranes and the biochemical processes that they encapsulated. PMID:23397957

  10. Development of a submerged anaerobic membrane bioreactor for concurrent extraction of volatile fatty acids and biohydrogen production.

    PubMed

    Trad, Zaineb; Akimbomi, Julius; Vial, Christophe; Larroche, Christian; Taherzadeh, Mohammad J; Fontaine, Jean-Pierre

    2015-11-01

    The aim of this work was to study an externally-submerged membrane bioreactor for the cyclic extraction of volatile fatty acids (VFAs) during anaerobic fermentation, combining the advantages of submerged and external technologies for enhancing biohydrogen (BioH2) production from agrowaste. Mixing and transmembrane pressure (TMP) across a hollow fiber membrane placed in a recirculation loop coupled to a stirred tank were investigated, so that the loop did not significantly modify the hydrodynamic properties in the tank. The fouling mechanism, due to cake layer formation, was reversible. A cleaning procedure based on gas scouring and backwashing with the substrate was defined. Low TMP, 10(4)Pa, was required to achieve a 3Lh(-1)m(-2) critical flux. During fermentation, BioH2 production was shown to restart after removing VFAs with the permeate, so as to enhance simultaneously BioH2 production and the recovery of VFAs as platform molecules. PMID:26253913

  11. Small Mismatches in Fatty Acyl Tail Lengths Can Effect Non Steroidal Anti-Inflammatory Drug Induced Membrane Fusion.

    PubMed

    Majumdar, Anupa; Sarkar, Munna

    2016-06-01

    Biological membranes are made up of a variety of lipids with diverse physicochemical properties. The lipid composition modulates different lipidic parameters, such as hydration, dynamics, lipid packing, curvature strain, etc. Changes in these parameters affect various membrane-mediated processes, such as membrane fusion which is an integral step in many biological processes. Packing defects, which originate either from mismatch in the headgroup region or in the hydrophobic acyl tail region, play a major role in modulating membrane dynamics. In this study, we demonstrate how even a small mismatch in the fatty acyl chain length, achieved by incorporation of low concentrations (up to 30 mol %) of dipalmitoylphosphatidylcholine (DPPC) into dimyristoylphosphatidylcholine (DMPC) small unilamellar vesicles (SUVs), alters several lipidic parameters like packing, dynamics, and headgroup hydration. This in turn affects non steroidal anti-inflammatory drug (NSAID) induced membrane fusion. Dynamic light scattering, differential scanning calorimetry, second-derivative absorption spectrophotometry, and steady-state and time-resolved fluorescence have been used to elucidate the effect of small mismatch in the tails in DMPC/DPPC mixed vesicles and how it modulates membrane fusion induced by the oxicam NSAIDs, meloxicam (Mx), piroxicam (Px), and tenoxicam (Tx). Fusion kinetics was monitored using fluorescence based fusion assays. At low DPPC concentration of 10 mol %, additional fluidization promotes lipid mixing to some extent for Mx, but at higher mol % of DPPC, subsequent increase in rigidity of membrane interior along with increase in headgroup hydration, synergistically inhibits fusion to various extents for the three different drugs, Mx, Px, and Tx. PMID:27153337

  12. NMR studies reveal the role of biomembranes in modulating ligand binding and release by intracellular bile acid binding proteins.

    PubMed

    Pedò, Massimo; Löhr, Frank; D'Onofrio, Mariapina; Assfalg, Michael; Dötsch, Volker; Molinari, Henriette

    2009-12-18

    Bile acid molecules are transferred vectorially between basolateral and apical membranes of hepatocytes and enterocytes in the context of the enterohepatic circulation, a process regulating whole body lipid homeostasis. This work addresses the role of the cytosolic lipid binding proteins in the intracellular transfer of bile acids between different membrane compartments. We present nuclear magnetic resonance (NMR) data describing the ternary system composed of the bile acid binding protein, bile acids, and membrane mimetic systems, such as anionic liposomes. This work provides evidence that the investigated liver bile acid binding protein undergoes association with the anionic membrane and binding-induced partial unfolding. The addition of the physiological ligand to the protein-liposome mixture is capable of modulating this interaction, shifting the equilibrium towards the free folded holo protein. An ensemble of NMR titration experiments, based on nitrogen-15 protein and ligand observation, confirm that the membrane and the ligand establish competing binding equilibria, modulating the cytoplasmic permeability of bile acids. These results support a mechanism of ligand binding and release controlled by the onset of a bile salt concentration gradient within the polarized cell. The location of a specific protein region interacting with liposomes is highlighted. PMID:19836400

  13. Increased Long Chain acyl-Coa Synthetase Activity and Fatty Acid Import Is Linked to Membrane Synthesis for Development of Picornavirus Replication Organelles

    PubMed Central

    Scott, Alison J.; Ford, Lauren A.; Pei, Zhengtong; Watkins, Paul A.; Ernst, Robert K.; Belov, George A.

    2013-01-01

    All positive strand (+RNA) viruses of eukaryotes replicate their genomes in association with membranes. The mechanisms of membrane remodeling in infected cells represent attractive targets for designing future therapeutics, but our understanding of this process is very limited. Elements of autophagy and/or the secretory pathway were proposed to be hijacked for building of picornavirus replication organelles. However, even closely related viruses differ significantly in their requirements for components of these pathways. We demonstrate here that infection with diverse picornaviruses rapidly activates import of long chain fatty acids. While in non-infected cells the imported fatty acids are channeled to lipid droplets, in infected cells the synthesis of neutral lipids is shut down and the fatty acids are utilized in highly up-regulated phosphatidylcholine synthesis. Thus the replication organelles are likely built from de novo synthesized membrane material, rather than from the remodeled pre-existing membranes. We show that activation of fatty acid import is linked to the up-regulation of cellular long chain acyl-CoA synthetase activity and identify the long chain acyl-CoA syntheatse3 (Acsl3) as a novel host factor required for polio replication. Poliovirus protein 2A is required to trigger the activation of import of fatty acids independent of its protease activity. Shift in fatty acid import preferences by infected cells results in synthesis of phosphatidylcholines different from those in uninfected cells, arguing that the viral replication organelles possess unique properties compared to the pre-existing membranes. Our data show how poliovirus can change the overall cellular membrane homeostasis by targeting one critical process. They explain earlier observations of increased phospholipid synthesis in infected cells and suggest a simple model of the structural development of the membranous scaffold of replication complexes of picorna-like viruses, that may be

  14. Proteome and Membrane Fatty Acid Analyses on Oligotropha carboxidovorans OM5 Grown under Chemolithoautotrophic and Heterotrophic Conditions

    PubMed Central

    Paul, Debarati; Kumar, Ranjit; Nanduri, Bindu; French, Todd; Pendarvis, Ken; Brown, Ashli; Lawrence, Mark L.; Burgess, Shane C.

    2011-01-01

    Oligotropha carboxidovorans OM5 T. (DSM 1227, ATCC 49405) is a chemolithoautotrophic bacterium able to utilize CO and H2 to derive energy for fixation of CO2. Thus, it is capable of growth using syngas, which is a mixture of varying amounts of CO and H2 generated by organic waste gasification. O. carboxidovorans is capable also of heterotrophic growth in standard bacteriologic media. Here we characterize how the O. carboxidovorans proteome adapts to different lifestyles of chemolithoautotrophy and heterotrophy. Fatty acid methyl ester (FAME) analysis of O. carboxidovorans grown with acetate or with syngas showed that the bacterium changes membrane fatty acid composition. Quantitative shotgun proteomic analysis of O. carboxidovorans grown in the presence of acetate and syngas showed production of proteins encoded on the megaplasmid for assimilating CO and H2 as well as proteins encoded on the chromosome that might have contributed to fatty acid and acetate metabolism. We found that adaptation to chemolithoautotrophic growth involved adaptations in cell envelope, oxidative homeostasis, and metabolic pathways such as glyoxylate shunt and amino acid/cofactor biosynthetic enzymes. PMID:21386900

  15. Response of Halomonas campisalis to saline stress: changes in growth kinetics, compatible solute production and membrane phospholipid fatty acid composition.

    PubMed

    Aston, John E; Peyton, Brent M

    2007-09-01

    The haloalkaliphile Halomonas campisalis, isolated near Soap Lake, Washington, was grown under both aerobic and denitrifying conditions from 0 to 260 g L(-1) NaCl, with optimal growth occurring at 20 and 30 g L(-1) NaCl, respectively. Halomonas campisalis was observed to produce high concentrations of compatible solutes, most notably ectoine (up to 500 mM within the cytoplasm), but hydroxyectoine and glycine betaine were also detected. The types and amounts of compatible solutes produced depended on salinity and specific growth rate, as well as on the terminal electron acceptor available (O(2) or NO(3) (-)). A decrease in ectoine production was observed with NO(3) (-) as compared with O(2) as the terminal electron acceptor. In addition, changes in the phospholipid fatty acid composition were measured with changing salinity. An increase in trans fatty acids was observed in the absence of salinity, and may be a response to membrane instability. Cyclic fatty acids were also observed to increase, both in the absence of salinity, and at very high salinities, indicating cell stress at these conditions. PMID:17651393

  16. Effect of acetaldehyde generated from ethanol by ADH-transfected CHO cells on their membrane fatty acid profiles.

    PubMed

    Meskar, A; Holownia, A; Bardou, L G; Menez, J F

    1996-01-01

    Ethanol has been previously shown to reduce the unsaturated fatty acid content of cell membranes. It is not known, however, if the observed deleterious effects are due to ethanol itself or its metabolite, acetaldehyde. The present study was undertaken to assess the effect of acetaldehyde produced from ethanol by alcohol-deyhdrogenase-transfected Chinese hamster ovary Cells on the membrane lipids and the lipid peroxidation measured by free and bound malondialdehyde (MDA). The effects of ethanol alone was assessed in the presence of 4-methylpyrazole (4-MP), an inhibitor of alcohol dehydrogenase. After 8 days of incubation, total cellular lipids were extracted, subjected to TLC, and analyzed by gas chromatography. MDA concentration were determined by thiobarbituric acid reaction followed by HPLC detection. The level of acetaldehyde in the culture medium increased with concentration of ethanol from 5 to 20 mM as did the lipid peroxidation. Total cholesterol, phospholipids, and triglycerids all increased with increasing concentration of acetaldehyde. These effects were due to acetaldehyde as they were blocked by 4-MP. Some changes in fatty acid profiles were observed by effect of ethanol itself. PMID:8949957

  17. Membrane omega-3 fatty acids modulate the oligomerisation kinetics of adenosine A2A and dopamine D2 receptors

    PubMed Central

    Guixà-González, Ramon; Javanainen, Matti; Gómez-Soler, Maricel; Cordobilla, Begoña; Domingo, Joan Carles; Sanz, Ferran; Pastor, Manuel; Ciruela, Francisco; Martinez-Seara, Hector; Selent, Jana

    2016-01-01

    Membrane levels of docosahexaenoic acid (DHA), an essential omega-3 polyunsaturated fatty acid (ω-3 PUFA), are decreased in common neuropsychiatric disorders. DHA modulates key cell membrane properties like fluidity, thereby affecting the behaviour of transmembrane proteins like G protein-coupled receptors (GPCRs). These receptors, which have special relevance for major neuropsychiatric disorders have recently been shown to form dimers or higher order oligomers, and evidence suggests that DHA levels affect GPCR function by modulating oligomerisation. In this study, we assessed the effect of membrane DHA content on the formation of a class of protein complexes with particular relevance for brain disease: adenosine A2A and dopamine D2 receptor oligomers. Using extensive multiscale computer modelling, we find a marked propensity of DHA for interaction with both A2A and D2 receptors, which leads to an increased rate of receptor oligomerisation. Bioluminescence resonance energy transfer (BRET) experiments performed on living cells suggest that this DHA effect on the oligomerisation of A2A and D2 receptors is purely kinetic. This work reveals for the first time that membrane ω-3 PUFAs play a key role in GPCR oligomerisation kinetics, which may have important implications for neuropsychiatric conditions like schizophrenia or Parkinson’s disease. PMID:26796668

  18. Membrane omega-3 fatty acids modulate the oligomerisation kinetics of adenosine A2A and dopamine D2 receptors

    NASA Astrophysics Data System (ADS)

    Guixà-González, Ramon; Javanainen, Matti; Gómez-Soler, Maricel; Cordobilla, Begoña; Domingo, Joan Carles; Sanz, Ferran; Pastor, Manuel; Ciruela, Francisco; Martinez-Seara, Hector; Selent, Jana

    2016-01-01

    Membrane levels of docosahexaenoic acid (DHA), an essential omega-3 polyunsaturated fatty acid (ω-3 PUFA), are decreased in common neuropsychiatric disorders. DHA modulates key cell membrane properties like fluidity, thereby affecting the behaviour of transmembrane proteins like G protein-coupled receptors (GPCRs). These receptors, which have special relevance for major neuropsychiatric disorders have recently been shown to form dimers or higher order oligomers, and evidence suggests that DHA levels affect GPCR function by modulating oligomerisation. In this study, we assessed the effect of membrane DHA content on the formation of a class of protein complexes with particular relevance for brain disease: adenosine A2A and dopamine D2 receptor oligomers. Using extensive multiscale computer modelling, we find a marked propensity of DHA for interaction with both A2A and D2 receptors, which leads to an increased rate of receptor oligomerisation. Bioluminescence resonance energy transfer (BRET) experiments performed on living cells suggest that this DHA effect on the oligomerisation of A2A and D2 receptors is purely kinetic. This work reveals for the first time that membrane ω-3 PUFAs play a key role in GPCR oligomerisation kinetics, which may have important implications for neuropsychiatric conditions like schizophrenia or Parkinson’s disease.

  19. Decreased Membrane Integrity in Aging Typha latifolia L.Pollen (Accumulation of Lysolipids and Free Fatty Acids).

    PubMed Central

    Van Bilsen, DGJL.; Hoekstra, F. A.

    1993-01-01

    Aging of cattail (Typha latifolia L.) pollen was studied at 24[deg]C under conditions of 40 and 75% relative humidity (RH). The decline of viability coincides with increased leakage at imbibition; both processes develop much faster at the higher humidity condition. During aging phospholipids are deesterified and free fatty acids (FFAs) and lysophospholipids (LPLs) accumulate, again, much more rapidly at 75% RH than at 40% RH. The fatty acid composition of the remaining phospholipids hardly changes during aging, which suggests limited involvement of lipid peroxidation in the degradation process. Tests with phospholipase A2 revealed that the saturated fatty acids occur at the sn-1 position of the glycerol backbone of the phospholipids. The fatty acid composition of the LPLs is similar to that of the phospholipids from which they were formed, indicating that the deesterification occurs at random. This favors involvement of free radicals instead of phospholipases in the deesterification process. Liposome studies were carried out to characterize components in the lipid fraction that might account for the leakage associated with aging. Entrapped carboxyfluorescein leaked much more from liposomes when they were partly made up from total lipids from aged pollen than from nonaged pollen. The components causing the leakage were found in both the polar and the neutral lipid fractions. Further purification and subsequent interchanging of the FFAs and LPLs between extracts from aged and nonaged pollen revealed that in neutral lipid extracts the FFAs are entirely responsible for the leakage, whereas in the phospholipid fraction the LPLs are largely responsible for the leakage. The leakage from the liposomes is not caused by fusion. We suggest that the observed loss of viability and increased leakage during aging are due to the nonenzymic accumulation of FFAs and LPLs in the pollen membranes. PMID:12231723

  20. Dietary fatty acids modulate associations between genetic variants and circulating fatty acids in plasma and erythrocyte membranes: meta-analysis of 9 studies in the CHARGE consortium

    PubMed Central

    Smith, Caren E.; Follis, Jack L.; Nettleton, Jennifer A.; Foy, Millennia; Wu, Jason H.Y.; Ma, Yiyi; Tanaka, Toshiko; Manichakul, Ani W.; Wu, Hongyu; Chu, Audrey Y.; Steffen, Lyn M.; Fornage, Myriam; Mozaffarian, Dariush; Kabagambe, Edmond K.; Ferruci, Luigi; da Chen, Yii-Der I; Rich, Stephen S.; Djoussé, Luc; Ridker, Paul M.; Tang, Weihong; McKnight, Barbara; Tsai, Michael Y.; Bandinelli, Stefania; Rotter, Jerome I.; Hu, Frank B.; Chasman, Daniel I.; Psaty, Bruce M.; Arnett, Donna K.; King, Irena B.; Sun, Qi; Wang, Lu; Lumley, Thomas; Chiuve, Stephanie E.; Siscovick, David S; Ordovás, José M.; Lemaitre, Rozenn N.

    2015-01-01

    Scope Tissue concentrations of omega-3 fatty acids may reduce cardiovascular disease risk, and genetic variants are associated with circulating fatty acids concentrations. Whether dietary fatty acids interact with genetic variants to modify circulating omega-3 fatty acids is unclear. Objective We evaluated interactions between genetic variants and fatty acid intakes for circulating alpha-linoleic acid (ALA), eicosapentaenoic acid (EPA), docosahexaenoic acid (DHA) and docosapentaenoic acid (DPA). Methods and Results We conducted meta-analyses (N to 11,668) evaluating interactions between dietary fatty acids and genetic variants (rs174538 and rs174548 in FADS1 (fatty acid desaturase 1), rs7435 in AGPAT3 (1-acyl-sn-glycerol-3-phosphate), rs4985167 in PDXDC1 (pyridoxal-dependent decarboxylase domain-containing 1), rs780094 in GCKR (glucokinase regulatory protein) and rs3734398 in ELOVL2 (fatty acid elongase 2)). Stratification by measurement compartment (plasma vs. erthyrocyte) revealed compartment-specific interactions between FADS1 rs174538 and rs174548 and dietary ALA and linoleic acid for DHA and DPA. Conclusion Our findings reinforce earlier reports that genetically-based differences in circulating fatty acids may be partially due to differences in the conversion of fatty acid precursors. Further, fatty acids measurement compartment may modify gene-diet relationships, and considering compartment may improve the detection of gene-fatty acids interactions for circulating fatty acid outcomes. PMID:25626431

  1. Retinoic acid binding protein in normal and neopolastic rat prostate.

    PubMed

    Gesell, M S; Brandes, M J; Arnold, E A; Isaacs, J T; Ueda, H; Millan, J C; Brandes, D

    1982-01-01

    Sucrose density gradient analysis of cytosol from normal and neoplastic rat prostatic tissues exhibited a peak of (3H) retinoic acid binding in the 2S region, corresponding to the cytoplasmic retinoic acid binding protein (cRABP). In the Fisher-Copenhagen F1 rat, cRABP was present in the lateral lobe, but could not be detected in the ventral nor in the dorsal prostatic lobes. Four sublines of the R-3327 rat prostatic tumor contained similar levels of this binding protein. The absence of cRABP in the normal tissue of origin of the R-3327 tumor, the rat dorsal prostate, and reappearance in the neoplastic tissues follows a pattern described in other human and animal tumors. The occurrence of cRABP in the well-differentiated as well as in the anaplastic R-3327 tumors in which markers which reflect a state of differentiation and hormonal regulation, such as androgen receptor, 5 alpha reductase, and secretory acid phosphatase are either markedly reduced or absent, points to cRABP as a marker of malignant transformation. PMID:6283503

  2. Folic acid binds DNA and RNA at different locations.

    PubMed

    Bourassa, P; Tajmir-Riahi, H A

    2015-03-01

    We located multiple binding sites for folic acid on DNA and tRNA at physiological conditions, using FTIR, CD, fluorescence spectroscopic methods and molecular modeling. Structural analysis revealed that folic acid binds DNA and tRNA at multiple sites via hydrophilic, hydrophobic and H-bonding contacts with overall binding constants of Kfolic acid-DNA=1.1 (±0.3)×10(4) M(-1) and Kfolic acid-tRNA=6.4 (±0.5)×10(3) M(-1). Molecular modeling showed the participation of several nucleobases in folic acid complexes with DNA and tRNA, stabilized by H-bonding network. Two types of complexes were located for folic acid-tRNA adducts, one at the major groove and the other with TΨC loop, while acid binding occurs at major and minor grooves of DNA duplex. Folic acid complexation induced more alterations of DNA structure than tRNA. PMID:25555838

  3. Cell membrane fatty acid composition differs between normal and malignant cell lines.

    PubMed

    Meng, Xialong; Riordan, Neil H; Riordan, Hugh D; Mikirova, Nina; Jackson, James; González, Michael J; Miranda-Massari, Jorge R; Mora, Edna; Trinidad Castillo, Waleska

    2004-06-01

    Twenty-eight fatty acids (C8:0 to C24:l n-9) were measured by gas chromatography in four normal cell lines (C3H / 10T1 / 2, CCD-18Co, CCD-25SK and CCD-37Lu) and seven cancer cell lines (C-41, Caov-3, LS-180, PC-3, SK-MEL-28, SK-MES-1 and U-87 MG). Results show differences in the content and proportions of fatty acids when comparing cancer cell lines with their normal counterparts. Cancer cell lines showed lower C20: 4 n-6, C24:1 n-9, polyunsaturated fatty acids (PUFA's) and ratios of C20:4 n-6 to C20:5 n-3 and C16:0 to C18:1 n-9 and stearic to oleic (SA/OA) than their normal counterparts. All cancer cell lines had SA/OA ratios lower than 7.0 while normal cell lines had ratios greater than 0.7 (p<0.05). In addition, the ratios of total saturated fatty acids (SFA) to PUFA'S and the concentration of C18:1 n-9, C18:2 n-6, C20:5 n-3 were higher in cancer cell lines as compared to normal cell lines. A positive correlation was detected between C16:0 and longer SFA'S (r = +0.511, p<0.05) in normal cell lines whereas a negative correlation (r=0.608, p<0.05) was obtained for malignant cell lines. Moreover, cancerous cell lines exhibited a particular desaturation defect and an abnormal incorporation of C18:2 n-6 and C20-4 n-6 fatty acids. PMID:15377057

  4. Adaptation of anaerobically grown Thauera aromatica, Geobacter sulfurreducens and Desulfococcus multivorans to organic solvents on the level of membrane fatty acid composition

    PubMed Central

    Duldhardt, Ilka; Gaebel, Julia; Chrzanowski, Lukasz; Nijenhuis, Ivonne; Härtig, Claus; Schauer, Frieder; Heipieper, Hermann J.

    2010-01-01

    Summary The effect of different solvents and pollutants on the cellular fatty acid composition of three bacterial strains: Thauera aromatica, Geobacter sulfurreducens and Desulfococcus multivorans, representatives of diverse predominant anaerobic metabolisms was investigated. As the prevailing adaptive mechanism in cells of T. aromatica and G. sulfurreducens whose cellular fatty acids patterns were dominated by palmitic acid (C16:0) and palmitoleic acid (C16:1cis), the cells reacted by an increase in the degree of saturation of their membrane fatty acids when grown in the presence of sublethal concentrations of the chemicals. Next to palmitic acid C16:0, the fatty acid pattern of D. multivorans was dominated by anteiso‐branched fatty acids which are characteristic for several sulfate‐reducing bacteria. The cells responded to the solvents with an increase in the ratio of straight‐chain saturated (C14:0, C16:0, C18:0) to anteiso‐branched fatty acids (C15:0anteiso, C17:0anteiso, C17:1anteisoΔ9cis). The results show that anaerobic bacteria react with similar mechanisms like aerobic bacteria in order to adapt their membrane to toxic organic solvents. The observed adaptive modifications on the level of membrane fatty acid composition can only be carried out with de novo synthesis of the fatty acids which is strictly related to cell growth. As the growth rates of anaerobic bacteria are generally much lower than in the so far investigated aerobic bacteria, this adaptive response needs more time in anaerobic bacteria. This might be one explanation for the previously observed higher sensitivity of anaerobic bacteria when compared with aerobic ones. PMID:21255320

  5. Adaptation of anaerobically grown Thauera aromatica, Geobacter sulfurreducens and Desulfococcus multivorans to organic solvents on the level of membrane fatty acid composition.

    PubMed

    Duldhardt, Ilka; Gaebel, Julia; Chrzanowski, Lukasz; Nijenhuis, Ivonne; Härtig, Claus; Schauer, Frieder; Heipieper, Hermann J

    2010-03-01

    The effect of different solvents and pollutants on the cellular fatty acid composition of three bacterial strains: Thauera aromatica, Geobacter sulfurreducens and Desulfococcus multivorans, representatives of diverse predominant anaerobic metabolisms was investigated. As the prevailing adaptive mechanism in cells of T. aromatica and G. sulfurreducens whose cellular fatty acids patterns were dominated by palmitic acid (C16:0) and palmitoleic acid (C16:1cis), the cells reacted by an increase in the degree of saturation of their membrane fatty acids when grown in the presence of sublethal concentrations of the chemicals. Next to palmitic acid C16:0, the fatty acid pattern of D. multivorans was dominated by anteiso-branched fatty acids which are characteristic for several sulfate-reducing bacteria. The cells responded to the solvents with an increase in the ratio of straight-chain saturated (C14:0, C16:0, C18:0) to anteiso-branched fatty acids (C15:0anteiso, C17:0anteiso, C17:1anteisoΔ9cis). The results show that anaerobic bacteria react with similar mechanisms like aerobic bacteria in order to adapt their membrane to toxic organic solvents. The observed adaptive modifications on the level of membrane fatty acid composition can only be carried out with de novo synthesis of the fatty acids which is strictly related to cell growth. As the growth rates of anaerobic bacteria are generally much lower than in the so far investigated aerobic bacteria, this adaptive response needs more time in anaerobic bacteria. This might be one explanation for the previously observed higher sensitivity of anaerobic bacteria when compared with aerobic ones. PMID:21255320

  6. Efficient Targeting of Fatty-Acid Modified Oligonucleotides to Live Cell Membranes through Stepwise Assembly

    PubMed Central

    2014-01-01

    Lipid modifications provide efficient targeting of oligonucleotides to live cell membranes in a range of applications. Targeting efficiency is a function of the rate of lipid DNA insertion into the cell surface and its persistence over time. Here we show that increasing lipid hydrophobicity increases membrane persistence, but decreases the rate of membrane insertion due to the formation of nonproductive aggregates in solution. To ameliorate this effect, we split the net hydrophobicity of the membrane anchor between two complementary oligonucleotides. When prehybridized in solution, doubly anchored molecules also aggregate due to their elevated hydrophobicity. However, when added sequentially to cells, aggregation does not occur so membrane insertion is efficient. Hybridization between the two strands locks the complexes at the cell surface by increasing net hydrophobicity, increasing their total concentration and lifetime, and dramatically improving their utility in a variety of biomedical applications. PMID:25325667

  7. 2-Hydroxy Fatty Acid Enantiomers of Gb3 Impact Shiga Toxin Binding and Membrane Organization.

    PubMed

    Schütte, Ole M; Patalag, Lukas J; Weber, Lucas M C; Ries, Annika; Römer, Winfried; Werz, Daniel B; Steinem, Claudia

    2015-06-16

    Shiga toxin subunit B (STxB) binding to its cellular receptor Gb3 leads to the formation of protein-lipid clusters and bending of the membrane. A newly developed synthetic route allowed synthesizing the biologically most relevant Gb3-C24:1 2OH species with both, the natural (Gb3-R) as well as the unnatural (Gb3-S) configuration of the 2OH group. The derivatives bind STxB with identical nanomolar affinity, while the propensity to induce membrane tubules in giant unilamellar vesicles is more pronounced for Gb3-S. Fluorescence and atomic force microscopy images of phase-separated supported membranes revealed differences in the lateral organization of the protein on the membrane. Gb3-R favorably induces large and tightly packed protein clusters, while a lower protein density is found on Gb3-S doped membranes. PMID:26083916

  8. Cholesterol Asymmetry in Synaptic Plasma Membranes

    PubMed Central

    Wood, W. Gibson; Igbavboa, Urule; Müller, Walter E.; Eckert, Gunter P.

