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Nuno C Santos

Publications and source records attributed to Nuno C Santos.

14 recordsLinked to original sources
PubMed2006

Fluorescence spectroscopy evaluation of fibrinogen-beta-estradiol binding.

Fluorescence spectroscopy experiments were performed in order to study conformational changes induced by the binding of beta-estradiol to fibrinogen at different ligand concentrations. The association constant (Ka) obtained for the fibrinogen-beta-estradiol binding was 6.47x10(6)M(-1), indicating a high affinity interaction. Fluorescence quenching experiments showed that approximately 30% of the tryptophan residues in the protein quaternary structure are accessible to ionic quenchers. The extent of quenching in the absence and presence of beta-estradiol was maximum for cesium ions and minimum for iodide, suggesting the presence of negatively charged residues in the vicinity of the tryptophan residues. The quenching parameters obtained at different beta-estradiol concentrations show alterations that confirm a conformational change, possibly due to a discrete reorganization of tryptophan residues during fibrinogen-beta-estradiol binding. This binding may be responsible for the effects of beta-estradiol on the decrease of erythrocyte aggregation and on cardiovascular risk reduction.

Databases, Protein↗
PubMed2006

An extrasolar planetary system with three Neptune-mass planets.

Over the past two years, the search for low-mass extrasolar planets has led to the detection of seven so-called 'hot Neptunes' or 'super-Earths' around Sun-like stars. These planets have masses 5-20 times larger than the Earth and are mainly found on close-in orbits with periods of 2-15 days. Here we report a system of three Neptune-mass planets with periods of 8.67, 31.6 and 197 days, orbiting the nearby star HD 69830. This star was already known to show an infrared excess possibly caused by an asteroid belt within 1 au (the Sun-Earth distance). Simulations show that the system is in a dynamically stable configuration. Theoretical calculations favour a mainly rocky composition for both inner planets, while the outer planet probably has a significant gaseous envelope surrounding its rocky/icy core; the outer planet orbits within the habitable zone of this star.

Journal Article↗
PubMed2006

Fibrinogen-beta-estradiol binding studied by fluorescence spectroscopy: denaturation and pH effects.

Fibrinogen is a blood plasma protein that plays a crucial role in hemostasis. It is known that erythrocyte aggregation increases in the presence of fibrinogen, and that beta-estradiol decreases erythrocyte aggregation with a constant fibrinogen concentration. In this work, we have used intrinsic tryptophan fluorescence to obtain information on the conformational changes of fibrinogen upon the recently proposed interaction with beta-estradiol. To evaluate the effect on the conformational changes during fibrinogen-beta-estradiol binding, fluorescence experiments were performed using guanidine hydrochloride (0-6 M) as denaturant, at different pH values. The results obtained for pH 6.5 and 8.0 showed no effect during the binding. The main differences were observed between pH 4.2 and 7.4, in the absence and in the presence of two different denaturant concentrations (1 and 5 M). A red shift of the fluorescence emission from 344 to 354 nm is observed when denaturant concentration is above 3 M for all studied pH values. This phenomenon may be explained by the loss of compact structure of the protein in the presence of denaturant, with tryptophan residues exposure to the aqueous environment and alteration of fibrinogen-beta-estradiol binding. These results demonstrate that the binding sites of fibrinogen are strongly dependent on the conformational state of the protein.

Binding Sites↗
PubMed2006

Changes in blood cell membrane properties in HIV type-1-infected patients.

To evaluate the possible HIV-1 infection-induced changes in cell membrane properties and in calcium signaling, membrane fluidity, acetylcholinesterase (AChE, a glycosylphosphatidylinositol-anchored protein) activity, and intracellular calcium concentration ([Ca2(+)](int)) were evaluated in lymphocytes and erythrocytes of infected individuals, previous to their engagement in antiretroviral therapy. Membrane fluidity was assessed by fluorescence spectroscopy measurements, using the fluorescence probes 1,6-diphenyl-1,3,5-hexatriene (DPH) and 1-[4-(trimethylamino)-phenyl]-6-phenyl-1,3,5-hexatriene (TMA-DPH). AChE activity was determined by the colorimetric Ellman's method and [Ca2(+)](int) using the fluorescent fura-2 acetoxymethyl ester. When compared with the control group, lymphocytes of infected patients presented significantly decreased membrane fluidity, decreased AChE activity, and increased [Ca2(+)](int). Erythrocytes from HIV-infected patients presented decreased [Ca2(+)](int) when compared with the control group and decreased membrane fluidity near the lipid/water interface. Our data show that HIV-1 infection leads to biochemical and biophysical changes in the membrane itself and in membrane protein activity in lymphocytes (average of infected and noninfected subpopulations) and even in erythrocytes. The present observations are in agreement with a process of facilitated propagation of the infection to new cells, stimulation of virion production, and maintenance of a reservoir of erythrocyte-bound infectious virus.

