Federica Campana PhD defense


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Federica Campana PhD defense

  1. 1. Tutor: Dottoranda:Prof. Stefano Piotto Piotto Federica CampanaCo-Tutor:Prof. Pablo V. EscribáDepartment of Biology, University of the Balearic IslandsSpainUniversità degli Studi di SalernoDottorato di Ricerca in Scienza e Tecnologie per l’Industria Chimica, Farmaceutica e AlimentareXI CICLOMolecular dynamics investigations of drug-cellmembrane interactions
  2. 2. Structure and function of lipidmembranesMembrane fluidizers alter membranephysical stateMembrane physical state modulatesthe activity of embedded proteinsCHOL content influences the effect ofmembrane fluidizersEffect of fatty acids insidemembranesOverview
  3. 3. Membrane properties depend on:temperaturepressureelectrical fieldpHsalt concentrationpresence of proteinsprotein conformationThe physical state of a biologicalmembrane depends on allthermodynamic variables.Membrane physical stateIt is involved in regulating theactivity of all proteins that areembedded and, consequently, theexpression of genes involved instress responses.
  4. 4. Objectives
  5. 5. Escribá, P. V. (2006) Trends in Molecular Medicine. 12:34-43Membrane Lipid Therapy (MLT)
  6. 6. Gα monomer Gβγ dimerBiogenic aminesAmino acids and ionsLipidsPeptides and proteinsOthersGαGβGγGPCRs and G proteins
  7. 7. GαGβGγ
  8. 8. G protein lipid moietiesGeranylgeranyol (GG) Myristic alcohol (MOH) Palmitic alcohol (POH)Myristic acid (MA) Palmitic acid (PA)
  9. 9. -418 2517-3 -13GG MOH POHFreeenergyofbinding(kcal/mol)POPCPOPC-POPELipid moieties affinity for different membrane compositions
  10. 10. Effect of lipid moieties on membranesAn increase in the proportion of PE gradually decreases Gαmonomer binding to model membranes.Heterotrimeric Gαβγ subunits have a greater affinity for non-lamellar phases.
  11. 11. Effect of hydroxylamine derivatives inmodulating membrane physical state
  12. 12. Vigh, L., Maresca, B., Harwood, J. L. (1998) TIBS. 23:369-74
  13. 13. Preservation of the chemical architecture of a cell or of an organism under stressful conditions istermed homeostasis.One of the best known mechanisms protecting cells from various stresses is the heat-shockresponse, which results in the induction of the synthesis of heat-shock proteins (HSPs or stressproteins).Hydroxylamine derivatives, interacting with lipid bilayers, promote the formation of chaperonemolecules in eukaryotic cells and induce the expression of heat-shock genes.NNO NCl OHNNO NNH2 OHBimoclomol BGP-15 NG-094HSP co-inducers
  14. 14. BGP-15 affinity for different CHOL concentrationsThe permeation of BGP-15 is mildlyinfluenced by the composition.Docking of BGP-15 is enhanced by highcholesterol level.BGP-15 affects both the level and the sizedistribution of CHOL-rich membranemicrodomains.BGP-15 activation of HSP involves theRac1 signaling cascade.Membrane CHOL profoundly affects thetargeting of Rac1 to membranes.BGP-15 inhibit the rapid HSF1 acetylationobserved in the early phase of heatstress, thereby promoting a prolongedduration of HSF1 binding to HSE on hspgenes.
  15. 15. Ability of HSP co-inducers to modify the physical state ofmembranes46.63 46.1643.2345.94SM/CHOL SM/CHOL/BGP-15 SM/CHOL/NG-094 SM/CHOL/BMCThickness-974566-1018570-981763-1011496SM/CHOL SM/CHOL/BGP-15 SM/CHOL/NG-094 SM/CHOL/BMCTotal energy0.920.890.840.92SM/CHOL SM/CHOL/BGP-15 SM/CHOL/NG-094 SM/CHOL/BMCCHOL Alignment
  16. 16. Effect of HSP co-inducers on membrane spatial distribution
  17. 17. CHOL content in lipid rafts influencesthe effect of HSP co-inducers
  18. 18. Affinity for CHOL concentration in membranes
  19. 19. Transparent atoms = more staticOpaque atoms = more mobileMembrane fluidityPure membrane Doped membraneNG-094 +SM/CHOL 60:40BGP-15 +SM/CHOL 80:20
  20. 20. BGP-15 and MβCD work together to induce HSP70HSP70 without BGP-15HSP70 with BGP-15Effect of cholesterol removal in HEK293 lines (Crul et al, unpublished results)
  21. 21. Hydroxy arachidonic acid, a newpotential non steroidal anti-inflammatory molecule
  22. 22. The COX functions as a membrane-associated homodimer, catalyzing the committed step in theconversion of AA to prostaglandin H2 (PGH2), following AAs release from membrane phospholipds.The COX enzymeLopez, D. H., Fiol-de Roque, M. A., Noguera-Salva, M. A., Teres, S., Campana, F., Piotto, S., Castro, J. A., Mohaibes, R. J., Escribá P.V., Busquets. X. 2-Hydroxy Arachidonic Acid: A New Non-Steroidal Anti-Inflammatory Drug. British Journal of Pharmacology. Submitted.
  23. 23. COX-1COX-2Docking on COX isoforms
  24. 24. 8.29 7.94 8.52 10.25 11.09 10.93AA AArOH AAsOH AA AArOH AAsOHBindingenergy(kcal/mol)COX-1 COX-2Affinity for COX isoforms
  25. 25. AA AA-OHFukui Indices for Radical Attackatom Mulliken Hirshfeld atom Mulliken HirshfeldC ( 1) 0.076 0.073 C ( 1) 0.121 0.110C ( 2) -0.023 0.014 C ( 2) -0.027 0.015H ( 47) 0.000 0.000 H ( 47) -0.005 -0.002H ( 48) 0.002 0.001 H ( 48) 0.007 0.003H ( 49) 0.014 0.007 H ( 49) 0.011 0.005O ( 50) 0.087 0.085 O ( 50) 0.108 0.111O ( 51) 0.027 0.038 O ( 51) 0.056 0.065H ( 52) 0.028 0.018 H ( 52) 0.013 0.008H ( 53) 0.034 0.022 H ( 53) 0.033 0.020H ( 54) 0.032 0.023 H ( 54) 0.042 0.032H ( 55) 0.019 0.014The presence of αOH reduces theprobability of extraction of thehydrogen on C13 of almost 60%The Fukui function explains the inibitor capabilities of AAxOH
  26. 26. Prof. Stefano Piotto PiottoProf.ssa Simona ConcilioProf. Pio IannelliDott.ssa Lucia SessaLab. 12Acknowledgement