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M U Palma

Publications and source records attributed to M U Palma.

18 recordsLinked to original sources
PubMed2004

Irreversible formation of intermediate BSA oligomers requires and induces conformational changes.

Understanding the relation between protein conformational changes and aggregation, and the physical mechanisms leading to such processes, is of primary importance, due to its direct relation to a vast class of severe pathologies. Growing evidence also suggests that oligomeric intermediates, which may occur early in the aggregation pathway, can be themselves pathogenic. The possible cytotoxicity of oligomers of non-disease-associated proteins adds generality to such suggestion and to the interest of studies of oligomer formation. Here we study the early stages of aggregation of Bovine Serum Albumin (BSA), a non pathogenic protein which has proved to be a useful model system. Dynamic light scattering and circular dichroism measurements in kinetic experiments following step-wise temperature rises, show that the "intermediate" form, which initiates large-scale aggregation, is the result of structural and conformational changes and concurrent formation of oligomers, of average size in the range of 100-200 A. Two distinct thresholds are observed. Beyond the first one oligomerization starts and causes partial irreversibility of conformational changes. Beyond the second threshold, additional secondary structural changes occurring in proteins being recruited progress on the same time scale of oligomerization. The concurrent behavior causes a mutual stabilization of oligomerization, and of structural and conformational changes, evidenced by a progressive increase of their irreversibility. This process interaction appears to be pivotal in producing irreversible oligomers.

Animals↗
PubMed2004

The role of pH on instability and aggregation of sickle hemoglobin solutions.

Understanding the physical basis of protein aggregation covers strong physical and biomedical interests. Sickle hemoglobin (HbS) is a point-mutant form of normal human adult hemoglobin (HbA). It is responsible for the first identified "molecular disease," as its propensity to aggregation is responsible for sickle cell disease. At moderately higher than physiological pH value, this propensity is inhibited: The rate of aggregate nucleation becomes exceedingly small and solubility after polymerization increases. These order-of-magnitude effects on polymer nucleation rates and concurrent relatively modest changes of solubility after polymerization are here shown to be related to both pH-induced changes of location and shape of the liquid-liquid demixing (LLD) region. This allows establishment of a self-consistent contact between the thermodynamics of the solution as such (i.e., the LLD region), the kinetics of fiber nucleation, the theory of percolation, and the thermodynamics of gelation. The observed pH-induced changes are largely attributable to strong perturbations of hydrophobic hydration configurations and related free energy by electric charges. Similar mechanisms of effective control of aggregate nucleation rates by means of agents such as cosolutes, pH, salts, and additives, shifting the LLD and associated regions of anomalous fluctuations, promise to be relevant to the whole field of protein aggregation pathologies.

Hemoglobin, Sickle↗
PubMed2003

Time scale of protein aggregation dictated by liquid-liquid demixing.

The growing impact of protein aggregation pathologies, together with the current high need for extensive information on protein structures are focusing much interest on the physics underlying the nucleation and growth of protein aggregates and crystals. Sickle Cell Hemoglobin (HbS), a point-mutant form of normal human Hemoglobin (HbA), is the first recognized and best-studied case of pathologically aggregating protein. Here we reanalyze kinetic data on nucleation of deoxy-HbS aggregates by referring them to the (concentration-dependent) temperature T(s) characterizing the occurrence of the phase transition of liquid-liquid demixing (LLD) of the solution. In this way, and by appropriate scaling of kinetic data at different concentrations, so as to normalize their spans, the apparently disparate sets of data are seen to fall on a master curve. Expressing the master curve vs. the parameter epsilon = (T - T(s)) / T(s), familiar from phase transition theory, allows eliciting the role of anomalously large concentration fluctuations associated with the LLD phase transition and also allows decoupling quantitatively the role of such fluctuations from that of microscopic, inter-protein interactions leading to nucleation. Referring to epsilon shows how in a narrow temperature span, that is at T - T(s), nucleation kinetics can undergo orders-of-magnitude changes, unexpected in terms of ordinary chemical kinetics. The same is true for similarly small changes of other parameters (pH, salts, precipitants), capable of altering T(s) and consequently epsilon. This offers the rationale for understanding how apparently minor changes of parameters can dramatically affect protein aggregation and related diseases.

Hemoglobin, Sickle↗
PubMed1999

Interacting processes in protein coagulation.

