Laurdan Spectrum Decomposition as a Tool for the Analysis of Surface Bilayer Structure and Polarity: a Study with DMPG, Peptides and Cholesterol

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Crystallographic and spectroscopic characterization of a molecular hinge: Conformational changes in bothropstoxin I, a dimeric Lys49 phospholipase A2 homologue

Bothropstoxin I(BthTX-I) from the venom of Bothrops jararacussu is a myotoxic phospholipase A2 (PLA2) homologue which, although catalytically inactive due to an Asp49-->Lys substitution, disrupts the integrity of lipid membranes by a Ca2+-independent mechanism, the crystal structures of two dimeric farms of BthLTX-I which diffract X-rays eo resolutions of 3.1 and 2.1 Angstrom have been determined, the monomers in both structures are related by an almost perfect twofold axis of rotation and the dimer interfaces are defined by contacts between the N-terminal alpha-helical regions and the tips of the beta-wings of partner monomers. Significant differences in the relative orientation of the monomers in the two crystal forms results in open and closed dimer conformations, Spectroscopic Investigations of BthTX-I in solution have correlated these conformational differences with changes in the intrinsic fluorescence emission of the single tryptophan residues located at the dimer interface, the possible relevance of this structural transition in the Ca2+-independent membrane damaging activity is discussed. (C) 1998 Wiley-Liss, Inc.

New Insight into the Mechanism of Action of Wasp Mastoparan Peptides: Lytic Activity and Clustering Observed with Giant Vesicles

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Consumo regular de suco de laranja vermelha em portadores de hepatite C crônica

Pós-graduação em Alimentos e Nutrição - FCFAR

Simulações por dinâmica molecular fine-e coarse-grained das interações intermoleculares entre peptídeos antimicrobianos da família Mastoparano e membranas modelo

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Nanotechnology-based drug delivery systems and herbal medicines: a review

Herbal medicines have been widely used around the world since ancient times. The advancement of phytochemical and phytopharmacological sciences has enabled elucidation of the composition and biological activities of several medicinal plant products. The effectiveness of many species of medicinal plants depends on the supply of active compounds. Most of the biologically active constituents of extracts, such as flavonoids, tannins, and terpenoids, are highly soluble in water, but have low absorption, because they are unable to cross the lipid membranes of the cells, have excessively high molecular size, or are poorly absorbed, resulting in loss of bioavailability and efficacy. Some extracts are not used clinically because of these obstacles. It has been widely proposed to combine herbal medicine with nanotechnology, because nano-structured systems might be able to potentiate the action of plant extracts, reducing the required dose and side effects, and improving activity. Nanosystems can deliver the active constituent at a sufficient concentration during the entire treatment period, directing it to the desired site of action. Conventional treatments do not meet these requirements. The purpose of this study is to review nanotechnology- based drug delivery systems and herbal medicines.

Functional and Topological Studies with Trp-Containing Analogs of the Peptide StII(1-30) Derived From the N-Terminus of the Pore Forming Toxin Sticholysin II: Contribution to Understand its Orientation in Membrane

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Recent advances in nanoparticle carriers for coordination complexes

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Monitoring the positioning of short polycationic peptides in model lipid bilayers by combining hydrogen/deuterium exchange and electrospray ionization mass spectrometry

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Initiating structural studies of Lys49-PLA2 homologues complexed with an anionic detergent, a fatty acid and a natural lipid

