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Browsing by Author "Katsaras, John"
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Item Docosahexaenoic acid regulates the formation of lipid rafts: A unified view from experiment and simulation(Elsevier, 2018-10) Wassall, Stephen R.; Leng, Xiaoling; Canner, Samuel W.; Pennington, Edward Ross; Kinnun, Jacob J.; Cavazos, Andres T.; Dadoo, Sahil; Johnson, Dylan; Heberle, Frederick A.; Katsaras, John; Shaikh, Saame Raza; Physics, School of ScienceDocosahexaenoic acid (DHA, 22:6) is an n-3 polyunsaturated fatty acid (n-3 PUFA) that influences immunological, metabolic, and neurological responses through complex mechanisms. One structural mechanism by which DHA exerts its biological effects is through its ability to modify the physical organization of plasma membrane signaling assemblies known as sphingomyelin/cholesterol (SM/chol)-enriched lipid rafts. Here we studied how DHA acyl chains esterified in the sn-2 position of phosphatidylcholine (PC) regulate the formation of raft and non-raft domains in mixtures with SM and chol on differing size scales. Coarse grained molecular dynamics simulations showed that 1-palmitoyl-2-docosahexaenoylphosphatylcholine (PDPC) enhances segregation into domains more than the monounsaturated control, 1-palmitoyl-2-oleoyl-phosphatidylcholine (POPC). Solid state 2H NMR and neutron scattering experiments provided direct experimental evidence that substituting PDPC for POPC increases the size of raft-like domains on the nanoscale. Confocal imaging of giant unilamellar vesicles with a non-raft fluorescent probe revealed that POPC had no influence on phase separation in the presence of SM/chol whereas PDPC drove strong domain segregation. Finally, monolayer compression studies suggest that PDPC increases lipid-lipid immiscibility in the presence of SM/chol compared to POPC. Collectively, the data across model systems provide compelling support for the emerging model that DHA acyl chains of PC lipids tune the size of lipid rafts, which has potential implications for signaling networks that rely on the compartmentalization of proteins within and outside of rafts.Item Interaction of α-tocopherol with a polyunsaturated lipid studied by MD simulations(Office of the Vice Chancellor for Research, 2013-04-05) Leng, Xiaoling; Williams, Justin A.; Marquardt, Drew; Kučerka, Norbert; Katsaras, John; Atkinson, Jeffrey; Harroun, Thad A.; Feller, Scott E.; Wassall, Stephen R.Polyunsaturated phospholipids are essential components of neural membranes and their effect on membrane architecture is proposed to be the molecular origin of a myriad of health benefits. A downside of polyunsaturated phospholipids is that they are highly susceptible to oxidation due to the presence of multiple double bonds. α-Tocopherol is the most biologically active component in a family of phenolic compounds that comprise vitamin E, which is the major lipid soluble antioxidant in cell membranes. To investigate whether α-tocopherol preferentially interacts with polyunsaturated phospholipids to optimize protection against oxidation, we performed MD simulations on 1-stearoyl-2-docosahexaenoylphosphatiylcholine (SDPC, 18:0-22:6PC) and 1-stearoyl-2-oleoylphosphatidylcholine (SOPC, 18:0-18:1PC) bilayers containing α-tocopherol. SDPC with a docosahexaenoyl sn-2 chain is polyunsaturated, while SOPC with an oleoyl sn-2 chain serves as a monounsaturated control. The simulations were run under constant pressure for 200 ns on a system that comprised 80 phospholipid molecules, 20 α-tocopherol molecules and 2165 water molecules. We discovered significant differences between the two systems. Analysis of the simulations indicates that the α-tocopherol has a strong interaction with the polyunsaturated fatty acid. The flip-flop of α-tocopherol across the bilayer is also much faster in SDPC than in SOPC. Solid state NMR, neutron scattering and complementary experiments are now underway to test the predictions from the MD simulations.Item Lipid bilayer thickness determines cholesterol's location in model membranes(RSC, 2016-12) Marquardt, Drew; Heberle, Frederick A.; Greathouse, Denise V.; Koeppe, Roger E., II.; Standaert, Robert F.; Van Oosten, Brad J.; Harroun, Thad A.; Kinnun, Jacob J.; Williams, Justin A.; Wassall, Stephen R.; Katsaras, John; Department of Physics, School of ScienceCholesterol is an essential biomolecule of animal cell membranes, and an important precursor for the biosynthesis of certain hormones and vitamins. It is also thought to play a key role in cell signaling processes associated with functional plasma membrane microdomains (domains enriched in cholesterol), commonly referred to as rafts. In all of these diverse biological phenomena, the transverse location of cholesterol in the membrane is almost certainly an important structural feature. Using a combination of neutron scattering and solid-state 2H NMR, we have determined the location and orientation of cholesterol in phosphatidylcholine (PC) model membranes having fatty acids of different lengths and degrees of unsaturation. The data establish that cholesterol reorients rapidly about the bilayer normal in all the membranes studied, but is tilted and forced to span the bilayer midplane in the very thin bilayers. The possibility that cholesterol lies flat in the middle of bilayers, including those made from PC lipids containing polyunsaturated fatty acids (PUFAs), is ruled out. These results support the notion that hydrophobic thickness is the primary determinant of cholesterol's location in membranes.Item The Molecular Structure of Sphingomyelin in Fluid Phase Bilayers Determined by the Joint Analysis of Small-Angle Neutron and X-ray Scattering Data(American Chemical Society, 2020-06-25) Doktorova, Milka; Kučerka, Norbert; Kinnun, Jacob J.; Pan, Jianjun; Marquardt, Drew; Scott, Haden L.; Venable, Richard M.; Pastor, Richard