    2010-01-01

    Lipids are essential for the structural and functional integrity of membranes. Membrane lipids are not randomly distributed but are localized in different domains. A common characteristic of these membrane domains is their association with cholesterol. Lipid rafts and caveolae are examples of cholesterol enriched domains, which have attracted keen interest. However, two other important cholesterol domains are the exofacial and cytofacial leaflets of the plasma membrane. The two leaflets that make up the bilayer differ in their fluidity, electrical charge, lipid distribution, and active sites of certain proteins. The synaptic plasma membrane (SPM) cytofacial leaflet contains over 85% of the total SPM cholesterol as compared with the exofacial leaflet. This asymmetric distribution of cholesterol is not fixed or immobile but can be modified by different conditions in vivo: 1) chronic ethanol consumption; 2) statins; 3) aging; and 4) apoE isoform. Several potential candidates have been proposed as mechanisms involved in regulation of SPM cholesterol asymmetry: apoE, low-density-lipoprotein receptor, sterol carrier protein-2, fatty acid binding proteins, polyunsaturated fatty acids, p-glycoprotein and caveolin-1. This review examines cholesterol asymmetry in SPM, potential mechanisms of regulation and impact on membrane structure and function. PMID:21214553

  9. Effects of decreased pH on membrane structural organization of Escherichia coli grown in different fatty acid-supplemented media: a 31P NMR study.

    PubMed

    Ianzini, F; Guidoni, L; Simone, G; Viti, V; Yatvin, M B

    1990-04-01

    Total membranes from Escherichia coli cells grown in different fatty acid-supplemented media have been examined by 31P NMR at different pH values. The isolated inner and outer membranes were also studied and compared to the liposomes formed with the corresponding extracted lipids. While the liposomes show structures that are correlated with lipid composition, degree of fatty acid unsaturation, and pH, the membrane structure is mainly bilayer. The presence of two bilayer phases characterized by different chemical shift anisotropy values (delta nu csa) is detectable at neutral pH; a perturbation of the bilayer phase characterized by the smallest delta nu csa is produced by low pH. Moreover, an isotropic peak is always present in the membrane NMR spectra: its attribution to cardiolipin molecules is discussed on the basis of digestion experiments with phospholipase C. PMID:2181934

  10. FABP4 reversed the regulation of leptin on mitochondrial fatty acid oxidation in mice adipocytes

    PubMed Central

    Gan, Lu; Liu, Zhenjiang; Cao, Weina; Zhang, Zhenzhen; Sun, Chao

    2015-01-01

    Fatty acid binding protein 4 (FABP4), plays key role in fatty acid transportation and oxidation, and increases with leptin synergistically during adipose inflammation process. However, the regulation mechanism between FABP4 and leptin on mitochondrial fatty acid oxidation remains unclear. In this study, we found that FABP4 reduced the expression of leptin, CPT-1 and AOX1 in mice adipocytes. Conversely, FABP4 was down-regulated in a time-dependent manner by leptin treatment. Additionally, forced expression of FABP4 attenuated the expression of PGC1-α, UCP2, CPT-1, AOX1 and COX2 compared with leptin incubation. Moreover, mitochondrial membrane potential, fatty acid oxidation enzyme medium-chain acyl-CoA dehydrogenase (MCAD), long-chain acyl-CoA dehydrogenase (LCAD) and Cyt C levels were reduced in response to the overexpression of FABP4. These reductions correspond well with the reduced release of free fatty acid and the inactivation of mitochondrial complexes I and III by FABP4 overexpression. Furthermore, addition of the Akt/mTOR pathway-specific inhibitor (MK2206) blocked the mitochondrial fatty acid oxidation and respiration factors, whereas interference of FABP4 overcame these effects. Taken together, FABP4 could reverse the activation of the leptin-induced mitochondrial fatty acid oxidation, and the inhibition of Akt/mTOR signal pathway played a key role in this process. PMID:26310911

  11. Does acute lead (Pb) contamination influence membrane fatty acid composition and freeze tolerance in intertidal blue mussels in arctic Greenland?

    PubMed

    Thyrring, Jakob; Juhl, Bodil Klein; Holmstrup, Martin; Blicher, Martin E; Sejr, Mikael K

    2015-11-01

    In their natural habitats, organisms are exposed to multiple stressors. Heavy metal contamination stresses the cell membrane due to increased peroxidation of lipids. Likewise, sub-zero air temperatures potentially reduce membrane functionality in ectothermal animals. We tested if acute lead (Pb) exposure for 7 days would influence survival in intertidal blue mussels (Mytilus edulis) after exposure to realistic sub-zero air temperatures. A full factorial experiment with five tissue Pb concentrations between 0 and 3500 μg Pb/g and six sub-zero temperatures from 0 to -17 °C were used to test the hypothesis that sub-lethal effects of Pb may increase the lethality caused by freezing in blue mussels exposed to temperatures simulating Greenland winter conditions. We found a significant effect of temperature on mortality. However, the short-term exposure to Pb did not result in any effects of Pb, nor did we find interactions between Pb and temperature. We analysed the relative abundance of major phospholipid fatty acids (PLFAs) in the gill tissue, but we found no significant effect of Pb tissue concentration on PLFA composition. Results suggest that Pb accumulation has limited effects on freeze tolerance and does not induce membrane damage in terms of persistent lipid peroxidation. PMID:26438355

  12. Fatty acid fouling of forward osmosis membrane: Effects of pH, calcium, membrane orientation, initial permeate flux and foulant composition.

    PubMed

    Zhao, Pin; Gao, Baoyu; Yue, Qinyan; Liu, Pan; Shon, Ho Kyong

    2016-08-01

    Octanoic acid (OA) was selected to represent fatty acids in effluent organic matter (EOM). The effects of feed solution (FS) properties, membrane orientation and initial permeate flux on OA fouling in forward osmosis (FO) were investigated. The undissociated OA formed a cake layer quickly and caused the water flux to decline significantly in the initial 0.5hr at unadjusted pH3.56; while the fully dissociated OA behaved as an anionic surfactant and promoted the water permeation at an elevated pH of 9.00. Moreover, except at the initial stage, the sudden decline of water flux (meaning the occurrence of severe membrane fouling) occurred in two conditions: 1. 0.5mmol/L Ca(2+), active layer facing draw solution (AL-DS) and 1.5mol/L NaCl (DS); 2. No Ca(2+), active layer-facing FS (AL-FS) and 4mol/L NaCl (DS). This demonstrated that cake layer compaction or pore blocking occurred only when enough foulants were absorbed into the membrane surface, and the water permeation was high enough to compact the deposit inside the porous substrate. Furthermore, bovine serum albumin (BSA) was selected as a co-foulant. The water flux of both co-foulants was between the fluxes obtained separately for the two foulants at pH3.56, and larger than the two values at pH9.00. This manifested that, at pH3.56, BSA alleviated the effect of the cake layer caused by OA, and OA enhanced BSA fouling simultaneously; while at pH9.00, the mutual effects of OA and BSA eased the membrane fouling. PMID:27521936

  13. A high-fat diet and the threonine-encoding allele (Thr54) polymorphism of fatty acid–binding protein 2 reduce plasma triglyceride–rich lipoproteins

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The Thr54 allele of the fatty acid binding protein 2 (FABP2) DNA polymorphism is associated with increased triglyceride-rich lipoproteins and insulin resistance. We investigated whether the triglyceride-rich lipoprotein response to diets of varied fat content is affected by the fatty acid binding pr...

  14. Changes in membrane fatty acids composition of microbial cells induced by addiction of thymol, carvacrol, limonene, cinnamaldehyde, and eugenol in the growing media.

    PubMed

    Di Pasqua, Rosangela; Hoskins, Nikki; Betts, Gail; Mauriello, Gianluigi

    2006-04-01

    Major active compounds from essential oils are well-known to possess antimicrobial activity against both pathogen and spoilage microorganisms. The aim of this work was to determine the alteration of the membrane fatty acid profile as an adaptive mechanism of the cells in the presence of a sublethal concentration of antimicrobial compound in response to a stress condition. Methanolic solutions of thymol, carvacrol, limonene, cinnamaldehyde, and eugenol were added into growth media of Escherichia coli O157:H7, Salmonella enterica serovar typhimurium, Pseudomonas fluorescens, Brochothrix thermosphacta, and Staphylococcus aureus strains. Fatty acid extraction and gas chromatographic analysis were performed to assess changes in membrane fatty acid composition. Substantial changes were observed on the long chain unsaturated fatty acids when the E. coli and Salmonella strains grew in the presence of limonene and cinnamaldehyde and carvacrol and eugenol, respectively. All compounds influenced the fatty acid profile of B. thermosphacta, while Pseudomonas and S. aureus strains did not show substantial changes in their fatty acid compositions. PMID:16569070

  15. Hypobaric bacteriology: growth, cytoplasmic membrane polarization and total cellular fatty acids in Escherichia coli and Bacillus subtilis

    NASA Astrophysics Data System (ADS)

    Pokorny, N. J.; Boulter-Bitzer, J. I.; Hart, M. M.; Storey, L.; Lee, H.; Trevors, J. T.

    2005-10-01

    Escherichia coli JM109 (Gram-negative) and Bacillus subtilis (Gram-positive) were grown under hypobaric conditions for 19 days at 25 °C to study the effects of 33 and 67 kPa low pressures on selected physiological responses; growth, cytoplasmic membrane polarization (measure of cytoplasmic membrane fluidity) and total cellular fatty acids. In the first experiment, cytoplasmic membrane polarization in B. subtilis increased under both hypobaric conditions, indicating the membrane became more rigid or less fluid. This experiment was repeated and the effect of the hypobaric conditions was not evident as in the first experiment with B. subtilis. In addition, total cellular fatty acids analysis for B. subtilis showed that hypobaric conditions did not alter the ratio of saturated to unsaturated fatty acids. The cytoplasmic membrane remained in the same fluid state in hypobaric grown E. coli cell cultures as in the 101 kPa ambient control cells in both experiments. However, the saturated to unsaturated ratios were altered in E. coli under hypobaric conditions. It is important to note the ratios for E. coli were less than 1, while the ratios for Bacillus were in the 28 50 range. Growth of both species was also measured by colony forming units at the termination of the 19 day experiment. Both bacterial species were capable of growth under hypobaric conditions and no distinct trend emerged as to the effect of hypobaric pressure on bacterial growth and cytoplasmic membrane fluidity.

  16. Mitochondrial Carnitine Palmitoyltransferase 1a (CPT1a) Is Part of an Outer Membrane Fatty Acid Transfer Complex*

    PubMed Central

    Lee, Kwangwon; Kerner, Janos; Hoppel, Charles L.

    2011-01-01

    CPT1a (carnitine palmitoyltransferase 1a) in the liver mitochondrial outer membrane (MOM) catalyzes the primary regulated step in overall mitochondrial fatty acid oxidation. It has been suggested that the fundamental unit of CPT1a exists as a trimer, which, under native conditions, could form a dimer of the trimers, creating a hexamer channel for acylcarnitine translocation. To examine the state of CPT1a in the MOM, we employed a combined approach of sizing by mass and isolation using an immunological method. Blue native electrophoresis followed by detection with immunoblotting and mass spectrometry identified large molecular mass complexes that contained not only CPT1a but also long chain acyl-CoA synthetase (ACSL) and the voltage-dependent anion channel (VDAC). Immunoprecipitation with antisera against the proteins revealed a strong interaction between the three proteins. Immobilized CPT1a-specific antibodies immunocaptured not only CPT1a but also ACSL and VDAC, further strengthening findings with blue native electrophoresis and immunoprecipitation. This study shows strong protein-protein interaction between CPT1a, ACSL, and VDAC. We propose that this complex transfers activated fatty acids through the MOM. PMID:21622568

  17. Mitochondrial carnitine palmitoyltransferase 1a (CPT1a) is part of an outer membrane fatty acid transfer complex.

    PubMed

    Lee, Kwangwon; Kerner, Janos; Hoppel, Charles L

    2011-07-22

    CPT1a (carnitine palmitoyltransferase 1a) in the liver mitochondrial outer membrane (MOM) catalyzes the primary regulated step in overall mitochondrial fatty acid oxidation. It has been suggested that the fundamental unit of CPT1a exists as a trimer, which, under native conditions, could form a dimer of the trimers, creating a hexamer channel for acylcarnitine translocation. To examine the state of CPT1a in the MOM, we employed a combined approach of sizing by mass and isolation using an immunological method. Blue native electrophoresis followed by detection with immunoblotting and mass spectrometry identified large molecular mass complexes that contained not only CPT1a but also long chain acyl-CoA synthetase (ACSL) and the voltage-dependent anion channel (VDAC). Immunoprecipitation with antisera against the proteins revealed a strong interaction between the three proteins. Immobilized CPT1a-specific antibodies immunocaptured not only CPT1a but also ACSL and VDAC, further strengthening findings with blue native electrophoresis and immunoprecipitation. This study shows strong protein-protein interaction between CPT1a, ACSL, and VDAC. We propose that this complex transfers activated fatty acids through the MOM. PMID:21622568

  18. Fatty acyl donor selectivity in membrane bound O-acyltransferases and communal cell fate decision-making

    PubMed Central

    Tuladhar, Rubina; Lum, Lawrence

    2015-01-01

    The post-translational modification of proteins with lipid moieties confers spatial and temporal control of protein function by restricting their subcellular distribution or movement in the extracellular milieu. Yet, little is known about the significance of lipid selectivity to the activity of proteins targeted for such modifications. Membrane bound O-acyl transferases (MBOATs) are a superfamily of multipass enzymes that transfer fatty acids on to lipid or protein substrates. Three MBOATs constitute a subfamily with secreted signalling molecules for substrates, the Wnt, Hedgehog (Hh) and Ghrelin proteins. Given their important roles in adult tissue homoeostasis, all three molecules and their respective associated acyltransferases provide a framework for interrogating the role of extracellular acylation events in cell-to-cell communication. Here, we discuss how the preference for a fatty acyl donor in the Wnt acyltransferase porcupine (Porcn) and possibly in other protein lipidation enzymes may provide a means for coupling metabolic health at the single cell level to communal cell fate decision-making in complex multicellular organisms. PMID:25849923

  19. Fatty acyl donor selectivity in membrane bound O-acyltransferases and communal cell fate decision-making.

    PubMed

    Tuladhar, Rubina; Lum, Lawrence

    2015-04-01

    The post-translational modification of proteins with lipid moieties confers spatial and temporal control of protein function by restricting their subcellular distribution or movement in the extracellular milieu. Yet, little is known about the significance of lipid selectivity to the activity of proteins targeted for such modifications. Membrane bound O-acyl transferases (MBOATs) are a superfamily of multipass enzymes that transfer fatty acids on to lipid or protein substrates. Three MBOATs constitute a subfamily with secreted signalling molecules for substrates, the Wnt, Hedgehog (Hh) and Ghrelin proteins. Given their important roles in adult tissue homoeostasis, all three molecules and their respective associated acyltransferases provide a framework for interrogating the role of extracellular acylation events in cell-to-cell communication. Here, we discuss how the preference for a fatty acyl donor in the Wnt acyltransferase porcupine (Porcn) and possibly in other protein lipidation enzymes may provide a means for coupling metabolic health at the single cell level to communal cell fate decision-making in complex multicellular organisms. PMID:25849923

  20. Erythrocyte membrane fatty acids in multiple sclerosis patients and hot-nature dietary intervention with co-supplemented hemp-seed and evening-primrose oils.

    PubMed

    Rezapour-Firouzi, Soheila; Arefhosseini, Seyed Rafie; Ebrahimi-Mamaghani, Mehrangiz; Farhoudi, Mehdi; Baradaran, Behzad; Ali, Torbati Mohammad; Zamani, Fatemeh

    2013-01-01

    The risk of developing multiple sclerosis (MS) is associated with increased dietary intake of saturated fatty acids. For many years it has been suspected that this disease might be associated with an imbalance between unsaturated and saturated fatty acids. We determined erythrocyte membrane fatty acids levels in Hot nature dietary intervention with co-supplemented hemp seed and evening primrose oils in multiple sclerosis patients. To determine the erythrocyte membrane fatty acids levels and correlate it with expanded disability status scale (EDSS) at baseline after 6 months intervention in MS patients by gas chromatography, in this double blind, randomized trial, 100 RRMS patients with EDSS<6 were allocated into three groups: "Group A" that received co-supplemented hemp seed and evening primrose oils with advised Hot nature diet. "Group B" received olive oil and "Group C" received the co-supplemented oils. The results showed that the mean follow-up was 180 ± 2.9SD days (N=65, 23 M and 42 F aged 34.25 ± 8.07 years with disease duration of 6.80 ± 4.33 years). There was no significant difference in the study parameters at baseline. After 6 months, EDSS, Immunological parameters and the erythrocyte cell membrane with regard to specific fatty acids showed improvement in the group A and C, whereas there was worsening condition for the group B after the intervention. We concluded that Hot-nature dietary intervention with co-supplemented hemp seed and evening primrose oils caused an increase PUFAs in MS patients and improvement in the erythrocyte membrane fatty acids composition. This could be an indication of restored plasma stores, and a reflection of disease severity reduction. PMID:24311880

  1. Transfer of fatty acids from the 1-position of phosphatidyl-ethanolamine to the major outer membrane lipoprotein of E coli

    SciTech Connect

    Jackowski, S.; Rock, C.O.

    1986-05-01

    The fatty acids esterified to Braun's lipoprotein are derived from the phospholipid pool in E. coli. Mutants lacking acyl-CoA synthetase activity (fadD) incorporated extracellular fatty acids specifically into the 1-position of phosphatidylethanolamine (PtdEtn). This pathway was blocked by chloramphenicol and was depressed by preventing the acylation of the amino terminus of the lipoprotein with globomycin. Transfer of fatty acids to lipoprotein was investigated in fadD mutants harboring hybrid plasmids containing either the lipoprotein gene or a lipoprotein-..beta..-lactamase gene fusion under control of the lactose promoter. Labeling of the 1-position of the PtdEtn pool prior to induction of lipoprotein biosynthesis resulted in the transfer of fatty acids from PtdEtn to the lipoproteins. Induction of lipoprotein synthesis in the presence of exogenous (1-/sup 14/C)palmitate increased the amount of radioactivity entering the PtdEtn pool and efficiently labeled lipoprotein acyl moieties. Lipoprotein fatty acids derived from the 1-position of PtdEtn were resistant to hydroxylamine hydrolysis, and globomycin reduced the incorporation of exogenous (1-/sup 14/C)palmitic acid into lipoproteins by 80% suggesting that the fatty acid is attached to the amino terminus. These data illustrate the metabolic relationship between turnover of fatty acids in the 1-position of PtdEtn and the maturation of the major outer membrane lipoprotein.

  2. The removal of fatty acids from edible oil. Removal of the dispersed phase of a water-in-oil dispersion by a hydrophilic membrane

    SciTech Connect

    Keutrentjes, J.T.F.; Doornbusch, G.I.; Van't Riet, K. )

    1991-01-01

    Fatty acids can be extracted from an oil phase by forming a dispersed phase of saponified fatty acids/water/isopropanol in oil. This dispersion can be separated in the two phases by two membranes of opposite polarity in series. In this study the separation of the water phase from the dispersion by a hydrophilic membrane and the mechanisms underlying the flux characteristics are investigated. The permeation flux through a PAN ultrafiltration membrane is optimized with respect to the fatty acid/water/isopropanol ratio. It appears that a 1:6.5:3 (v/v) ratio gives the highest flux (95/(m{sup 2}{center dot}h{center dot}bar)). The dispersion at these conditions consists of a continuous oil phase as well as a continuous water phase between 20 and 65% water phase hold up. The flux/pressure curve shows a linear increase of the flux with pressure at low pressures (determined buy the membrane resistance), followed by a maximum flux value for the case where the volume of the water phase present in the inflow is limiting. It is not possible to remove the water phase with membranes below a water phase hold up to 20%. At this hold up value the transition between a bicontinuous and a discrete dispersion also occurs.

  3. Purified membrane and soluble folate binding proteins from cultured KB cells have similar amino acid compositions and molecular weights but differ in fatty acid acylation.

    PubMed Central

    Luhrs, C A; Pitiranggon, P; da Costa, M; Rothenberg, S P; Slomiany, B L; Brink, L; Tous, G I; Stein, S

    1987-01-01

    A membrane-associated folate binding protein (FBP) and a soluble FBP, which is released into the culture medium, have been purified from human KB cells using affinity chromatography. By NaDodSO4/PAGE, both proteins have an apparent Mr of approximately 42,000. However, in the presence of Triton X-100, the soluble FBP eluted from a Sephadex G-150 column with an apparent Mr of approximately 40,000 (similar to NaDodSO4/PAGE) but the membrane-associated FBP eluted with an apparent Mr of approximately 160,000, indicating that this species contains a hydrophobic domain that interacts with the detergent micelles. The amino acid compositions of both forms of FBP were similar, especially with respect to the apolar amino acids. In addition, the 18 amino acids at the amino termini of both proteins were identical. The membrane FBP, following delipidation with chloroform/methanol, contained 7.1 mol of fatty acid per mol of protein, of which 4.7 mol was amide-linked and 2.4 mol was ester-linked. The soluble FBP contained only 0.05 mol of fatty acid per mol of protein. These studies indicate that the membrane FBP of KB cells contains covalently bound fatty acids that may serve to anchor the protein in the cell membrane. Images PMID:3476960

  4. Changes in the Fatty Acid Profile and Phospholipid Molecular Species Composition of Human Erythrocyte Membranes after Hybrid Palm and Extra Virgin Olive Oil Supplementation.

    PubMed

    Pacetti, D; Gagliardi, R; Balzano, M; Frega, N G; Ojeda, M L; Borrero, M; Ruiz, A; Lucci, P

    2016-07-13

    This work aims to evaluate and compare, for the first time, the effects of extra virgin olive oil (EVOO) and hybrid palm oil (HPO) supplementation on the fatty acid profile and phospholipid (PL) molecular species composition of human erythrocyte membranes. Results supported the effectiveness of both HPO and EVOO supplementation (3 months, 25 mL/day) in decreasing the lipophilic index of erythrocytes with no significant differences between HPO and EVOO groups at month 3. On the other hand, the novel and rapid ultraperformance liquid chromatography-tandem mass spectrometry method used for PL analysis reveals an increase in the levels of phosphatidylcholine and phosphatidylethanolamine species esterified with polyunsaturated fatty acids. This work demonstrates the ability of both EVOO and HPO to increase the degree of unsaturation of erythrocyte membrane lipids with an improvement in membrane fluidity that could be associated with a lower risk of developing cardiovascular diseases. PMID:27315139

  5. Effect of Saturated Very Long-Chain Fatty Acids on the Organization of Lipid Membranes: A Study Combining (2)H NMR Spectroscopy and Molecular Dynamics Simulations.

    PubMed

    Paz Ramos, Adrian; Lagüe, Patrick; Lamoureux, Guillaume; Lafleur, Michel

    2016-07-21

    Little is known about the interaction of very long-chain saturated fatty acids (VLCFAs) with biological membranes. However, this could play an important role on interleaflet interactions and signal transduction mechanisms in cells. The aim of this work is to determine how VLCFA structurally adapts in fluid phospholipid bilayers, since both species must exhibit a significant hydrophobic mismatch. The membrane organization has been described by means of (2)H NMR and molecular dynamics simulations. Our results show that the protonation state affects the position and order of free fatty acids (FFAs) in phospholipid membranes. It was shown that the protonated FFA-C24 carboxyl group is located slightly under the POPC head group and therefore its acyl chain can interact with the lipids of the opposite leaflet. This interdigitation of the end of the acyl chain causes a second plateau observed in SC-D profiles, a very unusual feature in lipid systems. PMID:27351151

  6. Role of plasma membrane lipid composition on cellular homeostasis: learning from cell line models expressing fatty acid desaturases

    PubMed Central

    Jaureguiberry, María S.; Tricerri, M. Alejandra; Sanchez, Susana A.; Finarelli, Gabriela S.; Montanaro, Mauro A.; Prieto, Eduardo D.; Rimoldi, Omar J.

    2014-01-01

    Experimental evidence has suggested that plasma membrane (PM)-associated signaling and hence cell metabolism and viability depend on lipid composition and organization. The aim of the present work is to develop a cell model to study the endogenous polyunsaturated fatty acids (PUFAs) effect on PM properties and analyze its influence on cholesterol (Chol) homeostasis. We have previously shown that by using a cell line over-expressing stearoyl-CoA-desaturase, membrane composition and organization coordinate cellular pathways involved in Chol efflux and cell viability by different mechanisms. Now, we expanded our studies to a cell model over-expressing both Δ5 and Δ6 desaturases, which resulted in a permanently higher PUFA content in PM. Furthermore, this cell line showed increased PM fluidity, Chol storage, and mitochondrial activity. In addition, human apolipoprotein A-I-mediated Chol removal was less efficient in these cells than in the corresponding control. Taken together, our results suggested that the cell functionality is preserved by regulating PM organization and Chol exportation and homeostasis. PMID:24473084

  7. Membrane simulations mimicking acidic pH reveal increased thickness and negative curvature in a bilayer consisting of lysophosphatidylcholines and free fatty acids.

    PubMed

    Lähdesmäki, Katariina; Ollila, O H Samuli; Koivuniemi, Artturi; Kovanen, Petri T; Hyvönen, Marja T

    2010-05-01

    Phospholipids are key components of biological membranes and their lipolysis with phospholipase A(2) (PLA(2)) enzymes occurs in different cellular pH environments. Since no studies are available on the effect of pH on PLA(2)-modified phospholipid membranes, we performed 50-ns atomistic molecular dynamics simulations at three different pH conditions (pH 9.0, 7.5, and 5.5) using a fully PLA(2)-hydrolyzed phosphatidylcholine (PC) bilayer which consists solely of lysophosphatidylcholine and free fatty acid molecules. We found that a decrease in pH results in lateral squeezing of the membrane, i.e. in decreased surface area per headgroup. Thus, at the decreased pH, the lipid hydrocarbon chains had larger S(CD) order parameter values, and also enhanced membrane thickness, as seen in the electron density profiles across the membrane. From the lateral pressure profiles, we found that the values of spontaneous curvature of the two opposing monolayers became negative when the pH was decreased. At low pH, protonation of the free fatty acid headgroups reduces their mutual repulsion and accounts for the pH dependence of all the above-mentioned properties. The altered structural characteristics may significantly affect the overall surface properties of biomembranes in cellular vesicles, lipid droplets, and plasma lipoproteins, play an important role in membrane fission and fusion, and modify interactions between membrane lipids and the proteins embedded within them. PMID:20132791

  8. Blood-brain barrier and brain fatty acid uptake: Role of arachidonic acid and PGE2.

    PubMed

    Murphy, Eric J

    2015-12-01

    How do fatty acids enter the brain and what role, if any, do membrane and cytosolic fatty acid binding proteins have on facilitating this process? This is a fundamental question that many lipid neurochemists will freely admit they cannot answer in any kind of definitive manner. A study by Dalvi and colleagues in this issue of the Journal of Neurochemistry now adds to our knowledge in this field. Among other important observations, their experiments demonstrate that a physiological level of arachidonic acid (ARA), that could be associated with many different physiological and pathophysiological states, increases permeability in a model of the human blood brain barrier (BBB) in the absence of cytokines. This last point is very important as it suggests increases in BBB permeability may occur in situations other than those associated with increases in tumor necrosis factor a (TNFα) and interleukin1b (IL1β), giving additional options for developing drugs impacting BBB permeability. PMID:26383055

  9. Characterization of phosphonic acid binding to zinc oxide

    SciTech Connect

    Hotchkiss, Peter J.; Malicki, Michał; Giordano, Anthony J.; Armstrong, Neal R.; Marder, Seth R.