Lys49-Phospholipase A(2) (Lys49-PLA(2) - EC 3.1.1.4) homologues damage membranes by a Ca2+-independent mechanism which does not involve catalytic activity. Both MjTX-II from Bothrops moojeni and BthTX-I from Bothrops jararacussu are dimeric in solution and in the crystalline states, and a model for the Ca2+-independent membrane damaging mechanism has been suggested in which flexibility at the dimer interface region pert-nits quaternary structural transitions between open and closed membrane bound dimer conformations which results in the perturbation of membrane phospholipids and disruption of the bilayer structure [1]. With the aim of gaining insights into the structural determinants involved in protein/lipid association, we report here the crystallization and preliminary X-ray analysis of the (i) MjTX-II/SDS complex at a resolution of 2.78Angstrom, (ii) MjTX-II/STE complex at a resolution of 1.8 Angstrom and (W) BthTX-I/DMPC complex at 2.72Angstrom. These complexes were crystallized by the hanging drop vapour-diffusion technique in (i) HEPES buffer (pH 7.5) 1.8M ammonium sulfate with 2% (w/v) polyethyleneglycol 400, in (ii) 0.6-0.8 M sodium citrate as the precipitant (pH 6.0-6.5) and in (iii) sodium citrate buffer (pH 5.8) and PEG 4000 and 20% isopropanol, respectively. Single crystals of these complexes have been obtained and X-ray diffraction data have been collected at room temperature using a R-AXIS IV imaging plate system and graphite monochromated Cu Kalpha X-ray radiation generated by a Rigaku RU300 rotating anode generator for (i) and (W) and using using a Synchrotron Radiation Source (Laboratorio Nacional de Luz Sincrotron, LNLS, Campinas, Brazil) for (ii).

Tertiary structural changes of the alpha-hemolysin from Staphylococcus aureus on association with liposome membranes

The interaction of alpha-hemolysin (also called alpha-toxin) from Staphylococcus aureus with mixed egg-yolk phosphatidylcholine/cholesterol liposomes has been investigated using the intrinsic tryptophan fluorescence emission (ITFE) signal. The ITFE intensity of alpha-hemolysin, which was obtained using a novel purification protocol, showed a triphasic increase on incubation with liposomes at low protein/lipid ratios. The first, rapid phase results in an increase in ITFE of 10%, which reflects rapid conformation changes in the alpha-hemolysin on association with the liposome membrane, the second phase of the ITFE increase is associated with a red shift from 334 to 339 nm in the maximum emission wavelength, suggesting the transition to a partially unfolded intermediate in the oligomerization process. The third phase of the ITFE intensity change demonstrates a temporal correlation with the appearance of SDS-stable oligomers. The results demonstrate the feasibility of identification of intermediate protein conformations in complex membrane-associated processes by manipulation of the liposomal membrane composition. (C) 1998 Academic Press.

CHOLESTEROL MODULATION OF LIPID PROTEIN INTERACTIONS IN LIVER MICROSOMAL MEMBRANE - A SPIN LABEL STUDY

ESR spectra of spin probes were used to monitor lipid-protein interactions in native and cholesterol-enriched microsomal membranes. In both systems composite spectra were obtained, one characteristic of bulk bilayer organization and another due to a motionally restricted population, which was ascribed to lipids in a protein microenvironment. Computer spectral subtractions revealed that cholesterol modulates the order/mobility of both populations in opposite ways, i.e., while the lipid bilayer region gives rise to more anisotropic spectra upon cholesterol enrichment, the spectra of the motionally restricted population become indicative of increased mobility and/or decreased order. These events were evidenced by measurement of both effective order parameters and correlation times. The percentages of the motionally restricted component were invariant in native and cholesterol-enriched microsomes. Variable temperature studies also indicated a lack of variation of the percentages of both spectral components, suggesting that the motionally restricted one was not due to protein aggregation. The results correlate well with the effect of cholesterol enrichment on membrane-bound enzyme kinetics and on the behavior of fluorescent probes [Castuma & Brenner (1986) Biochemistry 25, 4733-4738]. Several hypothesis are put forward to explain the molecular mechanism of the cholesterol-induced spectral changes.

Changes in lipid composition in hippocampus early and late after status epilepticus induced by kainic acid in wistar rats

Oxidative damage to biological membranes has been reported as a cause of alterations in many different diseases. We had previously reported lipid peroxidation in the kainic acid model of temporal epilepsy. In this study we evaluated earlier and later modifications in the lipid composition after status epileticus induced by kainic acid. Lipid composition was determined by thin-layer chromatography, in the cortex and hippocampus 12-14 h, 7-8, 75-80, or 140-150 days after the end of status epileticus. In the hippocampus there was a significant change in the lipid protein ratio after status epileticus and this was accompanied by an alteration in lipid composition in all tested times. These results suggested that lipid peroxidation induced by kainic acid could be accompanied by chronic changes in the lipid composition that could be related to the development of seizures.

Improved embryonic cryosurvival observed after in vitro supplementation with conjugated linoleic acid is related to changes in the membrane lipid profile

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)