Acetylcholinesterase↗
PubMed2005

Why are HIV-1 fusion inhibitors not effective against SARS-CoV? Biophysical evaluation of molecular interactions.

The envelope spike (S) glycoprotein of the severe acute respiratory syndrome associated coronavirus (SARS-CoV) mediates the entry of the virus into target cells. Recent studies point out to a cell entry mechanism of this virus similar to other enveloped viruses, such as HIV-1. As it happens with other viruses peptidic fusion inhibitors, SARS-CoV S protein HR2-derived peptides are potential therapeutic drugs against the virus. It is believed that HR2 peptides block the six-helix bundle formation, a key structure in the viral fusion, by interacting with the HR1 region. It is a matter of discussion if the HIV-1 gp41 HR2-derived peptide T20 (enfuvirtide) could be a possible SARS-CoV inhibitor given the similarities between the two viruses. We tested the possibility of interaction between both T20 (HIV-1 gp41 HR2-derived peptide) and T-1249 with S protein HR1- and HR2-derived peptides. Our biophysical data show a significant interaction between a SARS-CoV HR1-derived peptide and T20. However, the interaction is only moderate (K(B)=(1.1+/-0.3)x10(5) M(-1)). This finding shows that the reasoning behind the hypothesis that T20, already approved for clinical application in AIDS treatment, could inhibit the fusion of SARS-CoV with target cells is correct but the effect may not be strong enough for application.

Amino Acid Sequence↗
PubMed2005

Extrasolar planets: constraints for planet formation models.

Since 1995, more than 150 extrasolar planets have been discovered, most of them in orbits quite different from those of the giant planets in our own solar system. The number of discovered extrasolar planets demonstrates that planetary systems are common but also that they may possess a large variety of properties. As the number of detections grows, statistical studies of the properties of exoplanets and their host stars can be conducted to unravel some of the key physical and chemical processes leading to the formation of planetary systems.

Astronomical Phenomena↗
PubMed2005

Gramicidin D and dithiothreitol effects on erythrocyte exovesiculation.

The use of either diphenylhexatriene, trimethylamino-diphenylhexatriene, or heptadecyl-hydroxycoumarin (C17-HC) allows, simultaneously and with the same molecule, the induction of erythrocyte exovesiculation and labeling of the released vesicles with the fluorescent probe. This method was used to evaluate gramicidin D (a channel-forming peptide) and dithiothreitol (a reducing agent) effects on the human erythrocytes vesiculation process. The release of cholesterol and phospholipids in exovesicles at longer incubation times was only detectable in the presence of gramicidin or dithiothreitol. When C17-HC was used to induce the vesiculation, the presence of gramicidin or dithiothreitol lead to a drastic decrease on the [phospholipids]/[cholesterol] ratio. However, in the samples with dithiothreitol, this variation did not result in the expectable decrease of membrane fluidity. These effects can be related with the presence of lipid rafts, the transbilayer lipids reorientation induced by gramicidin or dithiothreitol, and the cholesterol-dependent gramicidin channels inactivation.

Cholesterol↗
PubMed2004

HIV fusion inhibitor peptide T-1249 is able to insert or adsorb to lipidic bilayers. Putative correlation with improved efficiency.

T-1249 is a HIV fusion inhibitor peptide under clinical trials. Its interaction with biological membrane models (large unilamellar vesicles) was studied using fluorescence spectroscopy. A gp41 peptide that includes one of the hydrophobic terminals of T-1249 was also studied. Both peptides partition extensively to liquid-crystalline POPC (1-palmitoyl-2-oleyl-sn-glycero-3-phosphocholine) (DeltaG = -7.0 kcal/mol and -8.7 kcal/mol, for T-1249 and terminal peptide, respectively) and are located at the interface of the membrane. T-1249 is essentially in a random coil conformation in this lipidic medium, although a small alpha-helix contribution is present. When other lipid compositions are used (DPPC, POPG + POPC, and POPC + cholesterol) (DPPC (1,2-dipalmitoyl-sn-glycero-3-phosphocholine) and POPG (1-palmitoyl-2-oleyl-sn-glycero-3-[phospho-rac-(1-glycerol)), partition decreases, the most severe effect being the presence of cholesterol. Partition experiments and fluorescence resonance energy transfer analysis show that T-1249 adsorbs to cholesterol-rich membranes. The improved clinical efficiency of T-1249 relative to enfuvirtide (T20) may be related to its bigger partition coefficient and ability to adsorb to rigid lipidic areas on the cell surface, where most receptors are inserted. Moreover, adsorption to the sterol-rich viral membrane helps to increase the local concentration of the inhibitor peptide at the fusion site.

Amino Acid Sequence↗
PubMed2004

An overview of the biophysical applications of atomic force microscopy.