    2011-01-24

    Radio Frequency (RF) sputter-deposited zinc oxide (ZnO) films have been modified with alkylphosphonic acids in order to study both the binding of the phosphonic acid (PA) group to the ZnO surface and the packing of the alkyl chain. The characterization of these PA-modified ZnO substrates by X-ray photoelectron spectroscopy (XPS), infrared reflection-absorption spectroscopy (IRRAS), atomic force microscopy (AFM) and contact angle measurements is presented herein. The surface modification procedure is straightforward and was adapted from earlier work. XPS analysis shows that oxygen plasma (OP) treatment creates reactive oxygen species on the surface of ZnO, allowing for a more robust binding of PAs to the ZnO surface. IRRAS analysis indicates that octadecylphosphonic acid binds to the ZnO surface in a predominantly tridentate fashion, forming dense, well-packed monolayers with alkyl chains in a fully anti-conformation. AFM and contact angle measurements indicate good surface coverage of the PAs with little to no multilayer formation.

  10. Membranes as Structural Antioxidants: RECYCLING OF MALONDIALDEHYDE TO ITS SOURCE IN OXIDATION-SENSITIVE CHLOROPLAST FATTY ACIDS.

    PubMed

    Schmid-Siegert, Emanuel; Stepushenko, Olga; Glauser, Gaetan; Farmer, Edward E

    2016-06-17

    Genetic evidence suggests that membranes rich in polyunsaturated fatty acids (PUFAs) act as supramolecular antioxidants that capture reactive oxygen species, thereby limiting damage to proteins. This process generates lipid fragmentation products including malondialdehyde (MDA), an archetypal marker of PUFA oxidation. We observed transient increases in levels of endogenous MDA in wounded Arabidopsis thaliana leaves, raising the possibility that MDA is metabolized. We developed a rigorous ion exchange method to purify enzymatically generated (13)C- and (14)C-MDA. Delivered as a volatile to intact plants, MDA was efficiently incorporated into lipids. Mass spectral and genetic analyses identified the major chloroplast galactolipid: α-linolenic acid (18:3)-7Z,10Z,13Z-hexadecatrienoic acid (16:3)-monogalactosyldiacylglycerol (18:3-16:3-MGDG) as an end-product of MDA incorporation. Consistent with this, the fad3-2 fad7-2 fad8 mutant that lacks tri-unsaturated fatty acids incorporated (14)C-MDA into 18:2-16:2-MGDG. Saponification of (14)C-labeled 18:3-16:3-MGDG revealed 84% of (14)C-label in the acyl groups with the remaining 16% in the head group. 18:3-16:3-MGDG is enriched proximal to photosystem II and is likely a major in vivo source of MDA in photosynthetic tissues. We propose that nonenzymatically generated lipid fragments such as MDA are recycled back into plastidic galactolipids that, in their role as cell protectants, can again be fragmented into MDA. PMID:27143359

  11. Characterization of 19 Genes Encoding Membrane-Bound Fatty Acid Desaturases and their Expression Profiles in Gossypium raimondii Under Low Temperature.

    PubMed

    Liu, Wei; Li, Wei; He, Qiuling; Daud, Muhammad Khan; Chen, Jinhong; Zhu, Shuijin

    2015-01-01

    To produce unsaturated fatty acids, membrane-bound fatty acid desaturases (FADs) can be exploited to introduce double bonds into the acyl chains of fatty acids. In this study, 19 membrane-bound FAD genes were identified in Gossypium raimondii through database searches and were classified into four different subfamilies based on phylogenetic analysis. All 19 membrane-bound FAD proteins shared three highly conserved histidine boxes, except for GrFAD2.1, which lost the third histidine box in the C-terminal region. In the G. raimondii genome, tandem duplication might have led to the increasing size of the FAD2 cluster in the Omega Desaturase subfamily, whereas segmental duplication appeared to be the dominant mechanism for the expansion of the Sphingolipid and Front-end Desaturase subfamilies. Gene expression analysis showed that seven membrane-bound FAD genes were significantly up-regulated and that five genes were greatly suppressed in G. raimondii leaves exposed to low temperature conditions. PMID:25894196

  12. Characterization of 19 Genes Encoding Membrane-Bound Fatty Acid Desaturases and their Expression Profiles in Gossypium raimondii Under Low Temperature

    PubMed Central

    He, Qiuling; Daud, Muhammad Khan; Chen, Jinhong; Zhu, Shuijin

    2015-01-01

    To produce unsaturated fatty acids, membrane-bound fatty acid desaturases (FADs) can be exploited to introduce double bonds into the acyl chains of fatty acids. In this study, 19 membrane-bound FAD genes were identified in Gossypium raimondii through database searches and were classified into four different subfamilies based on phylogenetic analysis. All 19 membrane-bound FAD proteins shared three highly conserved histidine boxes, except for GrFAD2.1, which lost the third histidine box in the C-terminal region. In the G. raimondii genome, tandem duplication might have led to the increasing size of the FAD2 cluster in the Omega Desaturase subfamily, whereas segmental duplication appeared to be the dominant mechanism for the expansion of the Sphingolipid and Front-end Desaturase subfamilies. Gene expression analysis showed that seven membrane-bound FAD genes were significantly up-regulated and that five genes were greatly suppressed in G. raimondii leaves exposed to low temperature conditions. PMID:25894196

  13. Interaction of free fatty acids with the erythrocyte membrane as affected by hyperthermia and ionizing radiation

    SciTech Connect

    Rybczynska, M.; Csordas, A. )

    1990-04-01

    The interference of hyperthermia and ionizing radiation, respectively, with the effects of capric (10:0), lauric (12:0), myristic (14:0), oleic (cis-18:1) and elaidic (trans-18:1) acids on the osmotic resistance of human erythrocytes was investigated. The results are summarized as follows: (A) not only at 37 degrees, but also at 42 and 47{degrees}C lauric acid (12:0) represents the minimum chain length for the biphasic behaviour of protecting against hypotonic hemolysis at a certain lower concentration range and hemolysis promotion at subsequent higher concentrations; (B) with increasing temperatures the protecting as well as the hemolytic effects occur at lower concentrations of the fatty acids; (C) the increase of temperature promotes the extent of hemolysis and reduces the extent of protection against hypotonic hemolysis; (D) Gamma-irradiation of erythrocytes selectively affects the concentration of oleic acid at which maximum protection against hypotonic hemolysis occurs, without altering the minimum concentration for 100% hemolysis.

  14. Retinoic acid-binding protein, rhombomeres and the neural crest.

    PubMed

    Maden, M; Hunt, P; Eriksson, U; Kuroiwa, A; Krumlauf, R; Summerbell, D

    1991-01-01

    We have investigated by immunocytochemistry the spatial and temporal distribution of cellular retinoic acid-binding protein (CRABP) in the developing nervous system of the chick embryo in order to answer two specific questions: do neural crest cells contain CRABP and where and when do CRABP-positive neuroblasts first arise in the neural tube? With regard to the neural crest, we have compared CRABP staining with HNK-1 staining (a marker of migrating neural crest) and found that they do indeed co-localise, but cephalic and trunk crest behave slightly differently. In the cephalic region in tissues such as the frontonasal mass and branchial arches, HNK-1 immunoreactivity is intense at early stages, but it disappears as CRABP immunoreactivity appears. Thus the two staining patterns do not overlap, but are complementary. In the trunk, HNK-1 and CRABP stain the same cell populations at the same time, such as those migrating through the anterior halves of the somites. In the neural tube, CRABP-positive neuroblasts first appear in the rhombencephalon just after the neural folds close and then a particular pattern of immunoreactivity appears within the rhombomeres of the hindbrain. Labelled cells are present in the future spinal cord, the posterior rhombencephalon up to rhombomere 6 and in rhombomere 4 thus producing a single stripe pattern. This pattern is dynamic and gradually changes as anterior rhombomeres begin to label. The similarity of this initial pattern to the arrangement of certain homeobox genes in the mouse stimulated us to examine the expression of the chicken Hox-2.9 gene. We show that at stage 15 the pattern of expression of this gene is closely related to that of CRABP. The relationship between retinoic acid, CRABP and homeobox genes is discussed. PMID:1707786

  15. Effects of sub-lethal concentrations of thyme and oregano essential oils, carvacrol, thymol, citral and trans-2-hexenal on membrane fatty acid composition and volatile molecule profile of Listeria monocytogenes, Escherichia coli and Salmonella enteritidis.

    PubMed

    Siroli, Lorenzo; Patrignani, Francesca; Gardini, Fausto; Lanciotti, Rosalba

    2015-09-01

    The aim of this work was to investigate the modifications of cell membrane fatty acid composition and volatile molecule profiles of Listeria monocytogenes, Salmonella enteritidis, Escherichia coli, during growth in the presence of different sub-lethal concentrations of thyme and oregano essential oils as well as carvacrol, thymol, trans-2-hexenal and citral. The results evidenced that the tested molecules induced noticeable modifications of membrane fatty acid profiles and volatile compounds produced during the growth. Although specific differences in relation to the species considered were identified, the tested compounds induced a marked increase of some membrane associated fatty acids, particularly unsaturated fatty acids, trans-isomers, and specific released free fatty acids. These findings can contribute to the comprehension of the stress response mechanisms used by different pathogenic microorganisms often involved in food-borne diseases in relation to the exposure to sub-lethal concentrations of natural antimicrobials. PMID:25842326

  16. Bacterial periplasmic sialic acid-binding proteins exhibit a conserved binding site

    SciTech Connect

    Gangi Setty, Thanuja; Cho, Christine; Govindappa, Sowmya; Apicella, Michael A.; Ramaswamy, S.

    2014-07-01

    Structure–function studies of sialic acid-binding proteins from F. nucleatum, P. multocida, V. cholerae and H. influenzae reveal a conserved network of hydrogen bonds involved in conformational change on ligand binding. Sialic acids are a family of related nine-carbon sugar acids that play important roles in both eukaryotes and prokaryotes. These sialic acids are incorporated/decorated onto lipooligosaccharides as terminal sugars in multiple bacteria to evade the host immune system. Many pathogenic bacteria scavenge sialic acids from their host and use them for molecular mimicry. The first step of this process is the transport of sialic acid to the cytoplasm, which often takes place using a tripartite ATP-independent transport system consisting of a periplasmic binding protein and a membrane transporter. In this paper, the structural characterization of periplasmic binding proteins from the pathogenic bacteria Fusobacterium nucleatum, Pasteurella multocida and Vibrio cholerae and their thermodynamic characterization are reported. The binding affinities of several mutations in the Neu5Ac binding site of the Haemophilus influenzae protein are also reported. The structure and the thermodynamics of the binding of sugars suggest that all of these proteins have a very well conserved binding pocket and similar binding affinities. A significant conformational change occurs when these proteins bind the sugar. While the C1 carboxylate has been identified as the primary binding site, a second conserved hydrogen-bonding network is involved in the initiation and stabilization of the conformational states.

  17. Monounsaturated fatty acids reduce the barrier of stratum corneum lipid membranes by enhancing the formation of a hexagonal lateral packing.

    PubMed

    Mojumdar, Enamul H; Helder, Richard W J; Gooris, Gert S; Bouwstra, Joke A

    2014-06-10

    The effectiveness of the skin barrier underlies the outer layer of the skin: the stratum corneum (SC). However, in several skin diseases this barrier is impaired. In two inflammatory skin diseases, atopic eczema and Netherton syndrome, an increased level of monounsaturated fatty acids (MUFAs) has been observed as opposed to healthy skin. In the present study, we aimed to investigate the effect of MUFAs on the lipid organization and skin lipid barrier using an in vitro model membrane system, the stratum corneum substitute (SCS), mimicking the SC lipid composition and organization. To achieve our goal, the SCS has been prepared with increasing levels of MUFAs using various chain length. Permeation studies and trans-epidermal water loss measurements show that an increment of MUFAs reduces the lipid barrier in the SCS. The increased level of unsaturation exerts its effect by reducing the packing density in the lipid organization, while the lamellar phases are not affected. Our findings indicate that increased levels of MUFAs may contribute to the impaired skin barrier in diseased skin. PMID:24818519

  18. Fatty acids and plasmalogens of the phospholipids of the sperm membranes and their relation with the post-thaw quality of stallion spermatozoa.

    PubMed

    Macías García, B; González Fernández, L; Ortega Ferrusola, C; Morillo Rodríguez, A; Gallardo Bolaños, J M; Rodríguez Martinez, H; Tapia, J A; Morcuende, D; Peña, F J

    2011-03-15

    Fatty acids and plasmalogens were extracted from the phospholipids of the plasma membrane of stallion spermatozoa, to determine their relation with sperm quality after freezing and thawing. Sperm quality was rated using a quality index that combined the results of the analysis of sperm motility and velocity (CASA analysis), membrane status and mitochondrial membrane potential (flow cytometry) post thaw. Receiving operating system (ROC) curves were used to evaluate the value of specific lipid components of the sperm membrane herein studied as forecast of potential freezeability. From all parameters studied the ratio of percentage of C16 plasmalogens related to total phospholipids was the one with the better diagnostic value. For potentially bad freezers, the significant area under the ROC-curve was 0.74, with 75% sensitivity and 79.9% specificity for a cut off value of 26.9. Also the percentage of plasmalogens respect to total phospholipids gave good diagnostic value for bad freezers. On the other hand, the percentage of C18 fatty aldehydes related to total phospholipids of the sperm membrane properly forecasted freezeability with an area under the ROC curve of 0.70 with 70% sensitivity and 62.5% specificity for a cut off value of 0.32. PMID:21144567

  19. Lipids, curvature stress, and the action of lipid prodrugs: free fatty acids and lysolipid enhancement of drug transport across liposomal membranes.

    PubMed

    Jespersen, Henrik; Andersen, Jonas H; Ditzel, Henrik J; Mouritsen, Ole G

    2012-01-01

    Molecular shape and its impact on bilayer curvature stress are powerful concepts for describing the effects of lipids and fatty acids on fundamental membrane properties, such as passive permeability and derived properties like drug transport across liposomal membranes. We illustrate these relationships by studying the effects of fatty acids and lysolipids on the permeation of a potent anti-cancer drug, doxorubicin, across the bilayer of a liposome in which the drug is encapsulated. Using a simple fluorescence assay, we have systematically studied the passive permeation of doxorubicin across liposomal membranes in different lipid phases: the solid-ordered phase (DPPC bilayers), the liquid-disordered phase (POPC lipid bilayers), and the liquid-ordered phase induced by high levels of cholesterol (DOPC + cholesterol lipid bilayers). The effect of different free fatty acids (FA) and lysolipids (LL), separately and in combination, on permeability was assessed to elucidate the possible mechanism of phospholipase A(2)-triggered release in cancer tissue of liposomal doxorubicin formulations. In all cases, FAs applied separately lead to significant enhancement of permeability, most pronounced in liquid-disordered bilayers and less pronounced in solid and solid-ordered bilayers. LLs applied separately had only a marginal effect on permeability. FA and LL applied in combination lead to a synergistic enhancement of permeability in solid bilayers, whereas in liquid-disordered bilayers, the combined effect suppressed the otherwise strong permeability enhancement due to the FAs. PMID:21839138

  20. Improving volatile fatty acid yield from sludge anaerobic fermentation through self-forming dynamic membrane separation.

    PubMed

    Liu, Hongbo; Wang, Yuanyuan; Yin, Bo; Zhu, Yanfang; Fu, Bo; Liu, He

    2016-10-01

    Self-forming dynamic membrane (SFDM) separation was applied to the conventional sludge fermenter for improving VFA yields. Results indicated SFDM presented good performance in transferring products, retaining substrates, and enriching useful bacteria. The retention ratios of suspended solids, soluble COD, proteins, and polysaccharides reached 99%, 30%, 70%, and 40%, respectively, and more than 90% of the VFAs and ammonia could be transferred in a timely manner. The structure of the microbial community was optimized, which led to enhanced releases of hydrolytic enzymes and accelerated enrichments of functional bacteria. Protease and β-glucosidase activities increased from 1.0 to 5.0U/mL and 15.0 to 23.0μmol/L·h, respectively. VFA yield and sludge conversion ratio increased by 233.3% and 227.9%, respectively. Moreover, SFDM had good operation stability, including a short formation time, a long operation period, and a low transmembrane pressure. These results show VFA yield from sludge fermentation can be greatly improved by SFDM separation. PMID:27347803

  1. Evaluating Healthful Properties of Cereals and Cereal Fractions by Their Bile-Acid-Binding Potential

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The healthful, cholesterol-lowering (atherosclerosis amelioration) or detoxification of harmful metabolites (cancer prevention) potential of cereals and cereal fractions could be predicted by evaluating their in vitro bile acid binding under physiological conditions. Using equal dry matter per incu...

  2. Insertion of fluorescent fatty acid probes into the outer membranes of the pathogenic spirochaetes Treponema pallidum and Borrelia burgdorferi.

    PubMed

    Cox, D L; Radolf, J D

    2001-05-01

    The authors examined the ability of octadecanoyl (C(18)), hexadecanoyl (C(16)) and dodecanoyl (C(12)) fatty acid (FA) conjugates of 5-aminofluorescein (OAF, HAF and DAF, respectively) to insert into the outer membranes (OMs) of Treponema pallidum, Borrelia burgdorferi and Escherichia coli. Biophysical studies have demonstrated that these compounds stably insert into phospholipid bilayers with the acyl chain within the hydrophobic interior of the apical leaflet and the hydrophilic fluorescein moiety near the phospholipid head groups. Consistent with the known poor intrinsic permeability of the E. coli OM to hydrophobic compounds and surfactants, E. coli was not labelled with any of the FA probes. OAF inserted more readily into OMs of B. burgdorferi than into those of T. pallidum, although both organisms were completely labelled at concentrations at or below 2 microg ml(-1). Intact spirochaetes were labelled with OAF but not with antibodies against known periplasmic antigens, thereby confirming that the probe interacted exclusively with the spirochaetal OMs. Separate experiments in which organisms were cooled to 4 degrees C (i.e. below the OM phase-transition temperatures) indicated that labelling with OAF was due to insertion of the probe into the OMs. B. burgdorferi, but not T. pallidum, was labelled by relatively high concentrations of HAF and DAF. Taken as a whole, these findings support the prediction that the lack of lipopolysaccharide renders T. pallidum and B. burgdorferi OMs markedly more permeable to lipophilic compounds than their Gram-negative bacterial counterparts. The data also raise the intriguing possibility that these two pathogenic spirochaetes obtain long-chain FAs, nutrients they are unable to synthesize, by direct permeation of their OMs. PMID:11320119

  3. Summer-to-Winter Phenotypic Flexibility of Fatty Acid Transport and Catabolism in Skeletal Muscle and Heart of Small Birds.

    PubMed

    Zhang, Yufeng; King, Marisa O; Harmon, Erin; Swanson, David L

    2015-01-01

    Prolonged shivering in birds is mainly fueled by lipids. Consequently, lipid transport and catabolism are vital for thermogenic performance and could be upregulated along with thermogenic capacity as part of the winter phenotype. We investigated summer-to-winter variation in lipid transport and catabolism by measuring mRNA expression, protein levels, and enzyme activities for several key steps of lipid transport and catabolic pathways in pectoralis muscle and heart in two small temperate-zone resident birds, American goldfinches (Spinus tristis) and black-capped chickadees (Poecile atricapillus). Cytosolic fatty acid binding protein (FABPc; a key component of intramyocyte lipid transport) mRNA and/or protein levels were generally higher in winter for pectoralis muscle and heart for both species. However, seasonal variation in plasma membrane lipid transporters, fatty acyl translocase, and plasma membrane fatty acid binding protein in pectoralis and heart differed between the two species, with winter increases for chickadees and seasonal stability or summer increases for goldfinches. Catabolic enzyme activities generally showed limited seasonal differences for both tissues and both species. These data suggest that FABPc is an important target of upregulation for the winter phenotype in pectoralis and heart of both species. Plasma membrane lipid transporters and lipid catabolic capacity were also elevated in winter for chickadees but not for goldfinches. Because the two species show differential regulation of distinct aspects of lipid transport and catabolism, these data are consistent with other recent studies documenting that different bird species or populations employ a variety of strategies to promote elevated winter thermogenic capacity. PMID:26658250

  4. Transport and signaling via the amino acid binding site of the yeast Gap1 amino acid transceptor.

    PubMed

    Van Zeebroeck, Griet; Bonini, Beatriz Monge; Versele, Matthias; Thevelein, Johan M

    2009-01-01

    Transporter-related nutrient sensors, called transceptors, mediate nutrient activation of signaling pathways through the plasma membrane. The mechanism of action of transporting and nontransporting transceptors is unknown. We have screened 319 amino acid analogs to identify compounds that act on Gap1, a transporting amino acid transceptor in yeast that triggers activation of the protein kinase A pathway. We identified competitive and noncompetitive inhibitors of transport, either with or without agonist action for signaling, including nontransported agonists. Using substituted cysteine accessibility method (SCAM) analysis, we identified Ser388 and Val389 as being exposed into the amino acid binding site, and we show that agonist action for signaling uses the same binding site as used for transport. Our results provide the first insight, to our knowledge, into the mechanism of action of transceptors. They indicate that signaling requires a ligand-induced specific conformational change that may be part of but does not require the complete transport cycle. PMID:19060912

  5. A Self-Assembled Aggregate Composed of a Fatty Acid Membrane and the Building Blocks of Biological Polymers Provides a First Step in the Emergence of Protocells.

    PubMed

    Black, Roy A; Blosser, Matthew C

    2016-01-01

    We propose that the first step in the origin of cellular life on Earth was the self-assembly of fatty acids with the building blocks of RNA and protein, resulting in a stable aggregate. This scheme provides explanations for the selection and concentration of the prebiotic components of cells; the stabilization and growth of early membranes; the catalysis of biopolymer synthesis; and the co-localization of membranes, RNA and protein. In this article, we review the evidence and rationale for the formation of the proposed aggregate: (i) the well-established phenomenon of self-assembly of fatty acids to form vesicles; (ii) our published evidence that nucleobases and sugars bind to and stabilize such vesicles; and (iii) the reasons why amino acids likely do so as well. We then explain how the conformational constraints and altered chemical environment due to binding of the components to the membrane could facilitate the formation of nucleosides, oligonucleotides and peptides. We conclude by discussing how the resulting oligomers, even if short and random, could have increased vesicle stability and growth more than their building blocks did, and how competition among these vesicles could have led to longer polymers with complex functions. PMID:27529283

  6. fat-1 transgene expression prevents cell culture-induced loss of membrane n-3 fatty acids in activated CD4+ T-cells.

    PubMed

    Fan, Yang-Yi; Kim, Wooki; Callaway, Evelyn; Smith, Roger; Jia, Qian; Zhou, Lan; McMurray, David N; Chapkin, Robert S

    2008-12-01

    In order to evaluate the effects of fatty acids on immune cell membrane structure and function, it is often necessary to maintain cells in culture. However, cell culture conditions typically reverse alterations in polyunsaturated fatty acid (PUFA) composition achieved by dietary lipid manipulation. Therefore, we hypothesized that T-cells from transgenic mice expressing the Caenorhabditis elegans n-3 desaturase (fat-1) gene would be resistant to the culture-induced loss of n-3 PUFA and, therefore, obviate the need to incorporate fatty acids or homologous serum into the medium. CD4+ T-cells were isolated from (i) control wild type (WT) mice fed a safflower oil-n-6 PUFA enriched diet (SAF) devoid of n-3 PUFA, (ii) fat-1 transgenic mice (enriched with endogenous n-3 PUFA) fed a SAF diet, or (iii) WT mice fed a fish oil (FO) based diet enriched in n-3 PUFA. T-cell phospholipids isolated from WT mice fed FO diet (enriched in n-3 PUFA) and fat-1 transgenic mice fed a SAF diet (enriched in n-6 PUFA) were both enriched in n-3 PUFA. As expected, the mol% levels of both n-3 and n-6 PUFA were decreased in cultures of CD4+ T-cells from FO-fed WT mice after 3d in culture. In contrast, the expression of n-3 desaturase prevented the culture-induced decrease of n-3 PUFA in CD4+ T-cells from the transgenic mice. Carboxyfluorescein succinidyl ester (CFSE) -labeled CD4+ T-cells from fat-1/SAF vs. WT/SAF mice stimulated with anti-CD3 and anti-CD28 for 3d, exhibited a reduced (P<0.05) number of cell divisions. We conclude that fat-1-containing CD4+ T-cells express a physiologically relevant, n-3 PUFA enriched, membrane fatty acid composition which is resistant to conventional cell culture-induced depletion. PMID:18977126

  7. Atomic resolution view into the structure–function relationships of the human myelin peripheral membrane protein P2

    PubMed Central

    Ruskamo, Salla; Yadav, Ravi P.; Sharma, Satyan; Lehtimäki, Mari; Laulumaa, Saara; Aggarwal, Shweta; Simons, Mikael; Bürck, Jochen; Ulrich, Anne S.; Juffer, André H.; Kursula, Inari; Kursula, Petri

    2014-01-01

    P2 is a fatty acid-binding protein expressed in vertebrate peripheral nerve myelin, where it may function in bilayer stacking and lipid transport. P2 binds to phospholipid membranes through its positively charged surface and a hydrophobic tip, and accommodates fatty acids inside its barrel structure. The structure of human P2 refined at the ultrahigh resolution of 0.93 Å allows detailed structural analyses, including the full organization of an internal hydrogen-bonding network. The orientation of the bound fatty-acid carboxyl group is linked to the protonation states of two coordinating arginine residues. An anion-binding site in the portal region is suggested to be relevant for membrane interactions and conformational changes. When bound to membrane multilayers, P2 has a preferred orientation and is stabilized, and the repeat distance indicates a single layer of P2 between membranes. Simulations show the formation of a double bilayer in the presence of P2, and in cultured cells wild-type P2 induces membrane-domain formation. Here, the most accurate structural and functional view to date on P2, a major component of peripheral nerve myelin, is presented, showing how it can interact with two membranes simultaneously while going through conformational changes at its portal region enabling ligand transfer. PMID:24419389

  8. Five Decades with Polyunsaturated Fatty Acids: Chemical Synthesis, Enzymatic Formation, Lipid Peroxidation and Its Biological Effects

    PubMed Central

    Catalá, Angel

    2013-01-01

    I have been involved in research on polyunsaturated fatty acids since 1964 and this review is intended to cover some of the most important aspects of this work. Polyunsaturated fatty acids have followed me during my whole scientific career and I have published a number of studies concerned with different aspects of them such as chemical synthesis, enzymatic formation, metabolism, transport, physical, chemical, and catalytic properties of a reconstructed desaturase system in liposomes, lipid peroxidation, and their effects. The first project I became involved in was the organic synthesis of [1-14C] eicosa-11,14-dienoic acid, with the aim of demonstrating the participation of that compound as a possible intermediary in the biosynthesis of arachidonic acid “in vivo.” From 1966 to 1982, I was involved in several projects that study the metabolism of polyunsaturated fatty acids. In the eighties, we studied fatty acid binding protein. From 1990 up to now, our laboratory has been interested in the lipid peroxidation of biological membranes from various tissues and different species as well as liposomes prepared with phospholipids rich in PUFAs. We tested the effect of many antioxidants such as alpha tocopherol, vitamin A, melatonin and its structural analogues, and conjugated linoleic acid, among others. PMID:24490074

  9. Prediction of nucleic acid binding probability in proteins: a neighboring residue network based score.

    PubMed

    Miao, Zhichao; Westhof, Eric

    2015-06-23

    We describe a general binding score for predicting the nucleic acid binding probability in proteins. The score is directly derived from physicochemical and evolutionary features and integrates a residue neighboring network approach. Our process achieves stable and high accuracies on both DNA- and RNA-binding proteins and illustrates how the main driving forces for nucleic acid binding are common. Because of the effective integration of the synergetic effects of the network of neighboring residues and the fact that the prediction yields a hierarchical scoring on the protein surface, energy funnels for nucleic acid binding appear on protein surfaces, pointing to the dynamic process occurring in the binding of nucleic acids to proteins. PMID:25940624

  10. Prediction of nucleic acid binding probability in proteins: a neighboring residue network based score

    PubMed Central

    Miao, Zhichao; Westhof, Eric

    2015-01-01

    We describe a general binding score for predicting the nucleic acid binding probability in proteins. The score is directly derived from physicochemical and evolutionary features and integrates a residue neighboring network approach. Our process achieves stable and high accuracies on both DNA- and RNA-binding proteins and illustrates how the main driving forces for nucleic acid binding are common. Because of the effective integration of the synergetic effects of the network of neighboring residues and the fact that the prediction yields a hierarchical scoring on the protein surface, energy funnels for nucleic acid binding appear on protein surfaces, pointing to the dynamic process occurring in the binding of nucleic acids to proteins. PMID:25940624

  11. Modeling Fatty Acid Transfer from Artery to Cardiomyocyte

    PubMed Central

    Arts, Theo; Reneman, Robert S.; Bassingthwaighte, James B.; van der Vusse, Ger J.

