Rezultate obinute in cadrul proiectului CEEX 168/2006 VIASAN:

Rezultate obinute in cadrul proiectului CEEX 168/2006 VIASAN:

S-au pus la punct metodele necesare desfasurarii studiilor in etapele viitoare ale proiectului de catre fiecare colaborator: protocoale de realizare a membranelor model (sferice si plane), protocoale de caracterizare electrofiziologica a membranelor model, parametrii de modelare a structurii moleculare a lipopolizaharidelor, colectie de tulpini bacteriene Gram-negative.

S-a caracterizat fluiditatea membranara a bistratului lipidic realizat cu diverse compozitii lipidice in prezenta si absenta ionilor de calciu (2mM) si s-a constatat rigidizarea usoara a membranei in prezenta ionilor d ecalciu;

S-au masurat parametrii electrici ai bistratului lipidic cu structura asemanatoare celei a membranei externa bacteriene;

S-au realizat parametrizarile necesare descrierii structurii atomice tridimensionale a moleculei de LPS pentru programele AMBE Rsi CHARMM;

S-a analizat influenta cationilor asupra transportului prin diverse molecule formatoare de canale membranare (alameticina, amfotericina, OmpF);

S-a studiat influenta mediilor de crestere asupra eficientei unor antibiotice beta-lactamice pe culturi bacteriene de E.coli;

S-a analizat procesul de insertie al porinei OmpF in bistratul lipidic folosind tehnici de fluorescenta;

in cadrul modelului teoretic s-a evidentiat dependenta mobilitatii atomilor din segmentele hidrofobe ale lipidelor de adancimea in bistrat si de distanta fata de molecula proteica din bistrat;

s-au parametrizat doua molecule de antibiotic (rifamicina si ceftazidima);

s-au pozitionat moleculele de antibiotic in porina inserata in bistrat si s-a analizat interactia antibiotic porina;

s-au selectat linii bacteriene de E coli rezistente sau sensibile la antibiotice betalactamice si s-au extras fragmente de membrana externa de la aceste linii celulare;

s-au obtinut proteolipozomi prin fuzionarea fragmentelor de membrana cu lipozomi unilamelari mici;

folosind proteolipozomii mici s-au realizat proteolipozomi gigant;

s-a probat insertia fragmentelor de membrana in proteolipozomi prin tehnica FRET;

s-a evidentiat activitatea de canal ionic a porinelor din framentele de membrana incorporate din proteolipozomi;

s-a demonstrat ca ampicilina nu influenteaza curentul transmembranar si capacitatea unui bistrat lipidic;

s-au comparat predictiile teoretice ale modelului teoretic privind mobilitatea atomilor in bistrat cu rezultatele experimentale privind fluiditatea membranara

S-au determinat valorile conductantei canalului OmpF in prezenta si absenta antibioticului;

S-a demonstrat independenta interactiei antibiotic porine (in prezenta anticorpilor anti-LPS) de fluiditatea lipidelor din bistratul membranei externe bacteriene;

S-au calculat valorile constantei de legare a antibioticului la porina;

s-a demonstrat influenta factorilor imunitari asupra aderentei bacteriilor la un subtrat celular sau inert in prezenta unor concentratii de antibiotic subinhibitorii;

s-au calculat parametrii interactiei anticorpi molecule de LPS;

s-au predictionat trei structuri m moleculare derivate din ceftazidima care consideram ca au eficienta antibacteriana sporita;

s-au propus unele imbunatatiri pentru protocoalele de testare a antibioticelor in faza preclinica;

Articole publicate sau acceptate pentru publicare in reviste cotate ISI

Roxana Chiriac, Tudor Luchian,

pH modulation of transport properties of alamethicin oligomers inserted in zwitterionic-based artificial lipid membranes, Biophys Chem. 2007 Nov;130(3):139-47.

Alina Asandei, Loredana Mereuta, Tudor Luchian, Influence of membrane potential upon reversible protonation of acidic residues from the OmpF eyelet, Biophysical Chemistry, 2008, in press

Balotescu C., Dracea O., Iordache C., Bucur M., Necula G., Ungureanu C., Cristea D., Lixandru M.S., Lazar V. Influence of different cultivation paramaters on the expression of genotypic and phenotypic resistance of some E. coli strains (accepted for publication in Roum. Biotech.Lett.).