The potentialities of the atomic force microscopy (AFM) make it a tool of undeniable value for the study of biologically relevant samples. AFM is progressively becoming a usual benchtop technique. In average, more than one paper is published every day on AFM biological applications. This figure overcomes materials science applications, showing that 17 years after its invention, AFM has completely crossed the limits of its traditional areas of application. Its potential to image the structure of biomolecules or bio-surfaces with molecular or even sub-molecular resolution, study samples under physiological conditions (which allows to follow in situ the real time dynamics of some biological events), measure local chemical, physical and mechanical properties of a sample and manipulate single molecules should be emphasized.

Animals↗
PubMed2004

Putative role of membranes in the HIV fusion inhibitor enfuvirtide mode of action at the molecular level.

Partition of the intrinsically fluorescent HIV fusion inhibitor enfuvirtide into lipidic membranes is relatively high (Delta G =6.6 kcal x mol(-1)) and modulated by cholesterol. A shallow position in the lipidic matrix makes it readily available for interaction with gp41. No conformational energetic barrier prevents enfuvirtide from being active in both aqueous solution and lipidic membranes. Lipidic membranes may play a key role in the enfuvirtide biochemical mode of action.

Enfuvirtide↗
PubMed2003

Quantifying molecular partition into model systems of biomembranes: an emphasis on optical spectroscopic methods.

Optical spectroscopies have been intensively used to determine partition coefficients by a plethora of methodologies. The present review is intended to give detailed and useful information for the determination of partition coefficients and addresses several relevant aspects, namely: (i) definition and calculation of the partition coefficient between aqueous and lipidic phases; (ii) partition coefficients vs. "binding" formalisms; (iii) advantages of spectroscopic methodologies over separation techniques; (iv) formalisms for various experimental approaches based on UV-Vis absorption or fluorescence parameters (fluorescence intensity, lifetime, anisotropy and quenching); (v) experimental hints, artifacts and model limitations; and (vi) a brief survey of nonoptical techniques.

Cell Membrane↗
PubMed2003

Multidisciplinary utilization of dimethyl sulfoxide: pharmacological, cellular, and molecular aspects.

DMSO is an amphipathic molecule with a highly polar domain and two apolar methyl groups, making it soluble in both aqueous and organic media. It is one of the most common solvents for the in vivo administration of several water-insoluble substances. Despite being frequently used as a solvent in biological studies and as a vehicle for drug therapy, the side-effects of DMSO (undesirable for these purposes) are apparent from its utilization in the laboratory (both in vivo and in vitro) and in clinical settings. DMSO is a hydrogen-bound disrupter, cell-differentiating agent, hydroxyl radical scavenger, intercellular electrical uncoupler, intracellular low-density lipoprotein-derived cholesterol mobilizing agent, cryoprotectant, solubilizing agent used in sample preparation for electron microscopy, antidote to the extravasation of vesicant anticancer agents, and topical analgesic. Additionally, it is used in the treatment of brain edema, amyloidosis, interstitial cystitis, and schizophrenia. Several systemic side-effects from the use of DMSO have been reported, namely nausea, vomiting, diarrhea, hemolysis, rashes, renal failure, hypertension, bradycardia, heart block, pulmonary edema, cardiac arrest, and bronchospasm. Looking at the multitude of effects of DMSO brought to light by these studies, it is easily understood how many researchers working with DMSO (or studying one of its specific effects) might not be fully aware of the experiences of other groups who are working with it but in a different context.

Amyloidosis↗
PubMed2003

Evaluation of lipopolysaccharide aggregation by light scattering spectroscopy.

Lipopolysaccharides (LPS) are cell wall components of Gram-negative bacteria. These molecules behave as bacterial endotoxins and their release into the bloodstream is a determinant of the development of a wide range of pathologies. These amphipathic molecules can self-aggregate into supramolecular structures with different shapes and sizes. The formation of these structures occurs when the LPS concentration is higher than the apparent critical micelle concentration (CMC(a)). Light scattering spectroscopy (both static and dynamic) was used to directly characterize the aggregation process of LPS from Escherichia coli serotype 026:B6. The results point to a CMC(a) value of 14 microg mL(-1) and the existence of premicelle LPS oligomers below this concentration. Both structures were characterized in terms of molecular weight (5.5 x 10(6) and 16 x 10(6) g mol(-1) below and above the CMC(a), respectively), interaction with the aqueous environment, gyration radius (56 and 105 nm), hydrodynamic radius, (60 and 95 nm) and geometry of the supramolecular structures (nearly spherical). Our data indicates that future in vitro experiments should be carried out both below and above the CMC(a). The search for drugs that interact with the aggregates, and thus change the CMC(a) and condition LPS interactions in the bloodstream, could be a new way to prevent certain bacterial-endotoxin-related pathologies.

Algorithms↗