    2015-01-01

    Despite the importance of oxidation of blood-borne long-chain fatty acids (Fa) in the cardiomyocytes for contractile energy of the heart, the mechanisms underlying the transfer of Fa from the coronary plasma to the cardiomyocyte is still incompletely understood. To obtain detailed insight into this transfer process, we designed a novel model of Fa transfer dynamics from coronary plasma through the endothelial cells and interstitium to the cardiomyocyte, applying standard physicochemical principles on diffusion and on the chemical equilibrium of Fa binding to carrier proteins Cp, like albumin in plasma and interstitium and Fatty Acid-Binding Proteins within endothelium and cardiomyocytes. Applying these principles, the present model strongly suggests that in the heart, binding and release of Fa to and from Cp in the aqueous border zones on both sides of the cell membranes form the major hindrance to Fa transfer. Although often considered, the membrane itself appears not to be a significant hindrance to diffusion of Fa. Proteins, residing in the cellular membrane, may facilitate transfer of Fa between Cp and membrane. The model is suited to simulate multiple tracer dilution experiments performed on isolated rabbit hearts administrating albumin and Fa as tracer substances into the coronary arterial perfusion line. Using parameter values on myocardial ultrastructure and physicochemical properties of Fa and Cp as reported in literature, simulated washout curves appear to be similar to the experimentally determined ones. We conclude therefore that the model is realistic and, hence, can be considered as a useful tool to better understand Fa transfer by evaluation of experimentally determined tracer washout curves. PMID:26675003

  12. Roles of N-Terminal Fatty Acid Acylations in Membrane Compartment Partitioning: Arabidopsis h-Type Thioredoxins as a Case Study[C][W

    PubMed Central

    Traverso, José A.; Micalella, Chiara; Martinez, Aude; Brown, Spencer C.; Satiat-Jeunemaître, Béatrice; Meinnel, Thierry; Giglione, Carmela

    2013-01-01

    N-terminal fatty acylations (N-myristoylation [MYR] and S-palmitoylation [PAL]) are crucial modifications affecting 2 to 4% of eukaryotic proteins. The role of these modifications is to target proteins to membranes. Predictive tools have revealed unexpected targets of these acylations in Arabidopsis thaliana and other plants. However, little is known about how N-terminal lipidation governs membrane compartmentalization of proteins in plants. We show here that h-type thioredoxins (h-TRXs) cluster in four evolutionary subgroups displaying strictly conserved N-terminal modifications. It was predicted that one subgroup undergoes only MYR and another undergoes both MYR and PAL. We used plant TRXs as a model protein family to explore the effect of MYR alone or MYR and PAL in the same family of proteins. We used a high-throughput biochemical strategy to assess MYR of specific TRXs. Moreover, various TRX–green fluorescent protein fusions revealed that MYR localized protein to the endomembrane system and that partitioning between this membrane compartment and the cytosol correlated with the catalytic efficiency of the N-myristoyltransferase acting at the N terminus of the TRXs. Generalization of these results was obtained using several randomly selected Arabidopsis proteins displaying a MYR site only. Finally, we demonstrated that a palmitoylatable Cys residue flanking the MYR site is crucial to localize proteins to micropatching zones of the plasma membrane. PMID:23543785

  13. Identification of amino acid residues that determine the substrate specificity of mammalian membrane-bound front-end fatty acid desaturases.

    PubMed

    Watanabe, Kenshi; Ohno, Makoto; Taguchi, Masahiro; Kawamoto, Seiji; Ono, Kazuhisa; Aki, Tsunehiro

    2016-01-01

    Membrane-bound desaturases are physiologically and industrially important enzymes that are involved in the production of diverse fatty acids such as polyunsaturated fatty acids and their derivatives. Here, we identified amino acid residues that determine the substrate specificity of rat Δ6 desaturase (D6d) acting on linoleoyl-CoA by comparing its amino acid sequence with that of Δ5 desaturase (D5d), which converts dihomo-γ-linolenoyl-CoA. The N-terminal cytochrome b5-like domain was excluded as a determinant by domain swapping analysis. Substitution of eight amino acid residues (Ser209, Asn211, Arg216, Ser235, Leu236, Trp244, Gln245, and Val344) of D6d with the corresponding residues of D5d by site-directed mutagenesis switched the substrate specificity from linoleoyl-CoA to dihomo-γ-linolenoyl-CoA. In addition, replacement of Leu323 of D6d with Phe323 on the basis of the amino acid sequence of zebra fish Δ5/6 bifunctional desaturase was found to render D6d bifunctional. Homology modeling of D6d using recent crystal structure data of human stearoyl-CoA (Δ9) desaturase revealed that Arg216, Trp244, Gln245, and Leu323 are located near the substrate-binding pocket. To our knowledge, this is the first report on the structural basis of the substrate specificity of a mammalian front-end fatty acid desaturase, which will aid in efficient production of value-added fatty acids. PMID:26590171

  14. Leukocyte Protease Binding to Nucleic Acids Promotes Nuclear Localization and Cleavage of Nucleic Acid Binding Proteins

    PubMed Central

    Thomas, Marshall P.; Whangbo, Jennifer; McCrossan, Geoffrey; Deutsch, Aaron; Martinod, Kimberly; Walch, Michael; Lieberman, Judy

    2014-01-01

    Killer lymphocyte granzyme (Gzm) serine proteases induce apoptosis of pathogen-infected cells and tumor cells. Many known Gzm substrates are nucleic acid binding proteins, and the Gzms accumulate in the target cell nucleus by an unknown mechanism. Here we show that human Gzms bind to DNA and RNA with nanomolar affinity. Gzms cleave their substrates most efficiently when both are bound to nucleic acids. RNase treatment of cell lysates reduces Gzm cleavage of RNA binding protein (RBP) targets, while adding RNA to recombinant RBP substrates increases in vitro cleavage. Binding to nucleic acids also influences Gzm trafficking within target cells. Pre-incubation with competitor DNA and DNase treatment both reduce Gzm nuclear localization. The Gzms are closely related to neutrophil proteases, including neutrophil elastase (NE) and cathepsin G (CATG). During neutrophil activation, NE translocates to the nucleus to initiate DNA extrusion into neutrophil extracellular traps (NETs), which bind NE and CATG. These myeloid cell proteases, but not digestive serine proteases, also bind DNA strongly and localize to nuclei and NETs in a DNA-dependent manner. Thus, high affinity nucleic acid binding is a conserved and functionally important property specific to leukocyte serine proteases. Furthermore, nucleic acid binding provides an elegant and simple mechanism to confer specificity of these proteases for cleavage of nucleic acid binding protein substrates that play essential roles in cellular gene expression and cell proliferation. PMID:24771851

  15. Cellular Retinoic Acid Binding Protein 2 Is Strikingly Downregulated in Human Esophageal Squamous Cell Carcinoma and Functions as a Tumor Suppressor

    PubMed Central

    Xiao, Weifan; Sun, Fenyong; Yuan, Hong; Pan, Qiuhui

    2016-01-01

    Esophageal squamous cell carcinoma (ESCC) is the predominant pathotype of esophageal carcinoma (EC) in China, especially in Henan province, with poor prognosis and limited 5-year survival rate. Cellular retinoic acid binding protein 2 (CRABP2) is a member of the retinoic acid (RA) and lipocalin/cytosolic fatty-acid binding protein family and plays a completely contrary role in tumorigenesis through the retinoid signaling pathway, depending on the nuclear RA receptors (RAR) and PPARbeta/delta receptors. Presently, the biological role of CRABP2 in the development of ESCC has never been reported. Here, we firstly evaluated the expression of CRABP2 at both mRNA and protein levels and showed that it was remarkably downregulated in clinical ESCC tissues and closely correlated with the occurrence position, pathology, TNM stage, size, infiltration depth and cell differentiation of the tumor. Additionally, the biological function assays demonstrated that CRABP2 acted as a tumor suppressor in esophageal squamous carcinogenesis by significantly inhibiting cell growth, inducing cell apoptosis and blocking cell metastasis both in vitro and in vivo. All in all, our finding simplicate that CRABP2 is possibly an efficient molecular marker for diagnosing and predicting the development of ESCC. PMID:26839961

  16. Free fatty-acid uptake by isolated rat hepatocytes.

    PubMed

    Renaud, G; Bouma, M E; Foliot, A; Infante, R

    1985-11-01

    In isolated rat hepatocytes, the rate of palmitic acid binding and uptake is directly related to the concentration of free fatty acid (FFA) in the medium. After their entry into the cell, FFA are immediately incorporated into cellular phospholipids and triglycerides and no accumulation of free fatty acids can be demonstrated inside the cell. The rate of free fatty-acid uptake remains unchanged after incubation in a 2 mM KCN containing medium, indicating that in the range of fatty-acid concentrations used in this study, this phenomenon does not require energy. PMID:2421669

  17. In Vitro bile acid binding of kale, mustard greens, broccoli, cabbage and green bell pepper improves with microwave cooking

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Bile acid binding potential of foods and food fractions has been related to lowering the risk of heart disease and that of cancer. Sautéing or steam cooking has been observed to significantly improve bile acid binding of green/leafy vegetables. It was hypothesized that microwave cooking could impr...

  18. Effect of polyvinyl alcohol hydrogel as a biocarrier on volatile fatty acids production of a two-stage thermophilic anaerobic membrane bioreactor.

    PubMed

    Chaikasem, Supawat; Abeynayaka, Amila; Visvanathan, Chettiyappan

    2014-09-01

    This work studied the effect of polyvinyl alcohol hydrogel (PVA-gel) beads, as an effective biocarrier for volatile fatty acid (VFA) production in hydrolytic reactor of a two-stage thermophilic anaerobic membrane bioreactor (TAnMBR). The two-stage TAnMBR, treating synthetic high strength particulate wastewater with influent chemical oxygen demand (COD) [16.4±0.8 g/L], was operated at 55 °C. Under steady state conditions, the reactor was operated at an organic loading rate of 8.2±0.4 kg COD/m(3) d. Operational performance of the system was monitored by assessing VFA composition and quantity, methane production and COD removal efficiency. Increment of VFA production was observed with PVA-gel addition. Hydrolytic effluent contained large amount of acetic acid and n-butyric acid. However, increase in VFA production adversely affected the methanogenic reactor performance due to lack of methanogenic archaea. PMID:24803272

  19. The Green Microalga Chlamydomonas reinhardtii Has a Single ω-3 Fatty Acid Desaturase That Localizes to the Chloroplast and Impacts Both Plastidic and Extraplastidic Membrane Lipids1[C][W

    PubMed Central

    Nguyen, Hoa Mai; Cuiné, Stéphan; Beyly-Adriano, Audrey; Légeret, Bertrand; Billon, Emmanuelle; Auroy, Pascaline; Beisson, Fred; Peltier, Gilles; Li-Beisson, Yonghua

    2013-01-01

    The ω-3 polyunsaturated fatty acids account for more than 50% of total fatty acids in the green microalga Chlamydomonas reinhardtii, where they are present in both plastidic and extraplastidic membranes. In an effort to elucidate the lipid desaturation pathways in this model alga, a mutant with more than 65% reduction in total ω-3 fatty acids was isolated by screening an insertional mutant library using gas chromatography-based analysis of total fatty acids of cell pellets. Molecular genetics analyses revealed the insertion of a TOC1 transposon 113 bp upstream of the ATG start codon of a putative ω-3 desaturase (CrFAD7; locus Cre01.g038600). Nuclear genetic complementation of crfad7 using genomic DNA containing CrFAD7 restored the wild-type fatty acid profile. Under standard growth conditions, the mutant is indistinguishable from the wild type except for the fatty acid difference, but when exposed to short-term heat stress, its photosynthesis activity is more thermotolerant than the wild type. A comparative lipidomic analysis of the crfad7 mutant and the wild type revealed reductions in all ω-3 fatty acid-containing plastidic and extraplastidic glycerolipid molecular species. CrFAD7 was localized to the plastid by immunofluorescence in situ hybridization. Transformation of the crfad7 plastidial genome with a codon-optimized CrFAD7 restored the ω-3 fatty acid content of both plastidic and extraplastidic lipids. These results show that CrFAD7 is the only ω-3 fatty acid desaturase expressed in C. reinhardtii, and we discuss possible mechanisms of how a plastid-located desaturase may impact the ω-3 fatty acid content of extraplastidic lipids. PMID:23958863

  20. The Vibrio cholerae Fatty Acid Regulatory Protein, FadR, Represses Transcription of plsB, the Gene Encoding the First Enzyme of Membrane Phospholipid Biosynthesis

    PubMed Central

    Feng, Youjun; Cronan, John E.

    2011-01-01

    SUMMARY Glycerol-3-phosphate (sn-glycerol-3-P, G3P) acyltransferase catalyzes the first committed step in the biosynthesis of membrane phospholipids, the acylation of G3P to form 1-acyl G3P (lysophosphatidic acid). The paradigm G3P acyltransferase is the Escherichia coli plsB gene product which acylates position-1 of G3P using fatty acids in thioester linkage to either acyl carrier protein (ACP) or CoA as acyl-donors. Although the Escherichia coli plsB gene was discovered about 30 years ago, no evidence for transcriptional control of its expression has been reported. However Kazakov and coworkers (Kazakov, A. E. et al. (2009) J Bacteriol, 191, 52–64) reported the presence of a putative FadR-binding site upstream of the candidate plsB genes of V. cholerae and three other Vibrio species suggesting that plsB might be regulated by FadR, a GntR-family transcription factor thus far known only to regulate fatty acid synthesis and degradation. We report that the V. cholerae plsB homologue restored growth of E. coli strain BB26-36 which is a G3P auxotroph due to an altered G3P acyltransferase activity. The plsB promoter was also mapped and the predicted FadR-binding palindrome was found to span positions -19 to -35, upstream of the transcription start site. Gel shift assays confirmed that both V. cholerae FadR and E. coli FadR bound the V. cholerae plsB promoter region and binding was reversed upon addition of long chain fatty acyl-CoA thioesters. The expression level of the V. cholerae plsB gene was elevated 2–3 fold in an E. coli fadR null mutant strain indicating that FadR acts as a repressor of V. cholerae plsB expression. In both E. coli and V. cholerae the β-galactosidase activity of transcriptional fusions of the V. cholerae plsB promoter to lacZ increased 2–3 fold upon supplementation of growth media with oleic acid. Therefore, V. cholerae coordinates fatty acid metabolism with 1-acyl G3P synthesis. PMID:21771112

  1. Recovery of valuable soluble compounds from washing waters generated during small fatty pelagic surimi processing by membrane processes.

    PubMed

    Dumay, J; Radier, S; Barnathan, G; Bergé, J P; Jaouen, P

    2008-04-01

    This work focuses on the treatment of washing waters coming from surimi manufacturing using ultrafiltration technology at a laboratory scale. Four membrane materials (poly-ether sulfone, polyacrilonytrile, poly vinylidene fluoride and regenerated cellulose) and 5 Molecular Weight Cut-Off (from 3 to 100 kDa) have been studied at bench laboratory scale using the pilot Rayflow 100, commercialised by Rhodia Orelis. The investigation deals with the ability for membranes to offer a high retention of biochemical compounds (proteins and lipids). Results obtained during adsorption tests showed that the regenerated cellulose material seems to be the most appropriate with regards to pore size reduction due to the protein-adsorption. During the ultrafiltration of the washing water, the regenerated cellulose material leads to the best results, followed by the polyacrylonitrile and poly-vinylidene fluoride materials. Poor results were obtained with polyether sulfone membrane. Compared to the other materials, the regenerated cellulose is the easiest to regenerate, with minimal cleaning water and no chemical treatment necessary. Biochemical characterization of the fractions generated during the ultrafiltration with the polyacrilonytrile, poly vinylidene fluoride and regenerated cellulose membranes showed that all the membranes provided a high recovery rate of the lipids and proteins. The 10 kDa regenerated cellulose membrane had the highest performance and was further evaluated. With such a treatment, the chemical oxygen demand was reduced by 75%. By performing hydrolysis followed by a centrifugation, biochemical composition of the sludge and liquid fraction were modified, producing an insoluble fraction containing fats and few proteins and a soluble fraction containing proteins and few fats. The sludge, initially insoluble, was mainly solubilized during hydrolysis, and lipids and peptides were concentrated by ultrafiltration. PMID:18619150

  2. Structural Basis of Fatty Acid Substrate Binding to Cyclooxygenase-2*

    PubMed Central

    Vecchio, Alex J.; Simmons, Danielle M.; Malkowski, Michael G.

    2010-01-01

    The cyclooxygenases (COX-1 and COX-2) are membrane-associated heme-containing homodimers that generate prostaglandin H2 from arachidonic acid (AA). Although AA is the preferred substrate, other fatty acids are oxygenated by these enzymes with varying efficiencies. We determined the crystal structures of AA, eicosapentaenoic acid (EPA), and docosahexaenoic acid (DHA) bound to Co3+-protoporphyrin IX-reconstituted murine COX-2 to 2.1, 2.4, and 2.65 Å, respectively. AA, EPA, and docosahexaenoic acid bind in different conformations in each monomer constituting the homodimer in their respective structures such that one monomer exhibits nonproductive binding and the other productive binding of the substrate in the cyclooxygenase channel. The interactions identified between protein and substrate when bound to COX-1 are conserved in our COX-2 structures, with the only notable difference being the lack of interaction of the carboxylate of AA and EPA with the side chain of Arg-120. Leu-531 exhibits a different side chain conformation when the nonproductive and productive binding modes of AA are compared. Unlike COX-1, mutating this residue to Ala, Phe, Pro, or Thr did not result in a significant loss of activity or substrate binding affinity. Determination of the L531F:AA crystal structure resulted in AA binding in the same global conformation in each monomer. We speculate that the mobility of the Leu-531 side chain increases the volume available at the opening of the cyclooxygenase channel and contributes to the observed ability of COX-2 to oxygenate a broad spectrum of fatty acid and fatty ester substrates. PMID:20463020

  3. The Yeast Eukaryotic Translation Initiation Factor 2B Translation Initiation Complex Interacts with the Fatty Acid Synthesis Enzyme YBR159W and Endoplasmic Reticulum Membranes

    PubMed Central

    Browne, Christopher M.; Samir, Parimal; Fites, J. Scott; Villarreal, Seth A.

    2013-01-01

    Using affinity purifications coupled with mass spectrometry and yeast two-hybrid assays, we show the Saccharomyces cerevisiae translation initiation factor complex eukaryotic translation initiation factor 2B (eIF2B) and the very-long-chain fatty acid (VLCFA) synthesis keto-reductase enzyme YBR159W physically interact. The data show that the interaction is specifically between YBR159W and eIF2B and not between other members of the translation initiation or VLCFA pathways. A ybr159wΔ null strain has a slow-growth phenotype and a reduced translation rate but a normal GCN4 response to amino acid starvation. Although YBR159W localizes to the endoplasmic reticulum membrane, subcellular fractionation experiments show that a fraction of eIF2B cofractionates with lipid membranes in a YBR159W-independent manner. We show that a ybr159wΔ yeast strain and other strains with null mutations in the VLCFA pathway cause eIF2B to appear as numerous foci throughout the cytoplasm. PMID:23263984

  4. The N-terminal SH4 region of the Src family kinase Fyn is modified by methylation and heterogeneous fatty acylation: role in membrane targeting, cell adhesion, and spreading.

    PubMed

    Liang, Xiquan; Lu, Yun; Wilkes, Meredith; Neubert, Thomas A; Resh, Marilyn D

    2004-02-27

    The N-terminal SH4 domain of Src family kinases is responsible for promoting membrane binding and plasma membrane targeting. Most Src family kinases contain an N-terminal Met-Gly-Cys consensus sequence that undergoes dual acylation with myristate and palmitate after removal of methionine. Previous studies of Src family kinase fatty acylation have relied on radiolabeling of cells with radioactive fatty acids. Although this method is useful for verifying that a given fatty acid is attached to a protein, it does not reveal whether other fatty acids or other modifying groups are attached to the protein. Here we use matrix-assisted laser desorption/ionization-time of flight (MALDI-TOF) mass spectrometry to identify fatty acylated species of the Src family kinase Fyn. Our results reveal that Fyn is efficiently myristoylated and that some of the myristoylated proteins are also heterogeneously S-acylated with palmitate, palmitoleate, stearate, or oleate. Furthermore, we show for the first time that Fyn is trimethylated at lysine residues 7 and/or 9 within its N-terminal region. Both myristoylation and palmitoylation were required for methylation of Fyn. However, a general methylation inhibitor had no inhibitory effect on myristoylation and palmitoylation of Fyn, suggesting that methylation occurs after myristoylation and palmitoylation. Lysine mutants of Fyn that could not be methylated failed to promote cell adhesion and spreading, suggesting that methylation is important for Fyn function. PMID:14660555

  5. Adaptation of Pseudomonas putida S12 to ethanol and toluene at the level of fatty acid composition of membranes

    SciTech Connect

    Heipieper, H.J.; De Bont, J.A.M.

    1994-12-01

    Many organic solvents are toxic to organisms because the partition preferentially in membranes. However, microorganisms can adapt to different organic substances. In this paper, the authors investigate the effects of both ethanol and toluene on these adaptation mechanisms in Pseudomonas putida S12 and demonstrated that isomerization allows a swifter adaptation than the growth-dependent system, which relies on changes in the degree of saturation. 34 refs., 4 figs.

  6. Evidence linking the Pseudomonas oleovorans alkane omega-hydroxylase, an integral membrane diiron enzyme, and the fatty acid desaturase family.

    PubMed

    Shanklin, John; Whittle, Edward

    2003-06-19

    Pseudomonas oleovorans alkane omega-hydroxylase (AlkB) is an integral membrane diiron enzyme that shares a requirement for iron and oxygen for activity in a manner similar to that of the non-heme integral membrane desaturases, epoxidases, acetylenases, conjugases, ketolases, decarbonylase and methyl oxidases. No overall sequence similarity is detected between AlkB and these desaturase-like enzymes by computer algorithms; however, they do contain a series of histidine residues in a similar relative positioning with respect to hydrophobic regions thought to be transmembrane domains. To test whether these conserved histidine residues are functionally equivalent to those of the desaturase-like enzymes we used scanning alanine mutagenesis to test if they are essential for activity of AlkB. These experiments show that alanine substitution of any of the eight conserved histidines results in complete inactivation, whereas replacement of three non-conserved histidines in close proximity to the conserved residues, results in only partial inactivation. These data provide the first experimental support for the hypotheses: (i) that the histidine motif in AlkB is equivalent to that in the desaturase-like enzymes and (ii) that the conserved histidine residues play a vital role such as coordinating the Fe ions comprising the diiron active site. PMID:12804773

  7. Atomic resolution view into the structure–function relationships of the human myelin peripheral membrane protein P2

    SciTech Connect

    Ruskamo, Salla; Yadav, Ravi P.; Sharma, Satyan; Lehtimäki, Mari; Laulumaa, Saara; Aggarwal, Shweta; Simons, Mikael; Bürck, Jochen; Ulrich, Anne S.; Juffer, André H.; Kursula, Inari; Kursula, Petri

    2014-01-01

    The structure of the human myelin peripheral membrane protein P2 has been refined at 0.93 Å resolution. In combination with functional experiments in vitro, in vivo and in silico, the fine details of the structure–function relationships in P2 are emerging. P2 is a fatty acid-binding protein expressed in vertebrate peripheral nerve myelin, where it may function in bilayer stacking and lipid transport. P2 binds to phospholipid membranes through its positively charged surface and a hydrophobic tip, and accommodates fatty acids inside its barrel structure. The structure of human P2 refined at the ultrahigh resolution of 0.93 Å allows detailed structural analyses, including the full organization of an internal hydrogen-bonding network. The orientation of the bound fatty-acid carboxyl group is linked to the protonation states of two coordinating arginine residues. An anion-binding site in the portal region is suggested to be relevant for membrane interactions and conformational changes. When bound to membrane multilayers, P2 has a preferred orientation and is stabilized, and the repeat distance indicates a single layer of P2 between membranes. Simulations show the formation of a double bilayer in the presence of P2, and in cultured cells wild-type P2 induces membrane-domain formation. Here, the most accurate structural and functional view to date on P2, a major component of peripheral nerve myelin, is presented, showing how it can interact with two membranes simultaneously while going through conformational changes at its portal region enabling ligand transfer.

  8. Molecular cloning, tissue distribution, and expression of a 14-kDa bile acid-binding protein from rat ileal cytosol.

    PubMed Central

    Gong, Y Z; Everett, E T; Schwartz, D A; Norris, J S; Wilson, F A

    1994-01-01

    A cDNA clone encoding the major intestinal cytosolic 14-kDa bile acid-binding protein (14-kDa I-BABP) was isolated from a rat ileal lambda gt22A library following immunoscreening using a monospecific antiserum raised against a 14-kDa polypeptide found in the rat ileal cytosol. One clone of 516 bp encoded a 128-amino acid protein with a predicted molecular mass of 14,544 Da. The deduced amino acid sequence of 14-kDa I-BABP showed 100% homology to rat intestinal 15-kDa protein (I-15P) and 72% homology to porcine 15-kDa gastrotropin, whereas comparison of I-BABP to rat 14-kDa fatty acid-binding proteins of liver, intestine, and heart revealed homologies of 44%, 25%, and 28%, respectively. Northern blot analysis revealed a single transcript of approximately 0.5 kb in ileum and ovary; however, the abundance of I-BABP mRNA was much greater in ileum than in ovary. No transcript was seen in RNA extracted from stomach, jejunum, colon, liver, adrenal, brain, heart, kidney, or testis. Transfection of the I-BABP cDNA into COS-7 cells resulted in the expression of a 14-kDa protein that was identical to the ileal cytosolic I-BABP as determined by immunoblotting. Photoaffinity labeling of expressed 14-kDa protein was saturable with respect to increasing concentrations of 7,7-azo[3H]taurocholate (Km, 83.3 microM; Vmax, 6.7 pmol/mg per 5 min). Taurocholate inhibited 7,7-azotaurocholate labeling by > 96% with lesser inhibition by taurochenodeoxycholate (83.1%), chenodeoxycholate (74.6%), cholate (50.5%), and progesterone (38.5%), whereas oleic acid and estradiol did not inhibit binding. Images PMID:8197128

  9. Liprotides made of α-lactalbumin and cis fatty acids form core-shell and multi-layer structures with a common membrane-targeting mechanism.