W.; Wassall, Stephen R.; Katsaras, John; Heberle, Frederick A.; Physics, School of ScienceWe have determined the fluid bilayer structure of palmitoyl sphingomyelin (PSM) and stearoyl sphingomyelin (SSM) by simultaneously analyzing small-angle neutron and X-ray scattering data. Using a newly developed scattering density profile (SDP) model for sphingomyelin lipids, we report structural parameters including the area per lipid, total bilayer thickness, and hydrocarbon thickness, in addition to lipid volumes determined by densitometry. Unconstrained all-atom simulations of PSM bilayers at 55 °C using the C36 CHARMM force field produced a lipid area of 56 Å2, a value that is 10% lower than the one determined experimentally by SDP analysis (61.9 Å2). Furthermore, scattering form factors calculated from the unconstrained simulations were in poor agreement with experimental form factors, even though segmental order parameter (SCD) profiles calculated from the simulations were in relatively good agreement with SCD profiles obtained from NMR experiments. Conversely, constrained area simulations at 61.9 Å2 resulted in good agreement between the simulation and experimental scattering form factors, but not with SCD profiles from NMR. We discuss possible reasons for the discrepancies between these two types of data that are frequently used as validation metrics for molecular dynamics force fields.Item α-Tocopherol is well designed to protect polyunsaturated fatty acids(Office of the Vice Chancellor for Research, 2014-04-11) Leng, Xiaoling; Williams, Justin A.; Marquardt, Drew; Kučerka, Norbert; Katsaras, John; Atkinson, Jeffrey; Harroun, Thad A.; Feller, Scott E.; Wassall, Stephen R.Polyunsaturated fatty acids (PUFA) are an influential constituent in cell membranes, but are extremely vulnerable to oxidation. The presumptive role for α-tocopherol (α-toc), the molecular form of vitamin E retained by the human body, is to protect PUFA-containing lipids from oxidation. To investigate whether α-toc preferentially interacts with PUFA in support of this function, we performed MD simulations on lipid bilayers composed of 1-stearoyl-2-docosahexaenoylphosphatidylcholine (SDPC, 18:0-22-6PC) and 1-stearoyl-2-oleoylphosphatidylcholine (SOPC, 18:0-18:1PC) in the presence of α-toc. SDPC with docosahexaenoic acid (DHA) for the sn-2 chain is polyunsaturated, while SOPC with oleic acid (OA) for the sn-2 chain serves as a monounsaturated control. The simulations were run at 37 °C under constant pressure for 200 ns on a system that comprised 80 phospholipid molecules, 20 α-toc molecules and 2165 water molecules. In qualitative agreement with our results from solid state 2H NMR and neutron scattering experiments, the simulations show that α-toc increases order inside the bilayer and that the chromanol headgroup sits near the surface in both SDPC and SOPC. Analyses of the density distribution of the lipid chains relative to α-toc show that the α-toc’s chromanol headgroup, the part of the molecule that protects against oxidation, would have more chance to interact with PUFA chains than saturated chains. A major prediction from our simulations is that α-toc undergoes flip-flop across the bilayer and that the rate is an order of magnitude greater in SDPC than SOPC. This is a remarkable finding that reveals a possible mechanism by which the chromanol group would not only wait at the membrane surface but would also patrol the membrane interior to meet lipid radicals and terminate the chain reaction by which lipid peroxidation proceeds.Item α-Tocopherol Is Well Designed to Protect Polyunsaturated Phospholipids: MD Simulations.(Elsevier, 2015-10-20) Leng, Xiaoling; Kinnun, Jacob J.; Marquardt, Drew; Ghefli, Mikel; Kučerka, Norbert; Katsaras, John; Atkinson, Jeffrey; Harroun, Thad A.; Feller, Scott E.; Wassall, Stephen R.; Department of Physics, School of ScienceThe presumptive function for alpha-tocopherol (αtoc) in membranes is to protect polyunsaturated lipids against oxidation. Although the chemistry of the process is well established, the role played by molecular structure that we address here with atomistic molecular-dynamics simulations remains controversial. The simulations were run in the constant particle NPT ensemble on hydrated lipid bilayers composed of SDPC (1-stearoyl-2-docosahexaenoylphosphatidylcholine, 18:0-22:6PC) and SOPC (1-stearoyl-2-oleoylphosphatidylcholine, 18:0-18:1PC) in the presence of 20 mol % αtoc at 37°C. SDPC with SA (stearic acid) for the sn-1 chain and DHA (docosahexaenoic acid) for the sn-2 chain is representative of polyunsaturated phospholipids, while SOPC with OA (oleic acid) substituted for the sn-2 chain serves as a monounsaturated control. Solid-state 2H nuclear magnetic resonance and neutron diffraction experiments provide validation. The simulations demonstrate that high disorder enhances the probability that DHA chains at the sn-2 position in SDPC rise up to the bilayer surface, whereby they encounter the chromanol group on αtoc molecules. This behavior is reflected in the van der Waals energy of interaction between αtoc and acyl chains, and illustrated by density maps of distribution for acyl chains around αtoc molecules that were constructed. An ability to more easily penetrate deep into the bilayer is another attribute conferred upon the chromanol group in αtoc by the high disorder possessed by DHA. By examining the trajectory of single molecules, we found that αtoc flip-flops across the SDPC bilayer on a submicrosecond timescale that is an order-of-magnitude greater than in SOPC. Our results reveal mechanisms by which the sacrificial hydroxyl group on the chromanol group can trap lipid peroxyl radicals within the interior and near the surface of a polyunsaturated membrane. At the same time, water-soluble reducing agents that regenerate αtoc can access the chromanol group when it locates at the surface.