Articole publicate sau acceptate la publicare in reviste cotate CNCSIS B+

M. Bacalum, M. Radu

Insertion of proteins in the lipid bilayer of liposomes revealed by FRET

Romanian Journal of Biophysics, Volume 17, Issue 2, Pages 129-138, (2007)

Moraru Alexandra; Svab Istvana Mihailescu Dan Florin

CHARMM force field parameterization of bacterial lipopolysaccharides

Rev. Roum Chimie (in press)

Comunicari la manifestari internationale

52nd Annual Meeting of the Biophysical Society and 16th IUPAB International Biophysics, 2-6.02, 2008, in Long Beach, CA. CHARMM Force FieldParameterization of Lipid A,

In cadrul workshopului paralel Workshop II: "Investigation of pore forming proteins in model membranes", The Annual International Conference of RSBMB, BUCHAREST, 29 31 May 2008.

Dracea O, Iordache C, Bucur M, Necula G, Ungureanu C, Balotescu C, Cristea D, Lixandru MS, Rau I, Lazr V, "Investigation of The Influence of Different Physico-Chemical Parameters on E. Coli Susceptibility to Antibiotics", In cadrul workshopului paralel Workshop II: "Investigation of pore forming proteins in model membranes", The Annual International Conference of RSBMB, BUCHAREST, 29 31 May 2008.

Marin A, Macri BM, Radu M, Winterhalter M, "The Effect of Ceftazidime on the Electrophysiological Properties of Ompf ", in cadrul workshopului paralel Workshop II: "Investigation of pore forming proteins in model membranes", The Annual International Conference of RSBMB, BUCHAREST, 29 31 May 2008.

Moraru A, vab I, Mihilescu DF, "CHARMm Force Field Parameterization of Bacterial Lipopolysaccharides", in cadrul workshopului paralel Workshop II: "Investigation of pore forming proteins in model membranes", The Annual International Conference of RSBMB, BUCHAREST, 29 31 May 2008.

Dracea O., Iordache C., Bucur M., Necula G., Ungureanu C., Balotescu C., Cristea D., Lixandru M.S., Lazar V., 2007, Investigation of the influence of different physicochemical parameters on Escherichia coli susceptibility to beta-lactams antibiotics. Abstract book of the 4th Congress of the European Society for Emerging Infections, Lisbon, Portugal 30 Sept.-3 Oct. 2007, P. 117.

Comunicari manifestari nationale

Florentina Petcu, Mihaela Bacalum, M. Radu

Adsorption and insertion of proteins in the lipid bilayer of liposomes revealed by FRET

The IX National Conference on Biophysics, Bucharest, Romania, May 11-14, 2007

Beatrice Macri, Mihai Radu

Giant liposomes - eukaryotic and bacterial model membranes

The IX National Conference on Biophysics, Bucharest, Romania, May 11-14, 2007

Roxana Chiriac, Alina Asandei, Loredana Mereuta, Tudor Luchian, pH modulation of ion transport through alamethicin channels formed in phosphatidylcholine artificial membranes, Conference of the Romanian Society of Pure and Applied Biophysics (RSPAB), IXth edition, with international participation, 2007, Bucharest, Romania

Tudor Luchian, Complementary interactions between the membrane dipole potential and protein pores inserted in artificial lipid bilayers), Conference of the Romanian Society of Pure and Applied Biophysics (RSPAB), IXth edition, with international participation, 2007, Bucharest, Romania

Roxana Chiriac and Tudor Luchian , Rafts-induced modulation of transport and kinetic activity of alamethicin in artificial lipid membranes, International conference on fundamental and applied research in physics, Iasi, October, 2007

Loredana Mereuta and Tudor Luchian, Membrane dipole potential-induced modulation of current fluctuations through the OmpF porin, International conference on fundamental and applied research in physics, Iasi, October, 2007

George Necula (Neurobiologie, master an II), indrumator: Prof. Dr. D. Mihailescu, Caracterizarea insertiei OmpF in membranele lipidice, Sesiunea de comunicari stiintifice a studentilor si cadrelor didactice din Fac. de Biologie, 24.05.2008

Mihaela Bacalum, Mihai Radu, A fluorescent study of the interactions between antibiotics and the Outer Membrane Porin F inserted in model membranes, Conferinta Nationala de Fizica, Magurele, 12-13 septembrie 2008

Beatrice Macri, Dan Mihailescu, Carmen Balotescu, Tudor, Luchian, Andrei Petrescu, Mihai Radu, Studiul mecanismelor de rezistenta la antibiotice prin impermeabilitate la bacteriile Gram-negative pe membrane naturale si reconstituite, Simpozion VIASAN 2008, Sinaia, 28-30 septembrie 2008

Alexandra Moraru, Istvan Svab, Dan Florin Mihailescu, Modelarea moleculara a lipopolizaharidelor bacteriene, prezentata la A XXVI-a Sesiune Anuala a Societatii Nationale de Biologie Celulara, 12 14 Iunie, 2008, Cluj-Napoca