    PubMed

    Frislev, Henriette S; Jessen, Christian M; Oliveira, Cristiano L P; Pedersen, Jan Skov; Otzen, Daniel E

    2016-07-01

    α-Lactalbumin (aLA) has been shown to form complexes with oleic acid (OA), which may target cancer cells. We recently showed that aLA and several other proteins all form protein-OA complexes called liprotides with a generic structure consisting of a micellar OA core surrounded by a shell of partially denatured protein. Here we report that a heat treatment and an alkaline treatment method both allow us to prepare liprotide complexes composed of aLA and a range of unsaturated fatty acids (FA), provided the FAs contain cis (but not trans) double bonds. All liprotides containing cis-FA form both small and large species, which all consist of partially denatured aLA, though the overall shape of the species differs. Small liprotides have a simple core-shell structure while the larger liprotides are multi-layered, i.e. they have an additional layer of both FA and aLA surrounding the outside of the core-shell structure. All liprotides can transfer their entire FA content to vesicles, releasing aLA as monomers and softening the lipid membrane. The more similar to OA, the more efficiently the different FAs induce hemolysis. We conclude that aLA can take up and transfer a wide variety of FA to membranes, provided they contain a cis-bond. This highlights liprotides as a general class of complexes where both protein and cis-FA component can be varied without departing from a generic (though sometimes multi-layered) core-shell structure. PMID:27068540

  10. Imaging heterogeneity of membrane and storage lipids in transgenic Camelina sativa seeds with altered fatty acid profiles.

    PubMed

    Horn, Patrick J; Silva, Jillian E; Anderson, Danielle; Fuchs, Johannes; Borisjuk, Ljudmilla; Nazarenus, Tara J; Shulaev, Vladimir; Cahoon, Edgar B; Chapman, Kent D

    2013-10-01

    Engineering compositional changes in oilseeds is typically accomplished by introducing new enzymatic step(s) and/or by blocking or enhancing an existing enzymatic step(s) in a seed-specific manner. However, in practice, the amounts of lipid species that accumulate in seeds are often different from what one would predict from enzyme expression levels, and these incongruences may be rooted in an incomplete understanding of the regulation of seed lipid metabolism at the cellular/tissue level. Here we show by mass spectrometry imaging approaches that triacylglycerols and their phospholipid precursors are distributed differently within cotyledons and the hypocotyl/radicle axis in embryos of the oilseed crop Camelina sativa, indicating tissue-specific heterogeneity in triacylglycerol metabolism. Phosphatidylcholines and triacylglycerols enriched in linoleic acid (C18:2) were preferentially localized to the axis tissues, whereas lipid classes enriched in gadoleic acid (C20:1) were preferentially localized to the cotyledons. Manipulation of seed lipid compositions by heterologous over-expression of an acyl-acyl carrier protein thioesterase, or by suppression of fatty acid desaturases and elongases, resulted in new overall seed storage lipid compositions with altered patterns of distribution of phospholipid and triacylglycerol in transgenic embryos. Our results reveal previously unknown differences in acyl lipid distribution in Camelina embryos, and suggest that this spatial heterogeneity may or may not be able to be changed effectively in transgenic seeds depending upon the targeted enzyme(s)/pathway(s). Further, these studies point to the importance of resolving the location of metabolites in addition to their quantities within plant tissues. PMID:23808562

  11. Bovine adipose triglyceride lipase is not altered and adipocyte fatty acid binding protein is increased by dietary flaxseed

    Technology Transfer Automated Retrieval System (TEKTRAN)

    In this paper, we report the full length coding sequence of bovine ATGL cDNA are reported and analyze its expression in bovine tissues. Similar to human, mouse, and pig ATGL sequences, bovine ATGL has a highly conserved patatin domain that is necessary for lipolytic function in mice and humans. Thi...

  12. Distinct Roles of Urinary Liver-Type Fatty Acid-Binding Protein in Non-Diabetic Patients with Anemia

    PubMed Central

    Imai, Naohiko; Yasuda, Takashi; Kamijo-Ikemori, Atsuko; Shibagaki, Yugo; Kimura, Kenjiro

    2015-01-01

    Background Various stresses including ischemia are known to up-regulate renal L-FABP gene expression and increase the urinary excretion of L-FABP. In diabetic patients with anemia, the urinary excretion of L-FABP is significantly increased. We studied the clinical significance of urinary L-FABP and its relationship with anemia in non-diabetic patients. Subjects and Methods A total of 156 patients were studied in this retrospective cross-sectional analysis. The associations between anemia and urinary L-FABP levels, and the predictors of urinary L-FABP levels in non-diabetic patients were evaluated. Results Urinary L-FABP levels were significantly higher in patients with anemia compared to those in patients without anemia. Similarly, the urinary L-FABP levels were significantly higher in patients with albuminuria compared to those in patients without albuminuria. Urinary L-FABP levels correlated with urinary albumin-to-creatinine ratios, estimated glomerular filtration rates, body mass index, and hemoglobin levels. Multivariate linear regression analysis determined that hemoglobin levels (β = -0.249, P = 0.001) and urinary albumin-to-creatinine ratios (β = 0.349, P < 0.001) were significant predictors of urinary L-FABP levels. Conclusions Urinary L-FABP is strongly associated with anemia in non-diabetic patients. PMID:26010898

  13. Enhanced secretion of glucosyltransferase by changes in potassium ion concentrations is accompanied by an altered pattern of membrane fatty acids in Streptococcus salivarius.

    PubMed Central

    Markevics, L J; Jacques, N A

    1985-01-01

    Growth of Streptococcus salivarius ATCC 25975 in a Na+-based medium containing 1 to 50 mM K+ enhanced extracellular glucosyltransferase production by 3.7-fold over the level of enzyme found in a K+-based medium containing 184 mM K+. Enzyme synthesis and secretion were further enhanced in a nonlinear manner with respect to the concentration of K+ in the medium when cells were cultured from an inoculum grown in the presence of 1 mM K+. This concentration of K+ was the minimum required to maintain a near-maximum growth rate for S. salivarius in medium where K+ was limited. A maximum sevenfold stimulation of glucosyltransferase production occurred at 18 mM K+ under these conditions. Analysis of the total membrane lipids showed that the composition of octadecanoic acid increased with decreasing K+ concentration essentially at the expense of the octadecenoic acid moiety. Extracellular glucosyltransferase production was found to be directly related to the ratio of these two fatty acids. Similar confirmatory results over a greater range of enzyme production were obtained with nonproliferating cell suspensions. PMID:3156125

  14. A Large-Scale Assessment of Nucleic Acids Binding Site Prediction Programs.

    PubMed

    Miao, Zhichao; Westhof, Eric

    2015-12-01

    Computational prediction of nucleic acid binding sites in proteins are necessary to disentangle functional mechanisms in most biological processes and to explore the binding mechanisms. Several strategies have been proposed, but the state-of-the-art approaches display a great diversity in i) the definition of nucleic acid binding sites; ii) the training and test datasets; iii) the algorithmic methods for the prediction strategies; iv) the performance measures and v) the distribution and availability of the prediction programs. Here we report a large-scale assessment of 19 web servers and 3 stand-alone programs on 41 datasets including more than 5000 proteins derived from 3D structures of protein-nucleic acid complexes. Well-defined binary assessment criteria (specificity, sensitivity, precision, accuracy…) are applied. We found that i) the tools have been greatly improved over the years; ii) some of the approaches suffer from theoretical defects and there is still room for sorting out the essential mechanisms of binding; iii) RNA binding and DNA binding appear to follow similar driving forces and iv) dataset bias may exist in some methods. PMID:26681179

  15. Cholesterol-lowering effect of rice bran protein containing bile acid-binding proteins.

    PubMed

    Wang, Jilite; Shimada, Masaya; Kato, Yukina; Kusada, Mio; Nagaoka, Satoshi

    2015-01-01

    Dietary plant protein is well known to reduce serum cholesterol levels. Rice bran is a by-product of rice milling and is a good source of protein. The present study examined whether feeding rats a high-cholesterol diet containing 10% rice bran protein (RBP) for 10 d affected cholesterol metabolism. Rats fed dietary RBP had lower serum total cholesterol levels and increased excretion of fecal steroids, such as cholesterol and bile acids, than those fed dietary casein. In vitro assays showed that RBP strongly bound to taurocholate, and inhibited the micellar solubility of cholesterol, compared with casein. Moreover, the bile acid-binding proteins of the RBP were eluted by a chromatographic column conjugated with cholic acid, and one of them was identified as hypothetical protein OsJ_13801 (NCBI accession No. EAZ29742) using MALDI-TOF mass spectrometry analysis. These results suggest that the hypocholesterolemic action of the RBP may be caused by the bile acid-binding proteins. PMID:25374002

  16. A Large-Scale Assessment of Nucleic Acids Binding Site Prediction Programs

    PubMed Central

    Miao, Zhichao; Westhof, Eric

    2015-01-01

    Computational prediction of nucleic acid binding sites in proteins are necessary to disentangle functional mechanisms in most biological processes and to explore the binding mechanisms. Several strategies have been proposed, but the state-of-the-art approaches display a great diversity in i) the definition of nucleic acid binding sites; ii) the training and test datasets; iii) the algorithmic methods for the prediction strategies; iv) the performance measures and v) the distribution and availability of the prediction programs. Here we report a large-scale assessment of 19 web servers and 3 stand-alone programs on 41 datasets including more than 5000 proteins derived from 3D structures of protein-nucleic acid complexes. Well-defined binary assessment criteria (specificity, sensitivity, precision, accuracy…) are applied. We found that i) the tools have been greatly improved over the years; ii) some of the approaches suffer from theoretical defects and there is still room for sorting out the essential mechanisms of binding; iii) RNA binding and DNA binding appear to follow similar driving forces and iv) dataset bias may exist in some methods. PMID:26681179

  17. Bile acid binding capacity of fish protein hydrolysates from discard species of the West Mediterranean Sea.

    PubMed

    Pérez-Gálvez, Raúl; García-Moreno, Pedro J; Morales-Medina, Rocío; Guadix, Antonio; Guadix, Emilia M

    2015-04-01

    Fish protein hydrolysates (FPH), produced from the six main discard species from the West Mediterranean Sea (sardine, horse mackerel, axillary seabream, bogue, small-spotted catshark and blue whiting) were tested for their bile acid binding capacity. This capacity is directly linked to the ability to inhibit bile reabsorption in the ileum and therefore to lower cholesterol levels in the bloodstream. From each species, FPH were obtained by three different enzymatic treatments employing two serine endoproteases (subtilisin and trypsin) sequentially or in combination. The results show statistically significant differences among the fish species, attaining interesting average values of bile acid binding capacity for blue whiting (27.32% relative to cholestyramine on an equal protein basis) and horse mackerel (27.42% relative to cholestyramine on an equal protein basis). The enzymatic treatments did not significantly affect the ability of a given species to bind bile acids. These results are similar to other protein sources, such as soy protein or casein, of proven hypocholesterolemic effect. It can be concluded that fish protein hydrolysates from these discard species are suitable as ingredients in the formulation of cholesterol-lowering supplements. PMID:25756593

  18. Two Arginine Residues of Streptococcus gordonii Sialic Acid-Binding Adhesin Hsa Are Essential for Interaction to Host Cell Receptors

    PubMed Central

    Urano-Tashiro, Yumiko; Takahashi, Yukihiro; Oguchi, Riyo; Konishi, Kiyoshi

    2016-01-01

    Hsa is a large, serine-rich protein of Streptococcus gordonii DL1 that mediates binding to α2-3-linked sialic acid termini of glycoproteins, including platelet glycoprotein Ibα, and erythrocyte membrane protein glycophorin A, and band 3. The binding of Hsa to platelet glycoprotein Ibα contributes to the pathogenesis of infective endocarditis. This interaction appears to be mediated by a second non-repetitive region (NR2) of Hsa. However, the molecular details of the interaction between the Hsa NR2 region and these glycoproteins are not well understood. In the present study, we identified the amino acid residues of the Hsa NR2 region that are involved in sialic acid recognition. To identify the sialic acid-binding site of Hsa NR2 region, we prepared various mutants of Hsa NR2 fused with glutathione transferase. Fusion proteins harboring Arg340 to Asn (R340N) or Arg365 to Asn (R365N) substitutions in the NR2 domain exhibited significantly reduced binding to human erythrocytes and platelets. A sugar-binding assay showed that these mutant proteins abolished binding to α2-3-linked sialic acid. Furthermore, we established S. gordonii DL1 derivatives that encoded the corresponding Hsa mutant protein. In whole-cell assays, these mutant strains showed significant reductions in hemagglutination, in platelet aggregation, and in adhesion to human leukocytes. These results indicate that the Arg340 and Arg365 residues of Hsa play an important role in the binding of Hsa to α2-3-linked sialic acid-containing glycoproteins. PMID:27101147

  19. Two Arginine Residues of Streptococcus gordonii Sialic Acid-Binding Adhesin Hsa Are Essential for Interaction to Host Cell Receptors.

    PubMed

    Urano-Tashiro, Yumiko; Takahashi, Yukihiro; Oguchi, Riyo; Konishi, Kiyoshi

    2016-01-01

    Hsa is a large, serine-rich protein of Streptococcus gordonii DL1 that mediates binding to α2-3-linked sialic acid termini of glycoproteins, including platelet glycoprotein Ibα, and erythrocyte membrane protein glycophorin A, and band 3. The binding of Hsa to platelet glycoprotein Ibα contributes to the pathogenesis of infective endocarditis. This interaction appears to be mediated by a second non-repetitive region (NR2) of Hsa. However, the molecular details of the interaction between the Hsa NR2 region and these glycoproteins are not well understood. In the present study, we identified the amino acid residues of the Hsa NR2 region that are involved in sialic acid recognition. To identify the sialic acid-binding site of Hsa NR2 region, we prepared various mutants of Hsa NR2 fused with glutathione transferase. Fusion proteins harboring Arg340 to Asn (R340N) or Arg365 to Asn (R365N) substitutions in the NR2 domain exhibited significantly reduced binding to human erythrocytes and platelets. A sugar-binding assay showed that these mutant proteins abolished binding to α2-3-linked sialic acid. Furthermore, we established S. gordonii DL1 derivatives that encoded the corresponding Hsa mutant protein. In whole-cell assays, these mutant strains showed significant reductions in hemagglutination, in platelet aggregation, and in adhesion to human leukocytes. These results indicate that the Arg340 and Arg365 residues of Hsa play an important role in the binding of Hsa to α2-3-linked sialic acid-containing glycoproteins. PMID:27101147

  20. In situ assay of fatty acid β-oxidation by metabolite profiling following permeabilization of cell membranes[S

    PubMed Central

    Ensenauer, Regina; Fingerhut, Ralph; Schriever, Sonja C.; Fink, Barbara; Becker, Marc; Sellerer, Nina C.; Pagel, Philipp; Kirschner, Andreas; Dame, Torsten; Olgemöller, Bernhard; Röschinger, Wulf; Roscher, Adelbert A.

    2012-01-01

    Quantitative analysis of mitochondrial FA β-oxidation (FAO) has drawn increasing interest for defining lipid-induced metabolic dysfunctions, such as in obesity-induced insulin resistance, and evaluating pharmacologic strategies to improve β-oxidation function. The aim was to develop a new assay to quantify β-oxidation function in intact mitochondria and with a low amount of cell material. Cell membranes of primary human fibroblasts were permeabilized with digitonin prior to a load with FFA substrate. Following 120 min of incubation, the various generated acylcarnitines were extracted from both cells and incubation medium by protein precipitation/desalting and subjected to solid-phase extraction. A panel of 30 acylcarnitines per well was quantified by MS/MS and normalized to citrate synthase activity to analyze mitochondrial metabolite flux. Pretreatment with bezafibrate and etomoxir revealed stimulating and inhibiting regulatory effects on β-oxidation function, respectively. In addition to the advantage of a much shorter assay time due to in situ permeabilization compared with whole-cell incubation systems, the method allows the detection of multiple acylcarnitines from an only limited amount of intact cells, particularly relevant to the use of primary cells. This novel approach facilitates highly sensitive, simple, and fast monitoring of pharmacological effects on FAO. PMID:22345709

  1. Changes in mitochondrial oxidative capacities during thermal acclimation of rainbow trout Oncorhynchus mykiss: roles of membrane proteins, phospholipids and their fatty acid compositions.

    PubMed

    Kraffe, Edouard; Marty, Yanic; Guderley, Helga

    2007-01-01

    diacylphosphatidylethanolamine/diacylphosphatidylcholine (diacylPE/diacylPC), whereas warm acclimation led to restructuring of fatty acids (FA) and to increases of plasmalogen forms of PE and PC. Modification of overall membrane unsaturation did not appear to be the primary aim of restructuring membrane FA during warm acclimation, as total mitochondrial phospholipids and the major phospholipid classes only showed slight shifts of their acyl composition with warm acclimation. On the other hand, natural lysophosphatidylcholine (LysoPC) showed dramatic changes in FA content, as 16:0 and 18:1n-9 doubled whereas 22:6n-3 decreased from around 50% to 32% in warm acclimated trout. Similarly, in cardiolipin (CL), the levels of 16:0 and 18:1n-7 halved while 18:2n-6 increased to over 20% of the FA with warm acclimation. Given the central role of CL in modulating the activity of CCO, F(0)F(1)-ATPase and ANT, these changes suggest that specific compositional changes in CL are important modulators of mitochondrial capacities. The many structural changes in membrane lipids contrast with the limited modifications of the membrane protein components examined and support the concept of lipid structure modulating mitochondrial capacities. PMID:17170158

  2. Point mutations in the S protein connect the sialic acid binding activity with the enteropathogenicity of transmissible gastroenteritis coronavirus.

    PubMed Central

    Krempl, C; Schultze, B; Laude, H; Herrler, G

    1997-01-01

    Enteropathogenic transmissible gastroenteritis virus (TGEV), a porcine coronavirus, is able to agglutinate erythrocytes because of sialic acid binding activity. Competitive inhibitors that may mask the sialic acid binding activity can be inactivated by sialidase treatment of virions. Here, we show that TGEV virions with efficient hemagglutinating activity were also obtained when cells were treated with sialidase prior to infection. This method was used to analyze TGEV mutants for hemagglutinating activity. Recently, mutants with strongly reduced enteropathogenicity that have point mutations or a deletion of four amino acids within residues 145 to 155 of the S protein have been described. Here, we show that in addition to their reduced pathogenicity, these mutants also have lost hemagglutinating activity. These results connect sialic acid binding activity with the enteropathogenicity of TGEV. PMID:9060696

  3. Neurologic syndrome associated with homozygous mutation at MAG sialic acid binding site.

    PubMed

    Roda, Ricardo H; FitzGibbon, Edmond J; Boucekkine, Houda; Schindler, Alice B; Blackstone, Craig

    2016-08-01

    The MAG gene encodes myelin-associated glycoprotein (MAG), an abundant protein involved in axon-glial interactions and myelination during nerve regeneration. Several members of a consanguineous family with a clinical syndrome reminiscent of Pelizaeus-Merzbacher disease and demyelinating leukodystrophy on brain MRI were recently found to harbor a homozygous missense p.Ser133Arg MAG mutation. Here, we report two brothers from a nonconsanguineous family afflicted with progressive cognitive impairment, neuropathy, ataxia, nystagmus, and gait disorder. Exome sequencing revealed the homozygous missense mutation p.Arg118His in MAG. This Arg118 residue in immunoglobulin domain 1 is critical for sialic acid binding, providing a compelling mechanistic basis for disease pathogenesis. PMID:27606346

  4. Effects of membrane polyunsaturated fatty acids on opiate peptide inhibition of basal and prostaglandin E1-stimulated cyclic AMP formation in intact N1E-115 neuroblastoma cells.

    PubMed

    Murphy, M G; Moak, C M; Rao, B G

    1987-12-01

    The effects of membrane polyunsaturated fatty acids (PUFA) on opiate peptide-mediated inhibition of basal and prostaglandin E1-stimulated cyclic AMP formation were examined in intact N1E-115 neuroblastoma cells. Addition of opiate peptides such as methionine 5-enkephalin (metEnk) to control cultures and to cultures that had been supplemented for 48 hr with 50 microM linoleic acid resulted in dose-dependent decreases in cAMP formation; these decreases were blocked by naloxone. Maximum inhibition of basal cyclase activity was 50-55% in both control and PUFA-enriched cells; however, half-maximal inhibition required ten times more metEnk in supplemented cultures than in controls. This is consistent with our observation that the affinity of binding of [tyrosyl-3',5'-3H(N)](2-D-alanine-5-D-leucine)enkephalin ([3H]DADLE) to intact PUFA-enriched cells was lower than that to control cells. Receptor density was not modified as a result of supplementation. Addition of prostaglandin E1 (PGE1) to the cells produced rapid dose-dependent increases in cAMP formation. Maximum responses were higher in PUFA-enriched than in control cells (1924 and 972 pmol cAMP formed/mg protein respectively). Also, the apparent value for EC50 for PGE1 was consistently lower in supplemented cultures. MetEnk reduced PGE1-stimulated cAMP formation by 45-55% in both control and supplemented cells, and values for IC50 were similar (approximately 30 nM) in both. In the presence of the opiate peptide, values for EC50 for PGE1 were similar in control and PUFA-enriched cultures (0.07 and 0.09 microM respectively). The data from these studies suggest that membrane PUFA increase the efficiency of coupling of receptors that stimulate cAMP formation and decrease the efficiency of those that mediate inhibition. PMID:2825714

  5. A transient increase in lipid peroxidation primes preadipocytes for delayed mitochondrial inner membrane permeabilization and ATP depletion during prolonged exposure to fatty acids.

    PubMed

    Rogers, Carlyle; Davis, Barbara; Neufer, P Darrell; Murphy, Michael P; Anderson, Ethan J; Robidoux, Jacques

    2014-02-01

    Preadipocytes are periodically subjected to fatty acid (FA) concentrations that are potentially cytotoxic. We tested the hypothesis that prolonged exposure of preadipocytes of human origin to a physiologically relevant mix of FAs leads to mitochondrial inner membrane (MIM) permeabilization and ultimately to mitochondrial crisis. We found that exposure of preadipocytes to FAs led to progressive cyclosporin A-sensitive MIM permeabilization, which in turn caused a reduction in MIM potential, oxygen consumption, and ATP synthetic capacity and, ultimately, death. Additionally, we showed that FAs induce a transient increase in intramitochondrial reactive oxygen species (ROS) and lipid peroxide production, lasting roughly 30 and 120min for the ROS and lipid peroxides, respectively. MIM permeabilization and its deleterious consequences including mitochondrial crisis and cell death were prevented by treating the cells with the mitochondrial FA uptake inhibitor etomoxir, the mitochondrion-selective superoxide and lipid peroxide antioxidants MitoTempo and MitoQ, or the lipid peroxide and reactive carbonyl scavenger l-carnosine. FAs also promoted a delayed oxidative stress phase. However, the beneficial effects of etomoxir, MitoTempo, and l-carnosine were lost by delaying the treatment by 2h, suggesting that the initial phase was sufficient to prime the cells for the delayed MIM permeabilization and mitochondrial crisis. It also suggested that the second ROS production phase is a consequence of this loss in mitochondrial health. Altogether, our data suggest that approaches designed to diminish intramitochondrial ROS or lipid peroxide accumulation, as well as MIM permeabilization, are valid mechanism-based therapeutic avenues to prevent the loss in preadipocyte metabolic fitness associated with prolonged exposure to elevated FA levels. PMID:24269897

  6. Structure of human NAPE-PLD: regulation of fatty-acid ethanolamide biosynthesis by bile acids

    PubMed Central

    Magotti, Paola; Bauer, Inga; Igarashi, Miki; Babagoli, Masih; Marotta, Roberto; Piomelli, Daniele; Garau, Gianpiero

    2015-01-01

    SUMMARY The fatty-acid ethanolamides (FAEs) are lipid mediators present in all organisms and involved in highly conserved biological functions such as innate immunity, energy balance and stress control. They are produced from membrane N-acylphosphatidylethanolamines (NAPEs) and include agonists for G protein-coupled receptors (e.g. cannabinoid receptors) and nuclear receptors (e.g. PPAR-α). Here we report the crystal structure of human NAPE-hydrolyzing phospholipase D (NAPE-PLD) at 2.65 Å resolution, a membrane enzyme that catalyzes FAE formation in mammals. NAPE-PLD forms homodimers partly separated by an internal ~9 Å-wide channel and uniquely adapted to associate with phospholipids. A hydrophobic cavity provides an entryway for NAPE into the active site, where a binuclear Zn2+ center orchestrates its hydrolysis. Bile acids bind with high affinity to selective pockets in this cavity, enhancing dimer assembly and enabling catalysis. These elements offer multiple targets for the design of small-molecule NAPE-PLD modulators with potential applications in inflammation and metabolic disorders. PMID:25684574

  7. Effect of the long-term feeding of dietary lipids on the learning ability, fatty acid composition of brain stem phospholipids and synaptic membrane fluidity in adult mice: a comparison of sardine oil diet with palm oil diet.

    PubMed

    Suzuki, H; Park, S J; Tamura, M; Ando, S

    1998-03-16

    The effect of 12 month feeding of 5% palm oil or sardine oil diet on the maze-learning ability, fatty acid composition of brain stem phospholipids and synaptic membrane fluidity in mice was studied. The time required to reach the maze exit and the number of times that a mouse strayed into blind alleys in the maze were measured three times every 4 days. The time and number of mice fed on the sardine oil diet were less than those of animals fed on the palm oil diet in the first and second trials. The results of fatty acid composition analysis of brain stem phosphatidylethanolamine showed that the percentage of docosahexaenoic acid (22:6, n-3; DHA) was higher, but the arachidonic acid (20:4, n-6; AA) and docosatetraenoic acid (22:4, n-6; DTA) were lower in the sardine oil diet fed-mice than in the palm oil diet fed-animals. Moreover, the microviscosity of the synaptic plasma membrane in the sardine oil diet group was lower than that in the palm oil diet group. These results suggest that the adult mice fed on the sardine oil diet for a long period maintain higher levels of docosahe xaenoic acid in brain phospholipids, synaptic membrane fluidity and maze-learning ability than animals fed on the palm oil diet. PMID:9593318

  8. Steam Cooking Significantly Improves in Vitro Bile Acid Binding of Beets, Eggplant, Asparagus, Carrots, Green Beans and Cauliflower

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The relative healthful potential of cooked beets, okra, eggplant, asparagus, carrots, green beans, cauliflower and turnips was evaluated by determining their in vitro bile acid binding using a mixture of bile acids secreted in human bile at a duodenal physiological pH of 6.3. Six treatments and two...

  9. BILE ACIDS REGULATE THE ONTOGENIC EXPRESSION OF ILEAL BILE ACID BINDING PROTEIN IN THE RAT VIA THE FARNESOID X RECEPTOR

    Technology Transfer Automated Retrieval System (TEKTRAN)

    In the rat, an increase in ileal bile acid binding protein (IBABP) expression occurs during the third postnatal week. In vitro studies suggest that bile acids (BAs) increase IBABP transcription by activating the BA receptor, farnesoid X receptor (FXR). Thus, we investigated the role of BAs on the on...

  10. Health promoting potential of cereals, grain fractions and beans as determined by their in vitro bile acid binding

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Health promoting potential (Cholesterol lowering and cancer risk reduction) of foods have been determined by in-vitro bile acid binding under physiological conditions. Lowered bile acids result in reduced fat absorption, conversion of cholesterol to bile acids and reduced cancer causing secondary b...

  11. Interaction of enterocyte FABPs with phospholipid membranes: clues for specific physiological roles.

    PubMed

    Falomir-Lockhart, Lisandro J; Franchini, Gisela R; Guerbi, María Ximena; Storch, Judith; Córsico, Betina

    2011-01-01

    Intestinal and liver fatty acid binding proteins (IFABP and LFABP, respectively) are cytosolic soluble proteins with the capacity to bind and transport hydrophobic ligands between different sub-cellular compartments. Their functions are still not clear but they are supposed to be involved in lipid trafficking and metabolism, cell growth, and regulation of several other processes, like cell differentiation. Here we investigated the interaction of these proteins with different models of phospholipid membrane vesicles in order to achieve further insight into their specificity within the enterocyte. A combination of biophysical and biochemical techniques allowed us to determine affinities of these proteins to membranes, the way phospholipid composition and vesicle size and curvature modulate such interaction, as well as the effect of protein binding on the integrity of the membrane structure. We demonstrate here that, besides their apparently opposite ligand transfer mechanisms, both LFABP and IFABP are able to interact with phospholipid membranes, but the factors that modulate such interactions are different for each protein, further implying different roles for IFABP and LFABP in the intracellular context. These results contribute to the proposed central role of intestinal FABPs in the lipid traffic within enterocytes as well as in the regulation of more complex cellular processes. PMID:21539932

  12. Effects of alpha-linolenic acid vs. docosahexaenoic acid supply on the distribution of fatty acids among the rat cardiac subcellular membranes after a short- or long-term dietary exposure

    PubMed Central

    Brochot, Amandine; Guinot, Marine; Auchere, Daniel; Macaire, Jean-Paul; Weill, Pierre; Grynberg, Alain; Rousseau-Ralliard, Delphine

    2009-01-01

    Background Previous work showed that the functional cardiac effect of dietary alpha-linolenic acid (ALA) in rats requires a long feeding period (6 months), although a docosahexaenoic (DHA) acid-supply affects cardiac adrenergic response after 2 months. However, the total cardiac membrane n-3 polyunsaturated fatty acid (PUFA) composition remained unchanged after 2 months. This delay could be due to a specific reorganization of the different subcellular membrane PUFA profiles. This study was designed to investigate the evolution between 2 and 6 months of diet duration of the fatty acid profile in sarcolemmal (SL), mitochondrial (MI), nuclear (NU) and sarcoplasmic reticulum (SR) membrane fractions. Methods Male Wistar rats were randomly assigned to 3 dietary groups (n = 10/diet/period), either n-3 PUFA-free diet (CTL), or ALA or DHA-rich diets. After 2 or 6 months, the subcellular cardiac membrane fractions were separated by differential centrifugations and sucrose gradients. Each membrane profile was analysed by gas chromatography (GC) after lipid extraction. Results As expected the n-3 PUFA-rich diets incorporated n-3 PUFA instead of n-6 PUFA in all the subcellular fractions, which also exhibited individual specificities. The diet duration increased SFA and decreased PUFA in SL, whereas NU remained constant. The SR and MI enriched in n-3 PUFA exhibited a decreased DHA level with ageing in the DHA and CTL groups. Conversely, the n-3 PUFA level remained unchanged in the ALA group, due to a significant increase in docosapentaenoic acid (DPA). N-3 PUFA rich diets lead to a better PUFA profile in all the fractions and significantly prevent the profile modifications induced by ageing. Conclusion With the ALA diet the n-3 PUFA content, particularly in SR and SL kept increasing between 2 and 6 months, which may partly account for the delay to achieve the modification of adrenergic response. PMID:19320987

  13. Insertional Inactivation of Branched-Chain α-Keto Acid Dehydrogenase in Staphylococcus aureus Leads to Decreased Branched-Chain Membrane Fatty Acid Content and Increased Susceptibility to Certain Stresses▿

    PubMed Central

    Singh, Vineet K.; Hattangady, Dipti S.; Giotis, Efstathios S.; Singh, Atul K.; Chamberlain, Neal R.; Stuart, Melissa K.; Wilkinson, Brian J.

    2008-01-01

    Staphylococcus aureus is a major community and nosocomial pathogen. Its ability to withstand multiple stress conditions and quickly develop resistance to antibiotics complicates the control of staphylococcal infections. Adaptation to lower temperatures is a key for the survival of bacterial species outside the host. Branched-chain α-keto acid dehydrogenase (BKD) is an enzyme complex that catalyzes the early stages of branched-chain fatty acid (BCFA) production. In this study, BKD was inactivated, resulting in reduced levels of BCFAs in the membrane of S. aureus. Growth of the BKD-inactivated mutant was progressively more impaired than that of wild-type S. aureus with decreasing temperature, to the point that the mutant could not grow at 12°C. The growth of the mutant was markedly stimulated by the inclusion of 2-methylbutyrate in the growth medium at all temperatures tested. 2-Methylbutyrate is a precursor of odd-numbered anteiso fatty acids and bypasses BKD. Interestingly, growth of wild-type S. aureus was also stimulated by including 2-methylbutyrate in the medium, especially at lower temperatures. The anteiso fatty acid content of the BKD-inactivated mutant was restored by the inclusion of 2-methylbutyrate in the medium. Fluorescence polarization measurements indicated that the membrane of the BKD-inactivated mutant was significantly less fluid than that of wild-type S. aureus. Consistent with this result, the mutant showed decreased toluene tolerance that could be increased by the inclusion of 2-methylbutyrate in the medium. The BKD-inactivated mutant was more susceptible to alkaline pH and oxidative stress conditions. Inactivation of the BKD enzyme complex in S. aureus also led to a reduction in adherence of the mutant to eukaryotic cells and its survival in a mouse host. In addition, the mutant offers a tool to study the role of membrane fluidity in the interaction of S. aureus with antimicrobial substances. PMID:18689519

  14. Fatty acids - trans fatty acids

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The data supporting a negative effect of dietary trans fatty acids on cardiovascular disease risk is consistent. The primary dietary sources of trans fatty acids include partially hydrogenated fat and rudiment fat. The adverse effect of trans fatty acids on plasma lipoprotein profiles is consisten...

  15. Nucleic acid-binding properties of the RRM-containing protein RDM1

    SciTech Connect

    Hamimes, Samia; Bourgeon, Dominique; Stasiak, Alicja Z.; Stasiak, Andrzej; Van Dyck, Eric . E-mail: Vandyck@iarc.fr

    2006-05-26

    RDM1 (RAD52 Motif 1) is a vertebrate protein involved in the cellular response to the anti-cancer drug cisplatin. In addition to an RNA recognition motif, RDM1 contains a small amino acid motif, named RD motif, which it shares with the recombination and repair protein, RAD52. RDM1 binds to single- and double-stranded DNA, and recognizes DNA distortions induced by cisplatin adducts in vitro. Here, we have performed an in-depth analysis of the nucleic acid-binding properties of RDM1 using gel-shift assays and electron microscopy. We show that RDM1 possesses acidic pH-dependent DNA-binding activity and that it binds RNA as well as DNA, and we present evidence from competition gel-shift experiments that RDM1 may be capable of discrimination between the two nucleic acids. Based on reported studies of RAD52, we have generated an RDM1 variant mutated in its RD motif. We find that the L{sub 119}GF {sup {yields}} AAA mutation affects the mode of RDM1 binding to single-stranded DNA.

  16. Structural determinants of human APOBEC3A enzymatic and nucleic acid binding properties

    PubMed Central

    Mitra, Mithun; Hercík, Kamil; Byeon, In-Ja L.; Ahn, Jinwoo; Hill, Shawn; Hinchee-Rodriguez, Kathyrn; Singer, Dustin; Byeon, Chang-Hyeock; Charlton, Lisa M.; Nam, Gabriel; Heidecker, Gisela; Gronenborn, Angela M.; Levin, Judith G.

    2014-01-01

    Human APOBEC3A (A3A) is a single-domain cytidine deaminase that converts deoxycytidine residues to deoxyuridine in single-stranded DNA (ssDNA). It inhibits a wide range of viruses and endogenous retroelements such as LINE-1, but it can also edit genomic DNA, which may play a role in carcinogenesis. Here, we extend our recent findings on the NMR structure of A3A and report structural, biochemical and cell-based mutagenesis studies to further characterize A3A’s deaminase and nucleic acid binding activities. We find that A3A binds ssRNA, but the RNA and DNA binding interfaces differ and no deamination of ssRNA is detected. Surprisingly, with only one exception (G105A), alanine substitution mutants with changes in residues affected by specific ssDNA binding retain deaminase activity. Furthermore, A3A binds and deaminates ssDNA in a length-dependent manner. Using catalytically active and inactive A3A mutants, we show that the determinants of A3A deaminase activity and anti-LINE-1 activity are not the same. Finally, we demonstrate A3A’s potential to mutate genomic DNA during transient strand separation and show that this process could be counteracted by ssDNA binding proteins. Taken together, our studies provide new insights into the molecular properties of A3A and its role in multiple cellular and antiviral functions. PMID:24163103

  17. Affinity regression predicts the recognition code of nucleic acid binding proteins

    PubMed Central

    Pelossof, Raphael; Singh, Irtisha; Yang, Julie L.; Weirauch, Matthew T.; Hughes, Timothy R.; Leslie, Christina S.

    2016-01-01

    Predicting the affinity profiles of nucleic acid-binding proteins directly from the protein sequence is a major unsolved problem. We present a statistical approach for learning the recognition code of a family of transcription factors (TFs) or RNA-binding proteins (RBPs) from high-throughput binding assays. Our method, called affinity regression, trains on protein binding microarray (PBM) or RNA compete experiments to learn an interaction model between proteins and nucleic acids, using only protein domain and probe sequences as inputs. By training on mouse homeodomain PBM profiles, our model correctly identifies residues that confer DNA-binding specificity and accurately predicts binding motifs for an independent set of divergent homeodomains. Similarly, learning from RNA compete profiles for diverse RBPs, our model can predict the binding affinities of held-out proteins and identify key RNA-binding residues. More broadly, we envision applying our method to model and predict biological interactions in any setting where there is a high-throughput ‘affinity’ readout. PMID:26571099

  18. Dynamics of the Peripheral Membrane Protein P2 from Human Myelin Measured by Neutron Scattering—A Comparison between Wild-Type Protein and a Hinge Mutant

    PubMed Central

    Laulumaa, Saara; Nieminen, Tuomo; Lehtimäki, Mari; Aggarwal, Shweta; Simons, Mikael; Koza, Michael M.; Vattulainen, Ilpo; Kursula, Petri; Natali, Francesca

    2015-01-01

    Myelin protein P2 is a fatty acid-binding structural component of the myelin sheath in the peripheral nervous system, and its function is related to its membrane binding capacity. Here, the link between P2 protein dynamics and structure and function was studied using elastic incoherent neutron scattering (EINS). The P38G mutation, at the hinge between the β barrel and the α-helical lid, increased the lipid stacking capacity of human P2 in vitro, and the mutated protein was also functional in cultured cells. The P38G mutation did not change the overall structure of the protein. For a deeper insight into P2 structure-function relationships, information on protein dynamics in the 10 ps to 1 ns time scale was obtained using EINS. Values of mean square displacements mainly from protein H atoms were extracted for wild-type P2 and the P38G mutant and compared. Our results show that at physiological temperatures, the P38G mutant is more dynamic than the wild-type P2 protein, especially on a slow 1-ns time scale. Molecular dynamics simulations confirmed the enhanced dynamics of the mutant variant, especially within the portal region in the presence of bound fatty acid. The increased softness of the hinge mutant of human myelin P2 protein is likely related to an enhanced flexibility of the portal region of this fatty acid-binding protein, as well as to its interactions with the lipid bilayer surface requiring conformational adaptations. PMID:26068118

  19. Towards the elucidation of molecular determinants of cooperativity in the liver bile acid binding protein.

    PubMed

    Pedò, Massimo; D'Onofrio, Mariapina; Ferranti, Pasquale; Molinari, Henriette; Assfalg, Michael

    2009-11-15

    Bile acid binding proteins (BABPs) are cytosolic lipid chaperones contributing to the maintenance of bile acid homeostasis and functional distribution within the cell. Liver BABPs act in parallel with ileal transporters to ensure vectorial transport of bile salts in hepatocytes and enterocytes, respectively. We describe the investigation of ligand binding to liver BABP, an essential step in the understanding of intracellular bile salt transport. Binding site occupancies were monitored in NMR titration experiments using (15)N-labelled ligand, while the relative populations of differently bound BABP forms were assessed by mass spectrometry. This site-specific information allowed the determination of intrinsic thermodynamic parameters and the identification of an extremely high cooperativity between two binding sites. Protein-observed NMR experiments revealed a global structural rearrangement which suggests an allosteric mechanism at the basis of the observed cooperativity. The view of a molecular tool capable of buffering against significant concentrations of free bile salts in a large range of solution conditions emerges from the observed pH-dependence of binding. We set to determine the molecular determinants of cooperativity by analysing the binding properties of a protein containing a mutated internal histidine. Both mass spectrometry and NMR experiments are consistent with an overall decreased binding affinity of the mutant, while the measured diffusion coefficients of ligand species reveal that the affinity loss concerns essentially one of the two binding sites. We therefore identified a mutation able to disrupt energetic communication functional to efficient binding and conclude that the buried histidine establishes contacts that stabilize the ternary complex. PMID:19603488

  20. Disulfide bridge regulates ligand-binding site selectivity in liver bile acid-binding proteins.

    PubMed

    Cogliati, Clelia; Tomaselli, Simona; Assfalg, Michael; Pedò, Massimo; Ferranti, Pasquale; Zetta, Lucia; Molinari, Henriette; Ragona, Laura

    2009-10-01

    Bile acid-binding proteins (BABPs) are cytosolic lipid chaperones that play central roles in driving bile flow, as well as in the adaptation to various pathological conditions, contributing to the maintenance of bile acid homeostasis and functional distribution within the cell. Understanding the mode of binding of bile acids with their cytoplasmic transporters is a key issue in providing a model for the mechanism of their transfer from the cytoplasm to the nucleus, for delivery to nuclear receptors. A number of factors have been shown to modulate bile salt selectivity, stoichiometry, and affinity of binding to BABPs, e.g. chemistry of the ligand, protein plasticity and, possibly, the formation of disulfide bridges. Here, the effects of the presence of a naturally occurring disulfide bridge on liver BABP ligand-binding properties and backbone dynamics have been investigated by NMR. Interestingly, the disulfide bridge does not modify the protein-binding stoichiometry, but has a key role in modulating recognition at both sites, inducing site selectivity for glycocholic and glycochenodeoxycholic acid. Protein conformational changes following the introduction of a disulfide bridge are small and located around the inner binding site, whereas significant changes in backbone motions are observed for several residues distributed over the entire protein, both in the apo form and in the holo form. Site selectivity appears, therefore, to be dependent on protein mobility rather than being governed by steric factors. The detected properties further establish a parallelism with the behaviour of human ileal BABP, substantiating the proposal that BABPs have parallel functions in hepatocytes and enterocytes. PMID:19754879

  1. Pressurized water extraction of β-glucan enriched fractions with bile acids-binding capacities obtained from edible mushrooms.

    PubMed

    Palanisamy, Marimuthu; Aldars-García, Laila; Gil-Ramírez, Alicia; Ruiz-Rodríguez, Alejandro; Marín, Francisco R; Reglero, Guillermo; Soler-Rivas, Cristina

    2014-01-01

    A pressurized water extraction (PWE) method was developed in order to extract β-glucans with bile acids-binding capacities from cultivated mushrooms (Agaricus bisporus, Lentinula edodes, and Pleurotus ostreatus) to be used as supplements to design novel foods with hypocholesterolemic properties. Extraction yields were higher in individual than sequential extractions being the optimal extraction parameters: 200°C, 5 cycles of 5 min each at 10.3 MPa. The crude polysaccharide (PSC) fractions, isolated from the PWE extracts contained mainly β-glucans (including chitooligosaccharides deriving from chitin hydrolysis), α-glucans, and other PSCs (hetero-/proteo-glucans) depending on the extraction temperature and mushroom strain considered. The observed bile acids-binding capacities of some extracts were similar to a β-glucan enriched fraction obtained from cereals. PMID:24399760

  2. Clinical benefit using sperm hyaluronic acid binding technique in ICSI cycles: a systematic review and meta-analysis.

    PubMed

    Beck-Fruchter, Ronit; Shalev, Eliezer; Weiss, Amir

    2016-03-01

    The human oocyte is surrounded by hyaluronic acid, which acts as a natural selector of spermatozoa. Human sperm that express hyaluronic acid receptors and bind to hyaluronic acid have normal shape, minimal DNA fragmentation and low frequency of chromosomal aneuploidies. Use of hyaluronic acid binding assays in intracytoplasmic sperm injection (ICSI) cycles to improve clinical outcomes has been studied, although none of these studies had sufficient statistical power. In this systematic review and meta-analysis, electronic databases were searched up to June 2015 to identify studies of ICSI cycles in which spermatozoa able to bind hyaluronic acid was selected. The main outcomes were fertilization rate and clinical pregnancy rate. Secondary outcomes included cleavage rate, embryo quality, implantation rate, spontaneous abortion and live birth rate. Seven studies and 1437 cycles were included. Use of hyaluronic acid binding sperm selection technique yielded no improvement in fertilization and pregnancy rates. A meta-analysis of all available studies showed an improvement in embryo quality and implantation rate; an analysis of prospective studies only showed an improvement in embryo quality. Evidence does not support routine use of hyaluronic acid binding assays in all ICSI cycles. Identification of patients that might benefit from this technique needs further study. PMID:26776822

  3. Relationship between binding affinities to cellular retinoic acid-binding protein and biological potency of a new series of retinoids.

    PubMed

    Sani, B P; Dawson, M I; Hobbs, P D; Chan, R L; Schiff, L J

    1984-01-01

    Binding affinities of a new and unusual series of retinoic acid analogues to cellular retinoic acid-binding protein, a possible mediator of their biological function in the control of differentiation and tumorigenesis, and to serum albumin, their plasma transport protein, were determined. Also, biological activity of these retinoids in the reversal of keratinization in hamster tracheal organ cultures was assessed and compared with their binding affinities. Analogues that possessed high biological activity showed high binding efficiency to cellular retinoic acid-binding protein. Those that were biologically less active were poor binders to the binding protein. Three retinoids, 4657-57, 3920-59, and 4445-75, which showed 90 to 100% binding efficiency of that of retinoic acid for cellular retinoic acid-binding protein expressed high biological activity detectable in the range of 10(-10) M as against 10(-11) M for retinoic acid. The correlation noticed in these two activities not only enhances the confidence in the two assay procedures but also paves the way for design and development of potential chemopreventive agents. No apparent differences were observed in the binding affinities of the retinoids to binding proteins of a normal tissue or a tumor tissue. No correlation existed between the binding affinities of these retinoids to serum albumin and their biological activity. Structure-activity relationships of the retinoids in relation to their binding affinities and biological activities have been discussed. PMID:6317169

  4. Steam cooking significantly improves in vitro bile acid binding of collard greens, kale, mustard greens, broccoli, green bell pepper, and cabbage.

    PubMed

    Kahlon, Talwinder Singh; Chiu, Mei-Chen M; Chapman, Mary H

    2008-06-01

    Bile acid binding capacity has been related to the cholesterol-lowering potential of foods and food fractions. Lowered recirculation of bile acids results in utilization of cholesterol to synthesize bile acid and reduced fat absorption. Secondary bile acids have been associated with increased risk of cancer. Bile acid binding potential has been related to lowering the risk of heart disease and that of cancer. Previously, we have reported bile acid binding by several uncooked vegetables. However, most vegetables are consumed after cooking. How cooking would influence in vitro bile acid binding of various vegetables was investigated using a mixture of bile acids secreted in human bile under physiological conditions. Eight replicate incubations were conducted for each treatment simulating gastric and intestinal digestion, which included a substrate only, a bile acid mixture only, and 6 with substrate and bile acid mixture. Cholestyramine (a cholesterol-lowering, bile acid binding drug) was the positive control treatment and cellulose was the negative control. Relative to cholestyramine, in vitro bile acid binding on dry matter basis was for the collard greens, kale, and mustard greens, 13%; broccoli, 10%; Brussels sprouts and spinach, 8%; green bell pepper, 7%; and cabbage, 5%. These results point to the significantly different (P < or = .05) health-promoting potential of collard greens = kale = mustard greens > broccoli > Brussels sprouts = spinach = green bell pepper > cabbage as indicated by their bile acid binding on dry matter basis. Steam cooking significantly improved the in vitro bile acid binding of collard greens, kale, mustard greens, broccoli, green bell pepper, and cabbage compared with previously observed bile acid binding values for these vegetables raw (uncooked). Inclusion of steam-cooked collard greens, kale, mustard greens, broccoli, green bell pepper, and cabbage in our daily diet as health-promoting vegetables should be emphasized. These green

  5. Circular RNA oligonucleotides. Synthesis, nucleic acid binding properties, and a comparison with circular DNAs.

    PubMed Central

    Wang, S; Kool, E T

    1994-01-01

    We report the synthesis and nucleic acid binding properties of two cyclic RNA oligonucleotides designed to bind single-stranded nucleic acids by pyr.pur.pyr-type triple helix formation. The circular RNAs are 34 nucleotides in size and were cyclized using a template-directed nonenzymatic ligation. To ensure isomeric 3'-5' purity in the ligation reaction, one nucleotide at the ligation site is a 2'-deoxyribose. One circle (1) is complementary to the sequence 5'-A12, and the second (2) is complementary to 5'-AAGAAAGAAAAG. Results of thermal denaturation experiments and mixing studies show that both circles bind complementary single-stranded DNA or RNA substrates by triple helix formation, in which two domains in a pyrimidine-rich circle sandwich a central purine-rich substrate. The affinities of these circles with their purine complements are much higher than the affinities of either the linear precursors or simple Watson-Crick DNA complements. For example, circle 1 binds rA12 (pH 7.0, 10 mM MgCl2, 100 mM NaCl) with a Tm of 48 degrees C and a Kd (37 degrees C) of 4.1 x 10(-9) M, while the linear precursor of the circle binds with a Tm of 34 degrees C and a Kd of 1.2 x 10(-6) M. The complexes of circle 2 are pH-dependent, as expected for triple helical complexes involving C(+)G.C triads, and mixing plots for both circles reveal one-to-one stoichiometry of binding either to RNA or DNA substrates. Comparison of circular RNAs with previously synthesized circular DNA oligonucleotides of the same sequence reveals similar behavior in the binding of DNA, but strikingly different behavior in the binding of RNA. The cyclic DNAs show high DNA-binding selectivity, giving relatively weaker duplex-type binding with complementary RNAs. The relative order of thermodynamic stability for the four types of triplex studied here is found to be DDD >> RRR > RDR >> DRD. The results are discussed in the context of recent reports of strong triplex dependence on RNA versus DNA backbones

  6. Circular RNA oligonucleotides. Synthesis, nucleic acid binding properties, and a comparison with circular DNAs.

    PubMed

    Wang, S; Kool, E T

    1994-06-25

    We report the synthesis and nucleic acid binding properties of two cyclic RNA oligonucleotides designed to bind single-stranded nucleic acids by pyr.pur.pyr-type triple helix formation. The circular RNAs are 34 nucleotides in size and were cyclized using a template-directed nonenzymatic ligation. To ensure isomeric 3'-5' purity in the ligation reaction, one nucleotide at the ligation site is a 2'-deoxyribose. One circle (1) is complementary to the sequence 5'-A12, and the second (2) is complementary to 5'-AAGAAAGAAAAG. Results of thermal denaturation experiments and mixing studies show that both circles bind complementary single-stranded DNA or RNA substrates by triple helix formation, in which two domains in a pyrimidine-rich circle sandwich a central purine-rich substrate. The affinities of these circles with their purine complements are much higher than the affinities of either the linear precursors or simple Watson-Crick DNA complements. For example, circle 1 binds rA12 (pH 7.0, 10 mM MgCl2, 100 mM NaCl) with a Tm of 48 degrees C and a Kd (37 degrees C) of 4.1 x 10(-9) M, while the linear precursor of the circle binds with a Tm of 34 degrees C and a Kd of 1.2 x 10(-6) M. The complexes of circle 2 are pH-dependent, as expected for triple helical complexes involving C(+)G.C triads, and mixing plots for both circles reveal one-to-one stoichiometry of binding either to RNA or DNA substrates. Comparison of circular RNAs with previously synthesized circular DNA oligonucleotides of the same sequence reveals similar behavior in the binding of DNA, but strikingly different behavior in the binding of RNA. The cyclic DNAs show high DNA-binding selectivity, giving relatively weaker duplex-type binding with complementary RNAs. The relative order of thermodynamic stability for the four types of triplex studied here is found to be DDD > RRR > RDR > DRD. The results are discussed in the context of recent reports of strong triplex dependence on RNA versus DNA backbones. Triplex

  7. Impact of UV-B on drought- or cadmium-induced changes in the fatty acid composition of membrane lipid fractions in wheat.

    PubMed

    Gondor, Orsolya Kinga; Szalai, Gabriella; Kovács, Viktória; Janda, Tibor; Pál, Magda

    2014-10-01

    UV-B radiation may have either a positive or negative impact under the same conditions in wheat, depending on the type of secondary abiotic stressor: Cd or drought. Supplemental UV-B prevented the wilting and leaf rolling induced by PEG treatment. In contrast, combined UV-B and Cd treatment resulted in pronounced oxidative stress. The opposite effect of UV-B radiation in the case of drought or cadmium stress may be related to the alteration induced in the fatty acid composition. UV-B caused changes in the unsaturation of leaf phosphatidylglycerol fractions, and the accumulation of flavonoid in the leaves may prevent the stress induced by subsequent drought treatment. However it resulted in pronounced injury despite the increased flavonoid content in roots exposed to Cd. This was manifested in a drastic decrease in the unsaturation of the leaf monogalactosyldiacylglycerol and the root phosphatidylglycerol and digalactosyldiacylglycerol fractions. Data on the flavonoid content and fatty acid composition showed that oxidative stress was induced by drought in the leaves, by Cd in the roots, and interestingly, by UV-B radiation in both the leaves and roots. The additive effect of the combined stresses was also detected in the roots. The results presented here suggest a relationship between the capacity of the plant to remodel the fatty acid composition and its resistance to various stress factors. PMID:25062444

  8. Measurements of the acid-binding capacity of ingredients used in pig diets

    PubMed Central

    2005-01-01

    Some feed ingredients bind more acid in the stomach than others and for this reason may be best omitted from pig starter foods if gastric acidity is to be promoted. The objective of this study was to measure the acid-binding capacity (ABC) of ingredients commonly used in pig starter foods. Ingredients were categorised as follows: (i) milk products (n = 6), (ii) cereals (n = 10), (iii) root and pulp products (n = 5), (iv) vegetable proteins (n = 11), (v) meat and fish meal (n = 2), (vi) medication (n = 3), (vii) amino acids (n = 4), (viii) minerals (n = 16), (ix) acid salts (n = 4), (x) acids (n = 10). A 0.5 g sample of food was suspended in 50 ml distilled de-ionised water with continuous stirring. This suspension was titrated with 0.1 mol/L HCl or 0.1 mol/L NaOH so that approximately 10 additions of titrant was required to reach pH 3.0. The pH readings after each addition were recorded following equilibration for three minutes. ABC was calculated as the amount of acid in milliequivalents (meq) required to lower the pH of 1 kg food to (a) pH 4.0 (ABC-4) and (b) pH 3.0 (ABC-3). Categories of food had significantly different (P < 0.01) ABC values. Mean ABC-4 and ABC-3 values of the ten categories were: (i) 623 (s.d. 367.0) and 936 (s.d. 460.2), (ii) 142 (s.d. 79.2) and 324 (s.d. 146.4), (iii) 368 (s.d. 65.3) and 804 (s.d. 126.7), (iv) 381 (s.d. 186.1) and 746 (s.d. 227.0), (v) 749 (s.d. 211.6) and 1508 (s.d. 360.8), (vi) 120 (s.d. 95.6) and 261 (s.d. 163.2), (vii) 177 (s.d. 60.7) and 1078 (s.d. 359.0), (viii) 5064 (s.d. 5525.1) and 7051 (s.d. 5911.6), (ix) 5057 (s.d. 1336.6) and 8945 (s.d. 2654.1) and (x) -5883 (s.d. 4220.5) and -2591 (s.d. 2245.4) meq HCl per kg, respectively. Within category, ABC-3 and ABC- 4 values were highly correlated: R2 values of 0.80 and greater for food categories i, iv, v, vi, vii and viii. The correlation between predicted and observed ABC values of 34 mixed diets was 0.83 for ABC-4 and 0.71 for ABC-3. It was concluded that complete diets

  9. Fatty Aldehyde and Fatty Alcohol Metabolism: Review and Importance for Epidermal Structure and Function

    PubMed Central

    Rizzo, William B.

    2014-01-01

    Normal fatty aldehyde and alcohol metabolism is essential for epidermal differentiation and function. Long-chain aldehydes are produced by catabolism of several lipids including fatty alcohols, sphingolipids, ether glycerolipids, isoprenoid alcohols and certain aliphatic lipids that undergo α- or ω-oxidation. The fatty aldehyde generated by these pathways is chiefly metabolized to fatty acid by fatty aldehyde dehydrogenase (FALDH, alternately known as ALDH3A2), which also functions to oxidize fatty alcohols as a component of the fatty alcohol:NAD oxidoreductase (FAO) enzyme complex. Genetic deficiency of FALDH/FAO in patients with Sjögren-Larsson syndrome (SLS) results in accumulation of fatty aldehydes, fatty alcohols and related lipids (ether glycerolipids, wax esters) in cultured keratinocytes. These biochemical changes are associated with abnormalities in formation of lamellar bodies in the stratum granulosum and impaired delivery of their precursor membranes to the stratum corneum (SC). The defective extracellular SC membranes are responsible for a leaky epidermal water barrier and ichthyosis. Although lamellar bodies appear to be the pathogenic target for abnormal fatty aldehyde/alcohol metabolism in SLS, the precise biochemical mechanisms are yet to be elucidated. Nevertheless, studies in SLS highlight the critical importance of FALDH and normal fatty aldehyde/alcohol metabolism for epidermal function. PMID:24036493

  10. Identification of a nucleic acid-binding region within the largest subunit of Drosophila melanogaster RNA polymerase II.

    PubMed Central

    Kontermann, R. E.; Kobor, M.; Bautz, E. K.

    1993-01-01

    The largest and the second-largest subunit of the multisubunit eukaryotic RNA polymerases are involved in interaction with the DNA template and the nascent RNA chain. Using Southwestern DNA-binding techniques and nitrocellulose filter binding assays of bacterially expressed fusion proteins, we have identified a region of the largest, 215-kDa, subunit of Drosophila RNA polymerase II that has the potential to bind nucleic acids nonspecifically. This nucleic acid-binding region is located between amino acid residues 309-384 and is highly conserved within the largest subunits of eukaryotic and bacterial RNA polymerases. A homology to a region of the DNA-binding cleft of Escherichia coli DNA polymerase I involved in binding of the newly synthesized DNA duplex provides indirect evidence that the nucleic acid-binding region of the largest subunit participates in interaction with double-stranded nucleic acids during transcription. The nonspecific DNA-binding behavior of the region is similar to that observed for the native enzyme in nitrocellulose filter binding assays and that of the separated largest subunit in Southwestern assays. A high content of basic amino acid residues is consistent with the electrostatic nature of nonspecific DNA binding by RNA polymerases. PMID:8443600

  11. Bile acid-binding activity of young persimmon (Diospyros kaki) fruit and its hypolipidemic effect in mice.

    PubMed

    Matsumoto, Kenji; Yokoyama, Shin-ichiro; Gato, Nobuki

    2010-02-01

    The hypolipidemic effects and bile acid-binding properties of young persimmon (Diospyros kaki) fruit were examined. In an animal experiment, male C57BL/6.Cr mice (n = 5) were fed an AIN-76-modified high fat diet supplemented with 2% or 5% (w/w) dried young persimmon fruit (YP) for 10 weeks. The intake of YP significantly enhanced fecal bile acid excretion and lowered the concentration of hepatic lipids and plasma cholesterol. Analysis of gene expression in liver tissue showed that 2% or 5% YP up-regulated the expression of the sterol regulatory element-binding protein-2 gene. In the 5% group, there were increased expressions of the genes for cholesterol 7alpha-hydroxylase and the low-density lipoprotein receptor. Next, the bile acid-binding ability of YP was analysed in vitro using cholic acid (CA). In 100-2000 microM CA solutions, 1% (w/v) YP adsorbed approximately 60% of CA, while dried mature persimmon fruit adsorbed approximately 20% of CA. The positive control, cholestyramine, adsorbed approximately 80% of CA in the 100-2000 microM CA solutions. A crude tannin extract from YP, which contained 54.7% condensed tannins, adsorbed approximately 78% of CA in the 2000 microM CA solutions. These results suggest that the ability of YP to bind bile acid contributes to its hypolipidemic effect in mice. PMID:19585467

  12. Inhibition of de novo Palmitate Synthesis by Fatty Acid Synthase Induces Apoptosis in Tumor Cells by Remodeling Cell Membranes, Inhibiting Signaling Pathways, and Reprogramming Gene Expression

    PubMed Central

    Ventura, Richard; Mordec, Kasia; Waszczuk, Joanna; Wang, Zhaoti; Lai, Julie; Fridlib, Marina; Buckley, Douglas; Kemble, George; Heuer, Timothy S.

    2015-01-01

    Inhibition of de novo palmitate synthesis via fatty acid synthase (FASN) inhibition provides an unproven approach to cancer therapy with a strong biological rationale. FASN expression increases with tumor progression and associates with chemoresistance, tumor metastasis, and diminished patient survival in numerous tumor types. TVB-3166, an orally-available, reversible, potent, and selective FASN inhibitor induces apoptosis, inhibits anchorage-independent cell growth under lipid-rich conditions, and inhibits in-vivo xenograft tumor growth. Dose-dependent effects are observed between 20–200 nM TVB-3166, which agrees with the IC50 in biochemical FASN and cellular palmitate synthesis assays. Mechanistic studies show that FASN inhibition disrupts lipid raft architecture, inhibits biological pathways such as lipid biosynthesis, PI3K–AKT–mTOR and β-catenin signal transduction, and inhibits expression of oncogenic effectors such as c-Myc; effects that are tumor-cell specific. Our results demonstrate that FASN inhibition has anti-tumor activities in biologically diverse preclinical tumor models and provide mechanistic and pharmacologic evidence that FASN inhibition presents a promising therapeutic strategy for treating a variety of cancers, including those expressing mutant K-Ras, ErbB2, c-Met, and PTEN. The reported findings inform ongoing studies to link mechanisms of action with defined tumor types and advance the discovery of biomarkers supporting development of FASN inhibitors as cancer therapeutics. Research in context Fatty acid synthase (FASN) is a vital enzyme in tumor cell biology; the over-expression of FASN is associated with diminished patient prognosis and resistance to many cancer therapies. Our data demonstrate that selective and potent FASN inhibition with TVB-3166 leads to selective death of tumor cells, without significant effect on normal cells, and inhibits in vivo xenograft tumor growth at well-tolerated doses. Candidate biomarkers for

  13. Bile acid-binding ability of kaki-tannin from young fruits of persimmon (Diospyros kaki) in vitro and in vivo.

    PubMed

    Matsumoto, Kenji; Kadowaki, Akio; Ozaki, Natsumi; Takenaka, Makiko; Ono, Hiroshi; Yokoyama, Shin-ichiro; Gato, Nobuki

    2011-04-01

    The bile acid-binding ability of a highly polymerized tannin (kaki-tannin) extracted from dried-young fruits of persimmon (Diospyros kaki) was examined. The kaki-tannin was composed mainly of epicatechin, epigallocatechin, epicatechin-3-O-gallate and epigallocatechin-3-O-gallate. Bile acid-binding ability of kaki-tannin was examined against cholic acid, glycocholic acid, taurocholic acid and deoxycholic acid in vitro, and its effect on fecal bile acid excretion in mice was also examined. Although the bile acid-binding ability of kaki-tannin was weaker than that of cholestyramine, kaki-tannin adsorbed all the bile acids tested and significantly promoted fecal bile acid excretion in mice when supplied at 1% (w/w) in the diet. PMID:20922818

  14. Increased placental fatty acid transporter 6 and binding protein 3 expression and fetal liver lipid accumulation in a mouse model of obesity in pregnancy.

    PubMed

    Díaz, Paula; Harris, Jessica; Rosario, Fredrick J; Powell, Theresa L; Jansson, Thomas

    2015-12-15

    Obesity in pregnancy is associated with increased fetal growth and adiposity, which, in part, is determined by transplacental nutrient supply. Trophoblast uptake and intracellular trafficking of lipids are dependent on placental fatty acid transport proteins (FATP), translocase (FAT/CD36), and fatty acid binding proteins (FABP). We hypothesized that maternal obesity in mice leads to increased placental expression of FAT/CD36, FATPs, and FABPs, and lipid accumulation in the fetal liver. C57/BL6J female mice were fed either a control (C; n = 10) or an obesogenic (OB; n = 10) high-fat, high-sugar diet before mating and throughout pregnancy. At E18.5, placentas and fetal livers were collected. Trophoblast plasma membranes (TPM) were isolated from placental homogenates. Expression of FAT/CD36 and FATP (TPM) and FABP (homogenates) was determined by immunoblotting. Gene expression was assessed by RT-quantitative PCR. Sections of fetal livers were stained for Oil Red O, and lipid droplets were quantified. TPM protein expression of FAT/CD36, FATP 2, and FATP 4 was comparable between C and OB groups. Conversely, TPM FATP 6 expression was increased by 35% in OB compared with C placentas without changes in mRNA expression. FABPs 1, 3-5 and PPARγ were expressed in homogenates, and FABP 3 expression increased 27% in OB compared with C placentas; however, no changes were observed in mRNA expression. Lipid droplet accumulation was 10-fold higher in the livers of fetuses from OB compared with C group. We propose that increased lipid transport capacity in obese mice promotes transplacental fatty acid transport and contributes to excess lipid accumulation in the fetal liver. PMID:26491104

  15. Tsetse Salivary Gland Proteins 1 and 2 Are High Affinity Nucleic Acid Binding Proteins with Residual Nuclease Activity

    PubMed Central

    Caljon, Guy; Ridder, Karin De; Stijlemans, Benoît; Coosemans, Marc; Magez, Stefan; De Baetselier, Patrick; Van Den Abbeele, Jan

    2012-01-01

    Analysis of the tsetse fly salivary gland EST database revealed the presence of a highly enriched cluster of putative endonuclease genes, including tsal1 and tsal2. Tsal proteins are the major components of tsetse fly (G. morsitans morsitans) saliva where they are present as monomers as well as high molecular weight complexes with other saliva proteins. We demonstrate that the recombinant tsetse salivary gland proteins 1&2 (Tsal1&2) display DNA/RNA non-specific, high affinity nucleic acid binding with KD values in the low nanomolar range and a non-exclusive preference for duplex. These Tsal proteins exert only a residual nuclease activity with a preference for dsDNA in a broad pH range. Knockdown of Tsal expression by in vivo RNA interference in the tsetse fly revealed a partially impaired blood digestion phenotype as evidenced by higher gut nucleic acid, hematin and protein contents. PMID:23110062

  16. Toxicity of synthetic herbicides containing 2,4-D and MCPA moieties towards Pseudomonas putida mt-2 and its response at the level of membrane fatty acid composition.

    PubMed

    Piotrowska, Aleksandra; Syguda, Anna; Chrzanowski, Łukasz; Heipieper, Hermann J

    2016-02-01

    One of the attempts to create more effective herbicidal compounds includes the use of ionic liquids. Herbicidal ionic liquids have more effective biological activity, they are less volatile, more thermally stable, and exhibit superior efficiency in comparison to typically employed herbicides, allowing the reduction of the herbicide dose applied per hectare. However, studies on the environmental toxicity of this group of compounds are very rarely available. Environmental toxicity is an important factor, showing the concentration of compounds that has negative effects on soil bacteria including those responsible for biodegradation processes. Therefore, potential toxicity of four herbicidal ionic liquids (HILs) precursors containing 2,4-D and MCPA moieties was tested with the well investigated model organism for toxicity and adaptation, Pseudomonas putida mt-2. Results were compared to those obtained for commercial 2,4-D and MCPA herbicides. Next to growth inhibition, given as EC50, changes in the isomerisation of cis to trans unsaturated fatty acids were applied as proxy for cellular stress adaptation to toxic substances. The results revealed that all investigated precursors of HILs showed lower toxicity compared to commercialized synthetic herbicides 2,4-D and MCPA. The collected data on toxicity of HILs together with their physico-chemical properties might be useful for assessing the potential risk of the environmental pollution as well as guidelines for setting the legislation for their future use. PMID:26347932

  17. Phylogenomic reconstruction of archaeal fatty acid metabolism

    PubMed Central

    Dibrova, Daria V.; Galperin, Michael Y.; Mulkidjanian, Armen Y.

    2014-01-01

    While certain archaea appear to synthesize and/or metabolize fatty acids, the respective pathways still remain obscure. By analyzing the genomic distribution of the key lipid-related enzymes, we were able to identify the likely components of the archaeal pathway of fatty acid metabolism, namely, a combination of the enzymes of bacterial-type β-oxidation of fatty acids (acyl-CoA-dehydrogenase, enoyl-CoA hydratase, and 3-hydroxyacyl-CoA dehydrogenase) with paralogs of the archaeal acetyl-CoA C-acetyltransferase, an enzyme of the mevalonate biosynthesis pathway. These three β-oxidation enzymes working in the reverse direction could potentially catalyze biosynthesis of fatty acids, with paralogs of acetyl-CoA C-acetyltransferase performing addition of C2 fragments. The presence in archaea of the genes for energy-transducing membrane enzyme complexes, such as cytochrome bc complex, cytochrome c oxidase, and diverse rhodopsins, was found to correlate with the presence of the proposed system of fatty acid biosynthesis. We speculate that because these membrane complexes functionally depend on fatty acid chains, their genes could have been acquired via lateral gene transfer from bacteria only by those archaea that already possessed a system of fatty acid biosynthesis. The proposed pathway of archaeal fatty acid metabolism operates in extreme conditions and therefore might be of interest in the context of biofuel production and other industrial applications. PMID:24818264

  18. In vitro bile-acid binding and fermentation of high, medium, and low molecular weight beta-glucan.

    PubMed

    Kim, Hyun Jung; White, Pamela J

    2010-01-13

    The impact of beta-glucan molecular weight (MW) on in vitro bile-acid binding and in vitro fermentation with human fecal flora was evaluated. beta-Glucan extracted from oat line 'N979-5-4' was treated with lichenase (1,3-1,4-beta-D-glucanase) to yield high (6.87x10(5) g/mol), medium (3.71x10(5) g/mol), and low (1.56x10(5) g/mol) MW fractions. The low MW beta-glucan bound more bile acid than did the high MW beta-glucan (p<0.05). If the positive control, cholestyramine, was considered to bind bile acid at 100%, the relative bile-acid binding of the original oat flour and the extracted beta-glucan with high, medium, and low MW was 15, 27, 24, and 21%, respectively. Significant effects of high, medium, and low MW beta-glucans on total SCFA were observed compared to the blank without substrate (p<0.05). There were no differences in pH changes and total gas production among high, medium, and low MW beta-glucans, and lactulose. The low MW beta-glucan produced greater amounts of SCFA than the high MW after 24 h of fermentation. Among the major SCFA, more propionate was produced from all MW fractions of extracted beta-glucans than from lactulose. In vitro fermentation of extracted beta-glucan fractions with different MW lowered pH and produced SCFA, providing potential biological function. PMID:20020684

  19. Lysine-functionalized nanodiamonds: synthesis, physiochemical characterization, and nucleic acid binding studies

    PubMed Central

    Kaur, Randeep; Chitanda, Jackson M; Michel, Deborah; Maley, Jason; Borondics, Ferenc; Yang, Peng; Verrall, Ronald E; Badea, Ildiko

    2012-01-01

    Purpose: Detonation nanodiamonds (NDs) are carbon-based nanomaterials that, because of their size (4–5 nm), stable inert core, alterable surface chemistry, fluorescence, and biocompatibility, are emerging as bioimaging agents and promising tools for the delivery of biochemical molecules into cellular systems. However, diamond particles possess a strong propensity to aggregate in liquid formulation media, restricting their applicability in biomedical sciences. Here, the authors describe the covalent functionalization of NDs with lysine in an attempt to develop nanoparticles able to act as suitable nonviral vectors for transferring genetic materials across cellular membranes. Methods: NDs were oxidized and functionalized by binding lysine moieties attached to a three-carbon-length linker (1,3-diaminopropane) to their surfaces through amide bonds. Raman and Fourier transform infrared spectroscopy, zeta potential measurement, dynamic light scattering, atomic force microscopic imaging, and thermogravimetric analysis were used to characterize the lysine-functionalized NDs. Finally, the ability of the functionalized diamonds to bind plasmid DNA and small interfering RNA was investigated by gel electrophoresis assay and through size and zeta potential measurements. Results: NDs were successfully functionalized with the lysine linker, producing surface loading of 1.7 mmol g−1 of ND. These modified NDs formed highly stable aqueous dispersions with a zeta potential of 49 mV and particle size of approximately 20 nm. The functionalized NDs were found to be able to bind plasmid DNA and small interfering RNA by forming nanosized “diamoplexes”. Conclusion: The lysine-substituted ND particles generated in this study exhibit stable aqueous formulations and show potential for use as carriers for genetic materials. PMID:22904623

  20. Changes in membrane fatty acid composition through proton-induced fabF mutation enhancing 1-butanol tolerance in E. coli

    NASA Astrophysics Data System (ADS)

    Jeong, Haeyoung; Kim, Sun Hong; Han, Sang Soo; Kim, Myung Hee; Lee, Keun Chul

    2012-07-01

    While a rational approach based on genomic data has become the preferred method for microbial strain development, radiation-induced random mutagenesis is still a robust method for organisms such as plants whose genome or target gene information is unavailable. We previously reported on a combined approach that consists of proton irradiation and a long-term experimental evolution to enhance 1-butanol tolerance of the E. coli C strain so that it can be used as a basal strain for the production of 1-butanol, a potential biofuel along with ethanol. Genome sequencing of one randomly chosen clone (PKH5000) from the endpoint population revealed eleven mutations occurring in the coding regions, and we found that a mutation (F74C) in fabF gene encoding β-ketoacyl-ACP synthases II is associated with a twofold increase in the major unsaturated fatty acid, cis-vaccenic acid. The increase of cis-vaccenic acid by wild-type FabF, which is more active at low temperatures or in the presence of organic compounds, is considered to be a protective mechanism against cold stress. A structural analysis of the FabF protein suggests that the F74C mutation may affect the enzyme activity through a change in flexibility around the catalytic site. The expression of a plasmid that harbors mutant fabF gene in the fabF knockout strain enhanced growth in a medium containing butanol with a concomitant elevation of the cis-vaccenic acid level. Among the eight available Keio knockout strains for genes that have amino acid substitution in the PKH5000 strain, the fabF mutant showed the slowest growth in the presence of 0.7% butanol. We propose that fabF, as probably the gene most responsible for butanol tolerance in wild-type form, contributes further when converted into a F74C missense mutation, which is beneficial as it increases the level of cis-vaccenic acid.

  1. Functional analysis of a dietary recombinant fatty acid binding protein 10 (FABP10) on the Epinephelus coioides in response to acute low temperature challenge.

    PubMed

    Luo, Sheng-Wei; Cai, Luo; Liu, Yuan; Wang, Wei-Na

    2014-02-01

    The effect of Ec-FABP10 (Epinephelus coiodes-FABP10) on growth performance, enzyme activity, respiratory burst, MDA level, ATP content, immune-related gene expression of juvenile orange-spotted grouper (E. coioides). The commercial diet supplemented with FABP10 protein was feed to orange-spotted grouper for six weeks. No significant difference was observed in the specific growth rates, while the survival rate in the FABP10 additive group was significantly higher. After the feeding trial, the groupers were exposed to acute low temperature challenge. The decreased level of respiratory burst activity was observed in the FABP10 additive group after the exposure to the acute low temperature stress, while the blood cell count increased significantly at 15 °C and a significant increase of ATP content was observed at 10 °C. Higher enzymatic activities of CAT and SOD were observed at 20 °C and 15 °C, respectively. Meanwhile, the lower level of MDA was observed after the exposure to acute low temperature challenge by comparing with the controls. Further transcript expression analyses of FABP10, SOD2, GPX4, HSPA4 and LIPC in liver by quantitative real-time PCR demonstrated that the up-regulated transcript expression of FABP10, SOD2, HSPA4 and LIPC was observed in FABP10 additive group at 15 °C, while the transcript expression of GPX4 increased significantly at 20 °C. Western blotting analysis confirmed that FABP10 protein expression strongly increased at 15 ± 0.5 °C in FABP10 additive group. These results showed that FABP10 additive diet could moderate the metabolic and immune abilities mainly via ROS pathway in the orange-spotted grouper. PMID:24412164

  2. Common Variants of the Liver Fatty Acid Binding Protein Gene Influence the Risk of Type 2 Diabetes and Insulin Resistance in Spanish Population

    PubMed Central

    Mansego, Maria Luisa; Martínez, Fernando; Martínez-Larrad, Maria Teresa; Zabena, Carina; Rojo, Gemma; Morcillo, Sonsoles; Soriguer, Federico; Martín-Escudero, Juan Carlos; Serrano-Ríos, Manuel; Redon, Josep; Chaves, Felipe Javier

    2012-01-01

    Summary The main objective was to evaluate the association between SNPs and haplotypes of the FABP1-4 genes and type 2 diabetes, as well as its interaction with fat intake, in one general Spanish population. The association was replicated in a second population in which HOMA index was also evaluated. Methods 1217 unrelated individuals were selected from a population-based study [Hortega study: 605 women; mean age 54 y; 7.8% with type 2 diabetes]. The replication population included 805 subjects from Segovia, a neighboring region of Spain (446 females; mean age 52 y; 10.3% with type 2 diabetes). DM2 mellitus was defined in a similar way in both studies. Fifteen SNPs previously associated with metabolic traits or with potential influence in the gene expression within the FABP1-4 genes were genotyped with SNPlex and tested. Age, sex and BMI were used as covariates in the logistic regression model. Results One polymorphism (rs2197076) and two haplotypes of the FABP-1 showed a strong association with the risk of DM2 in the original population. This association was further confirmed in the second population as well as in the pooled sample. None of the other analyzed variants in FABP2, FABP3 and FABP4 genes were associated. There was not a formal interaction between rs2197076 and fat intake. A significant association between the rs2197076 and the haplotypes of the FABP1 and HOMA-IR was also present in the replication population. Conclusions The study supports the role of common variants of the FABP-1 gene in the development of type 2 diabetes in Caucasians. PMID:22396741

  3. Severity of soybean meal induced distal intestinal inflammation, enterocyte proliferation rate, and fatty acid binding protein (Fabp2) staining differ between strains of rainbow trout (Oncorhynchus mykiss)

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Complete replacement of fishmeal in feeds for carnivorous fishes often causes reduced growth and can negatively affect health. Salmonids fed diets containing full fat or defatted soybean meal develop dose dependent inflammation in the distal intestine (DI). Little is known about the sensitivity of d...

  4. Development of a therapeutic monoclonal antibody that targets secreted fatty acid-binding protein aP2 to treat type 2 diabetes.

    PubMed

    Burak, M Furkan; Inouye, Karen E; White, Ariel; Lee, Alexandra; Tuncman, Gurol; Calay, Ediz S; Sekiya, Motohiro; Tirosh, Amir; Eguchi, Kosei; Birrane, Gabriel; Lightwood, Daniel; Howells, Louise; Odede, Geofrey; Hailu, Hanna; West, Shauna; Garlish, Rachel; Neale, Helen; Doyle, Carl; Moore, Adrian; Hotamisligil, Gökhan S

    2015-12-23

    The lipid chaperone aP2/FABP4 has been implicated in the pathology of many immunometabolic diseases, including diabetes in humans, but aP2 has not yet been targeted for therapeutic applications. aP2 is not only an intracellular protein but also an active adipokine that contributes to hyperglycemia by promoting hepatic gluconeogenesis and interfering with peripheral insulin action. Serum aP2 levels are markedly elevated in mouse and human obesity and strongly correlate with metabolic complications. These observations raise the possibility of a new strategy to treat metabolic disease by targeting serum aP2 with a monoclonal antibody (mAb) to aP2. We evaluated mAbs to aP2 and identified one, CA33, that lowered fasting blood glucose, improved systemic glucose metabolism, increased systemic insulin sensitivity, and reduced fat mass and liver steatosis in obese mouse models. We examined the structure of the aP2-CA33 complex and resolved the target epitope by crystallographic studies in comparison to another mAb that lacked efficacy in vivo. In hyperinsulinemic-euglycemic clamp studies, we found that the antidiabetic effect of CA33 was predominantly linked to the regulation of hepatic glucose output and peripheral glucose utilization. The antibody had no effect in aP2-deficient mice, demonstrating its target specificity. We conclude that an aP2 mAb-mediated therapeutic constitutes a feasible approach for the treatment of diabetes. PMID:26702093

  5. Steam Cooking Significantly Improves In Vitro Bile Acid Binding of Collard Greens, Kale, Mustard Greens, Broccoli, Green Bell Pepper and Cabbage

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Bile acid binding capacity has been related to the cholesterol-lowering potential of foods and food fractions. Lowering recirculating bile acids results in utilization of cholesterol to synthesize bile acid and reduced fat absorption. Secondary bile acids have been associated with increasing the r...

  6. Controlled growth of filamentous fatty acid vesicles under flow.

    PubMed

    Hentrich, Christian; Szostak, Jack W

    2014-12-16

    The earliest forms of cellular life would have required a membrane compartment capable of growth and division. Fatty acid vesicles are an attractive model of protocell membranes, as they can grow into filamentous vesicles that readily divide while retaining their contents. In order to study vesicle growth, we have developed a method for immobilizing multilamellar fatty acid vesicles on modified glass surfaces and inducing filamentous membrane growth under flow. Filament formation strictly depended on the presence of freshly neutralized fatty acid micelles in the flow chamber. Using light microscopy, we observed a strong dependence of initial growth velocity on initial vesicle size, suggesting that new fatty acid molecules were incorporated into the membrane over the entire external surface of the vesicle. We examined the influences of flow rate, fatty acid concentration, and salt concentration on filamentous growth and observed drastic shape changes, including membrane pearling, of preexisting membrane tubules in response to osmotic stress. These results illustrate the versatility of flow studies for exploring the process of fatty acid vesicle growth following exposure to free fatty acids. PMID:25402759

  7. Controlled Growth of Filamentous Fatty Acid Vesicles under Flow

    PubMed Central

    2014-01-01

    The earliest forms of cellular life would have required a membrane compartment capable of growth and division. Fatty acid vesicles are an attractive model of protocell membranes, as they can grow into filamentous vesicles that readily divide while retaining their contents. In order to study vesicle growth, we have developed a method for immobilizing multilamellar fatty acid vesicles on modified glass surfaces and inducing filamentous membrane growth under flow. Filament formation strictly depended on the presence of freshly neutralized fatty acid micelles in the flow chamber. Using light microscopy, we observed a strong dependence of initial growth velocity on initial vesicle size, suggesting that new fatty acid molecules were incorporated into the membrane over the entire external surface of the vesicle. We examined the influences of flow rate, fatty acid concentration, and salt concentration on filamentous growth and observed drastic shape changes, including membrane pearling, of preexisting membrane tubules in response to osmotic stress. These results illustrate the versatility of flow studies for exploring the process of fatty acid vesicle growth following exposure to free fatty acids. PMID:25402759

  8. Identification of Escherichia coli F4ac-binding proteins in porcine milk fat globule membrane.

    PubMed

    Novakovic, Predrag; Huang, Yanyun Y; Lockerbie, Betty; Shahriar, Farshid; Kelly, John; Gordon, John R; Middleton, Dorothy M; Loewen, Matthew E; Kidney, Beverly A; Simko, Elemir

    2015-04-01

    F4ac-positive enterotoxigenic Escherichia coli (ETEC) must attach to the intestinal mucosa to cause diarrhea in piglets. Prevention of bacterial attachment to the intestinal mucosa is the most effective defense against ETEC-induced diarrhea. Porcine milk fat globule membranes (MFGM) were shown to be able to inhibit attachment of ETEC to the intestinal brush border; however, the specific components of porcine MFGM that inhibited attachment of ETEC to enterocytes were not identified. Accordingly, the purpose of this study was to identify F4ac-binding MFGM proteins by overlay Western blot and affinity chromatography. The proteome of porcine MFGM was characterized and the following F4ac-binding proteins were detected by overlay Western blot and affinity chromatography: lactadherin, butyrophilin, adipophilin, acyl-CoA synthetase 3, and fatty acid-binding protein 3. The biological function of these proteins was not investigated but it is possible that their interaction with F4ac fimbria interferes with bacterial attachment and colonization. PMID:25852227

  9. Identification of Escherichia coli F4ac-binding proteins in porcine milk fat globule membrane

    PubMed Central

    Novakovic, Predrag; Huang, Yanyun Y.; Lockerbie, Betty; Shahriar, Farshid; Kelly, John; Gordon, John R.; Middleton, Dorothy M.; Loewen, Matthew E.; Kidney, Beverly A.; Simko, Elemir

    2015-01-01

    F4ac-positive enterotoxigenic Escherichia coli (ETEC) must attach to the intestinal mucosa to cause diarrhea in piglets. Prevention of bacterial attachment to the intestinal mucosa is the most effective defense against ETEC-induced diarrhea. Porcine milk fat globule membranes (MFGM) were shown to be able to inhibit attachment of ETEC to the intestinal brush border; however, the specific components of porcine MFGM that inhibited attachment of ETEC to enterocytes were not identified. Accordingly, the purpose of this study was to identify F4ac-binding MFGM proteins by overlay Western blot and affinity chromatography. The proteome of porcine MFGM was characterized and the following F4ac-binding proteins were detected by overlay Western blot and affinity chromatography: lactadherin, butyrophilin, adipophilin, acyl-CoA synthetase 3, and fatty acid-binding protein 3. The biological function of these proteins was not investigated but it is possible that their interaction with F4ac fimbria interferes with bacterial attachment and colonization. PMID:25852227

  10. Hepatocellular influx of [14C]oleate reflects membrane transport rather than intracellular metabolism or binding.

    PubMed Central

    Stremmel, W; Berk, P D

    1986-01-01

    When [14C]oleate bound to bovine serum albumin was incubated at 37 degrees C for 7 min with isolated rat hepatocytes in the absence of glucose, the cumulative oleate uptake curve had two components: a rapid, linear segment over the first 30 sec, followed by a slower, curvilinear component. At 173 microM [14C]oleate/albumin (1:1, mol/mol), the initial component had a slope (Vo) of 118 +/- 18 pmol per min per 5 X 10(4) hepatocytes (mean +/- SD). During this initial 30 sec, virtually no oleate was oxidized, and less than 11% was esterified. By 5 min, 79% was esterified; oxidation never exceeded 4%. Addition of 2 mM glucose significantly increased oleate esterification and thereby available oleate binding sites on cytosolic fatty acid binding protein but had no influence on Vo. Pretreatment with trypsin reduced Vo by 49 +/- 15%. These data indicate that the initial component of the oleate uptake curve reflects predominantly influx, whereas the subsequent component reflects a balance between influx, efflux, and intracellular metabolism. Influx is independent of intracellular binding, oxidation, and esterification and may reflect a membrane-associated carrier-mediated process. PMID:3458166

  11. Chicoric acid binds to two sites and decreases the activity of the YopH bacterial virulence factor

    PubMed Central

    Kuban-Jankowska, Alicja; Sahu, Kamlesh K.; Gorska, Magdalena; Tuszynski, Jack A.; Wozniak, Michal

    2016-01-01

    Chicoric acid (CA) is a phenolic compound present in dietary supplements with a large spectrum of biological properties reported ranging from antioxidant, to antiviral, to immunostimulatory properties. Due to the fact that chicoric acid promotes phagocytic activity and was reported as an allosteric inhibitor of the PTP1B phosphatase, we examined the effect of CA on YopH phosphatase from pathogenic bacteria, which block phagocytic processes of a host cell. We performed computational studies of chicoric acid binding to YopH as well as validation experiments with recombinant enzymes. In addition, we performed similar studies for caffeic and chlorogenic acids to compare the results. Docking experiments demonstrated that, from the tested compounds, only CA binds to both catalytic and secondary binding sites of YopH. Our experimental results showed that CA reduces activity of recombinant YopH phosphatase from Yersinia enterocolitica and human CD45 phosphatase. The inhibition caused by CA was irreversible and did not induce oxidation of catalytic cysteine. We proposed that inactivation of YopH induced by CA is involved with allosteric inhibition by interacting with essential regions responsible for ligand binding. PMID:26735581

  12. Chicoric acid binds to two sites and decreases the activity of the YopH bacterial virulence factor.

    PubMed

    Kuban-Jankowska, Alicja; Sahu, Kamlesh K; Gorska, Magdalena; Tuszynski, Jack A; Wozniak, Michal

    2016-01-19

    Chicoric acid (CA) is a phenolic compound present in dietary supplements with a large spectrum of biological properties reported ranging from antioxidant, to antiviral, to immunostimulatory properties. Due to the fact that chicoric acid promotes phagocytic activity and was reported as an allosteric inhibitor of the PTP1B phosphatase, we examined the effect of CA on YopH phosphatase from pathogenic bacteria, which block phagocytic processes of a host cell. We performed computational studies of chicoric acid binding to YopH as well as validation experiments with recombinant enzymes. In addition, we performed similar studies for caffeic and chlorogenic acids to compare the results. Docking experiments demonstrated that, from the tested compounds, only CA binds to both catalytic and secondary binding sites of YopH. Our experimental results showed that CA reduces activity of recombinant YopH phosphatase from Yersinia enterocolitica and human CD45 phosphatase. The inhibition caused by CA was irreversible and did not induce oxidation of catalytic cysteine. We proposed that inactivation of YopH induced by CA is involved with allosteric inhibition by interacting with essential regions responsible for ligand binding. PMID:26735581

  13. Cellular retinol-binding protein and retinoic acid-binding protein in rat testes: effect of retinol depletion.

    PubMed

    Ong, D E; Tsai, C H; Chytil, F

    1976-02-01

    Testes of rats contain two cellular binding proteins of interest in vitamin A metabolism. One protein binds retinoic acid with high specificity; the other binds retinol with high specificity. When the cellular retinol-binding protein was partially purified from rat testes, it exhibited fluorescence excitation and emission spectra similar to that of all-trans-retinol in hexane. Exposure of this preparation to UV light destroyed this fluorescence but spectra identical to the original were obtained after addition of retinol. Hexane extracts of the binding protein had fluorescence spectra identical to all-trans-retinol, suggesting that this compound is bound to the protein in vivo. Extracts of testes from retinol depleted rats were submitted to gel filtration but failed to show a retinol-like fluorescence at the elution position of retinol binding protein. This fluorescence was observed in the preparations from pair fed control animals. However, after addition of all-trans-retinol to the extracts from the depleted rats, fluorescence at that elution position was observed. This indicates that in testes of retinol depleted rats the cellular retinol binding protein is present but without bound retinol, in contrast to the non-depleted rats where 30-43% of the binding protein had bound retinol. The amounts of cellular retinol binding protein and retinoic acid binding protein in testes, as determined by sucrose gradient centrifugation, were found to be similar for retinol depleted and pair fed control rats. PMID:942996

  14. Techno-functional properties and in vitro bile acid-binding capacities of tamarillo (Solanum betaceum Cav.) hydrocolloids.

    PubMed

    Gannasin, Sri Puvanesvari; Adzahan, Noranizan Mohd; Mustafa, Shuhaimi; Muhammad, Kharidah

    2016-04-01

    Hydrocolloids were extracted from seed mucilage and the pulp fractions from red tamarillo (Solanum betaceum Cav.) mesocarp, and characterisation of their techno-functional properties and in vitro bile acid-binding capacities was performed. The seed mucilage hydrocolloids that were extracted, using either 1% citric acid (THC) or water (THW), had a good foaming capacity (32-36%), whereas the pulp hydrocolloids that were extracted, using 72% ethanol (THE) or 20mM HEPES buffer (THH), had no foaming capacity. The pulp hydrocolloid, however, possessed high oil-holding and water-holding capacities in the range of 3.3-3.6 g oil/g dry sample and 25-27 g water/g dry sample, respectively. This enabled the pulp hydrocolloid to entrap more bile acids (35-38% at a hydrocolloid concentration of 2%) in its gelatinous network in comparison to commercial oat fibre and other hydrocolloids studied. The exceptional emulsifying properties (80-96%) of both hydrocolloids suggest their potential applications as food emulsifiers and bile acid binders. PMID:26593571

  15. Structural basis for the assembly and nucleic acid binding of the TREX-2 transcription-export complex

    PubMed Central

    Ellisdon, Andrew M.; Dimitrova, Lyudmila; Hurt, Ed; Stewart, Murray

    2012-01-01

    The conserved TREX-2 transcription-export complex integrates transcription and processing of many actively-transcribed nascent mRNAs with the recruitment of export factors at nuclear pores and also contributes to transcriptional memory and genomic stability. We report the crystal structure of the Sac3–Thp1–Sem1 segment of Saccharomyces cerevisiae TREX-2 that interfaces with the gene expression machinery. Sac3–Thp1–Sem1 forms a novel PCI-domain complex characterized by the juxtaposition of Sac3 and Thp1 winged helix domains, forming a platform that mediates nucleic acid binding. Structure-guided mutations underline the essential requirement of the Thp1–Sac3 interaction for mRNA binding and for the coupling of transcription and processing with mRNP assembly and export. These results provide insight into how newly synthesized transcripts are efficiently transferred from TREX-2 to the principal mRNA export factor and, identify how Sem1 stabilizes PCI domain-containing proteins and promotes complex assembly. PMID:22343721

  16. Importance of the proline-rich multimerization domain on the oligomerization and nucleic acid binding properties of HIV-1 Vif.

    PubMed

    Bernacchi, Serena; Mercenne, Gaëlle; Tournaire, Clémence; Marquet, Roland; Paillart, Jean-Christophe

    2011-03-01

    The HIV-1 viral infectivity factor (Vif) is required for productive infection of non-permissive cells, including most natural HIV-1 targets, where it counteracts the antiviral activities of the cellular cytosine deaminases APOBEC-3G (A3G) and A3F. Vif is a multimeric protein and the conserved proline-rich domain (161)PPLP(164) regulating Vif oligomerization is crucial for its function and viral infectivity. Here, we expressed and purified wild-type Vif and a mutant protein in which alanines were substituted for the proline residues of the (161)PPLP(164) domain. Using dynamic light scattering, circular dichroism and fluorescence spectroscopy, we established the impact of these mutations on Vif oligomerization, secondary structure content and nucleic acids binding properties. In vitro, wild-type Vif formed oligomers of five to nine proteins, while Vif AALA formed dimers and/or trimers. Up to 40% of the unbound wild-type Vif protein appeared to be unfolded, but binding to the HIV-1 TAR apical loop promoted formation of β-sheets. Interestingly, alanine substitutions did not significantly affect the secondary structure of Vif, but they diminished its binding affinity and specificity for nucleic acids. Dynamic light scattering showed that Vif oligomerization, and interaction with folding-promoting nucleic acids, favor formation of high molecular mass complexes. These properties could be important for Vif functions involving RNAs. PMID:21076154

  17. Importance of the proline-rich multimerization domain on the oligomerization and nucleic acid binding properties of HIV-1 Vif

    PubMed Central

    Bernacchi, Serena; Mercenne, Gaëlle; Tournaire, Clémence; Marquet, Roland; Paillart, Jean-Christophe

    2011-01-01

    The HIV-1 viral infectivity factor (Vif) is required for productive infection of non-permissive cells, including most natural HIV-1 targets, where it counteracts the antiviral activities of the cellular cytosine deaminases APOBEC-3G (A3G) and A3F. Vif is a multimeric protein and the conserved proline-rich domain 161PPLP164 regulating Vif oligomerization is crucial for its function and viral infectivity. Here, we expressed and purified wild-type Vif and a mutant protein in which alanines were substituted for the proline residues of the 161PPLP164 domain. Using dynamic light scattering, circular dichroism and fluorescence spectroscopy, we established the impact of these mutations on Vif oligomerization, secondary structure content and nucleic acids binding properties. In vitro, wild-type Vif formed oligomers of five to nine proteins, while Vif AALA formed dimers and/or trimers. Up to 40% of the unbound wild-type Vif protein appeared to be unfolded, but binding to the HIV-1 TAR apical loop promoted formation of β-sheets. Interestingly, alanine substitutions did not significantly affect the secondary structure of Vif, but they diminished its binding affinity and specificity for nucleic acids. Dynamic light scattering showed that Vif oligomerization, and interaction with folding-promoting nucleic acids, favor formation of high molecular mass complexes. These properties could be important for Vif functions involving RNAs. PMID:21076154

  18. Bioactive Fatty Acids

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Oxygenated fatty acids are useful as specialty chemicals, plasticizers, and biomedicals. Microbial enzymes convert fatty acids to mono-, di-, and trihydroxy fatty acid products. Among them, Bacillus megaterium ALA2 converted n-6 and n-3 PUFAs to many new oxygenated fatty acids. Linoleic acid was ...

  19. Plant fatty acid hydroxylases

    DOEpatents

    Somerville, Chris; Broun, Pierre; van de Loo, Frank

    2001-01-01

    This invention relates to plant fatty acyl hydroxylases. Methods to use conserved amino acid or nucleotide sequences to obtain plant fatty acyl hydroxylases are described. Also described is the use of cDNA clones encoding a plant hydroxylase to produce a family of hydroxylated fatty acids in transgenic plants. In addition, the use of genes encoding fatty acid hydroxylases or desaturases to alter the level of lipid fatty acid unsaturation in transgenic plants is described.

  20. Dynamics of cellular retinoic acid binding protein I on multiple time scales with implications for ligand binding.

    PubMed

    Krishnan, V V; Sukumar, M; Gierasch, L M; Cosman, M

    2000-08-01

    Cellular retinoic acid binding protein I (CRABPI) belongs to the family of intracellular lipid binding proteins (iLBPs), all of which bind a hydrophobic ligand within an internal cavity. The structures of several iLBPs reveal minimal structural differences between the apo (ligand-free) and holo (ligand-bound) forms, suggesting that dynamics must play an important role in the ligand recognition and binding processes. Here, a variety of nuclear magnetic resonance (NMR) spectroscopy methods were used to systematically study the dynamics of both apo and holo CRABPI at various time scales. Translational and rotational diffusion constant measurements were used to study the overall motions of the proteins. Both apo and holo forms of CRABPI tend to self-associate at high (1.2 mM) concentrations, while at low concentrations (0.2 mM), they are predominantly monomeric. Rapid amide exchange rate and laboratory frame relaxation rate measurements at two spectrometer field strengths (500 and 600 MHz) were used to probe the internal motions of the individual residues. Several residues in the apo form, notably within the ligand recognition region, exhibit millisecond time scale motions that are significantly arrested in the holo form. In contrast, no significant differences in the high-frequency motions were observed between the two forms. These results provide direct experimental evidence for dynamics-induced ligand recognition and binding at a specifically defined time scale. They also exemplify the importance of dynamics in providing a more comprehensive understanding of how a protein functions. PMID:10924105

  1. Siglec-15, a member of the sialic acid-binding lectin, is a novel regulator for osteoclast differentiation

    SciTech Connect

    Hiruma, Yoshiharu; Hirai, Takehiro; Tsuda, Eisuke

    2011-06-10

    Highlights: {yields} Siglec-15 was identified as a gene overexpressed in giant cell tumor. {yields} Siglec-15 mRNA expression increased in association with osteoclast differentiation. {yields} Polyclonal antibody to Siglec-15 inhibited osteoclast differentiation in vitro. -- Abstract: Osteoclasts are tartrate-resistant acid phosphatase (TRAP)-positive multinucleated cells derived from monocyte/macrophage-lineage precursors and are critically responsible for bone resorption. In giant cell tumor of bone (GCT), numerous TRAP-positive multinucleated giant cells emerge and severe osteolytic bone destruction occurs, implying that the emerged giant cells are biologically similar to osteoclasts. To identify novel genes involved in osteoclastogenesis, we searched genes whose expression pattern was significantly different in GCT from normal and other bone tumor tissues. By screening a human gene expression database, we identified sialic acid-binding immunoglobulin-like lectin 15 (Siglec-15) as one of the genes markedly overexpressed in GCT. The mRNA expression level of Siglec-15 increased in association with osteoclast differentiation in cultures of mouse primary unfractionated bone marrow cells (UBMC), RAW264.7 cells of the mouse macrophage cell line and human osteoclast precursors (OCP). Treatment with polyclonal antibody to mouse Siglec-15 markedly inhibited osteoclast differentiation in primary mouse bone marrow monocyte/macrophage (BMM) cells stimulated with receptor activator of nuclear factor {kappa}B ligand (RANKL) or tumor necrosis factor (TNF)-{alpha}. The antibody also inhibited osteoclast differentiation in cultures of mouse UBMC and RAW264.7 cells stimulated with active vitamin D{sub 3} and RANKL, respectively. Finally, treatment with polyclonal antibody to human Siglec-15 inhibited RANKL-induced TRAP-positive multinuclear cell formation in a human OCP culture. These results suggest that Siglec-15 plays an important role in osteoclast differentiation.

  2. A Repetitive DNA Element Regulates Expression of the Helicobacter pylori Sialic Acid Binding Adhesin by a Rheostat-like Mechanism

    PubMed Central

    Vallström, Anna; Olofsson, Annelie; Öhman, Carina; Rakhimova, Lena; Borén, Thomas; Engstrand, Lars; Brännström, Kristoffer; Arnqvist, Anna

    2014-01-01

    During persistent infection, optimal expression of bacterial factors is required to match the ever-changing host environment. The gastric pathogen Helicobacter pylori has a large set of simple sequence repeats (SSR), which constitute contingency loci. Through a slipped strand mispairing mechanism, the SSRs generate heterogeneous populations that facilitate adaptation. Here, we present a model that explains, in molecular terms, how an intergenically located T-tract, via slipped strand mispairing, operates with a rheostat-like function, to fine-tune activity of the promoter that drives expression of the sialic acid binding adhesin, SabA. Using T-tract variants, in an isogenic strain background, we show that the length of the T-tract generates multiphasic output from the sabA promoter. Consequently, this alters the H. pylori binding to sialyl-Lewis x receptors on gastric mucosa. Fragment length analysis of post-infection isolated clones shows that the T-tract length is a highly variable feature in H. pylori. This mirrors the host-pathogen interplay, where the bacterium generates a set of clones from which the best-fit phenotypes are selected in the host. In silico and functional in vitro analyzes revealed that the length of the T-tract affects the local DNA structure and thereby binding of the RNA polymerase, through shifting of the axial alignment between the core promoter and UP-like elements. We identified additional genes in H. pylori, with T- or A-tracts positioned similar to that of sabA, and show that variations in the tract length likewise acted as rheostats to modulate cognate promoter output. Thus, we propose that this generally applicable mechanism, mediated by promoter-proximal SSRs, provides an alternative mechanism for transcriptional regulation in bacteria, such as H. pylori, which possesses a limited repertoire of classical trans-acting regulatory factors. PMID:24991812

  3. Regulation of B Cell Functions by the Sialic Acid-Binding Receptors Siglec-G and CD22

    PubMed Central

    Jellusova, Julia; Nitschke, Lars

    2011-01-01

    B cell antigen receptor (BCR) engagement can lead to many different physiologic outcomes. To achieve an appropriate response, the BCR signal is interpreted in the context of other stimuli and several additional receptors on the B cell surface participate in the modulation of the signal. Two members of the Siglec (sialic acid-binding immunoglobulin-like lectin) family, CD22 and Siglec-G have been shown to inhibit the BCR signal. Recent findings indicate that the ability of these two receptors to bind sialic acids might be important to induce tolerance to self-antigens. Sialylated glycans are usually absent on microbes but abundant in higher vertebrates and might therefore provide an important tolerogenic signal. Since the expression of the specific ligands for Siglec-G and CD22 is tightly regulated and since Siglecs are not only able to bind their ligands in trans but also on the same cell surface this might provide additional mechanisms to control the BCR signal. Although both Siglec-G and CD22 are expressed on B cells and are able to inhibit BCR mediated signaling, they also show unique biological functions. While CD22 is the dominant regulator of calcium signaling on conventional B2 cells and also seems to play a role on marginal zone B cells, Siglec-G exerts its function mainly on B1 cells and influences their lifespan and antibody production. Both Siglec-G and CD22 have also recently been linked to toll-like receptor signaling and may provide a link in the regulation of the adaptive and innate immune response of B cells. PMID:22566885

  4. RBscore&NBench: a high-level web server for nucleic acid binding residues prediction with a large-scale benchmarking database.

    PubMed

    Miao, Zhichao; Westhof, Eric

    2016-07-01

    RBscore&NBench combines a web server, RBscore and a database, NBench. RBscore predicts RNA-/DNA-binding residues in proteins and visualizes the prediction scores and features on protein structures. The scoring scheme of RBscore directly links feature values to nucleic acid binding probabilities and illustrates the nucleic acid binding energy funnel on the protein surface. To avoid dataset, binding site definition and assessment metric biases, we compared RBscore with 18 web servers and 3 stand-alone programs on 41 datasets, which demonstrated the high and stable accuracy of RBscore. A comprehensive comparison led us to develop a benchmark database named NBench. The web server is available on: http://ahsoka.u-strasbg.fr/rbscorenbench/. PMID:27084939

  5. Non-extractable procyanidins and lignin are important factors in the bile acid binding and radical scavenging properties of cell wall material in some fruits.

    PubMed

    Hamauzu, Yasunori; Mizuno, Yukari

    2011-03-01

    The cell wall components and the food functions of alcohol-insoluble solids (AIS) of Chinese quince, quince, hawthorn, apple, pear and blueberry fruits were analyzed. Chinese quince contained characteristically high contents of cellulose, lignin, and non-extractable procyanidins (NEPCs). On the other hand, the quince AIS contained the highest proportion of NEPCs, the highest mean degree of polymerization (mDP), the strongest radical scavenging activity, and strong bile acid binding activity. In fruit AIS, the lignin and NEPC contents both showed positive correlations with the bile acid binding and radical scavenging activities. The value for mDP × NEPC content was a good index for the radical scavenging activity. The results suggest that highly polymerized NEPCs and lignin are important factors of cell wall components of fruits to having a high functionality, and Chinese quince and quince are interesting fruits from this view point. PMID:21243435

  6. Endogenous fatty acids in olfactory hairs influence pheromone binding protein structure and function in Lymantria dispar.

    PubMed

    Nardella, Jason; Terrado, Mailyn; Honson, Nicolette S; Plettner, Erika

    2015-08-01

    The gypsy moth utilizes a pheromone, (7R,8S)-2-methyl-7,8-epoxyoctadecane, for mate location. The pheromone is detected by sensory hairs (sensilla) on the antennae of adult males. Sensilla contain the dendrites of olfactory neurons bathed in lymph, which contains pheromone binding proteins (PBPs). We have extracted and identified free fatty acids from lymph of sensory hairs, and we demonstrate that these function as endogenous ligands for gypsy moth PBP1 and PBP2. Homology modeling of both PBPs, and docking of fatty acids reveal multiple binding sites: one internal, the others external. Pheromone binding assays suggest that these fatty acids increase PBP-pheromone binding affinity. We show that fatty acid binding causes an increase in α-helix content in the N-terminal domain, but not in the C-terminal peptide of both proteins. The C-terminal peptide was shown to form a α-helix in a hydrophobic, homogeneous environment, but not in the presence of fatty acid micelles. Through partition assays we show that the fatty acids prevent adsorption of the pheromone on hydrophobic surfaces and facilitate pheromone partition into an aqueous phase. We propose that lymph is an emulsion of fatty acids and PBP that influence each other and thereby control the partition equilibria of hydrophobic odorants. PMID:26032337

  7. Restriction of dietary protein does not promote hepatic lipogenesis in lean or fatty pigs.

    PubMed

    Madeira, Marta S; Pires, Virgínia M R; Alfaia, Cristina M; Lopes, Paula A; Martins, Susana V; Pinto, Rui M A; Prates, José A M

    2016-04-01

    The influence of genotype (lean v. fatty) and dietary protein level (normal v. reduced) on plasma metabolites, hepatic fatty acid composition and mRNA levels of lipid-sensitive factors is reported for the first time, using the pig as an experimental model. The experiment was conducted on forty entire male pigs (twenty lean pigs of Large White×Landrace×Pietrain cross-breed and twenty fatty pigs of Alentejana purebreed) from 60 to 93 kg of live weight. Each pig genotype was divided into two subgroups, which were fed the following diets: a