A strong interest is currently focused on protein self-association and deposit. This usually involves conformational changes of the entire protein or of a fragment. It can occur even at low concentrations and is responsible for pathologies such as systemic amyloidosis, Alzheimer's and Prion diseases, and other neurodegenerative pathologies. Readily available proteins, exhibiting at low concentration self-association properties related to conformational changes, offer very convenient model systems capable of providing insight into this class of problems. Here we report experiments on bovine serum albumin, showing that the process of conformational change of this protein towards an intermediate form required for coagulation occurs simultaneously and interacts with two more processes: mesoscopic demixing of the solution and protein cross-linking. This pathway of three interacting processes allows coagulation even at very low concentrations, and it has been recently observed also in the case of a nonpeptidic polymer. It could therefore be a fairly common feature in polymer coagulation/gelation. Proteins 1999;37:116-120.

Alzheimer Disease↗
PubMed1998

Interaction of explicit solvent with hydrophobic/philic/charged residues of a protein: residue character vs. conformational context.

Molecular dynamics simulations of model solutes in explicit molecular water have recently elicited novel aspects of the strong nonpair additivity of the potential of mean force (PMF) and related solvent-induced forces (SIFs) and hydration. Here we present the results of the same type of work on SIFs acting on bovine pancreatic trypsin inhibitor (BPTI) at single residue/sidechain resolution. In this system, nonpair additivity and the consequent dependence of SIFs on the protein conformational context are sufficiently strong to overturn SIFs on some individual residues, relative to expectations based on their individual characters. This finding calls for a revisitation and offers a richer and diversified understanding of the role of hydrophobic/philic/charged groups in establishing the exquisite specificity of biomolecular folding and functional conformation. Its relevance is appreciated by noting that the work of a typical SIF acting on one residue, when displaced across a distance of 1 A, is the equivalent of up to a few kcal/mol, which is the range of the stability/function free energy of a protein.

Amino Acids↗
PubMed1997

Collective properties of hydration: long range and specificity of hydrophobic interactions.

We report results of molecular dynamics (MD) simulations of composite model solutes in explicit molecular water solvent, eliciting novel aspects of the recently demonstrated, strong many-body character of hydration. Our solutes consist of identical apolar (hydrophobic) elements in fixed configurations. Results show that the many-body character of PMF is sufficiently strong to cause 1) a remarkable extension of the range of hydrophobic interactions between pairs of solute elements, up to distances large enough to rule out pairwise interactions of any type, and 2) a SIF that drives one of the hydrophobic solute elements toward the solvent rather than away from it. These findings complement recent data concerning SIFs on a protein at single-residue resolution and on model systems. They illustrate new important consequences of the collective character of hydration and of PMF and reveal new aspects of hydrophobic interactions and, in general, of SIFs. Their relevance to protein recognition, conformation, function, and folding and to the observed slight yet significant nonadditivity of functional effects of distant point mutations in proteins is discussed. These results point out the functional role of the configurational and dynamical states (and related statistical weights) corresponding to the complex configurational energy landscape of the two interacting systems: biomolecule + water.

Computer Simulation↗
PubMed1996

Self-assembly of biopolymeric structures below the threshold of random cross-link percolation.

Self-assembly of extended structures via cross-linking of individual biomolecules often occurs in solutions at concentrations well below the estimated threshold for random cross-link percolation. This requires solute-solute correlations. Here we study bovine serum albumin. Its unfolding causes the appearance of an instability region of the sol, not observed for native bovine serum albumin. As a consequence, spinodal demixing of the sol is observed. The thermodynamic phase transition corresponding to this demixing is the determinative symmetry-breaking step allowing the subsequent occurrence of (correlated) cross-linking and its progress up to the topological phase transition of gelation. The occurrence of this sequence is of marked interest to theories of spontaneous symmetry-breaking leading to morphogenesis, as well as to percolation theories. The present results extend the validity of conclusions drawn from our previous studies of other systems, by showing in one single case, system features that we have hitherto observed separately in different systems. Time-resolved experimental observations of the present type also bring kinetic and diffusional processes and solute-solvent interactions into the picture of cross-link percolation.

Animals↗
PubMed1993

Self-assembly of bioelastomeric structures from solutions: mean-field critical behavior and Flory-Huggins free energy of interactions.

Elastic and quasi-elastic light scattering studies were performed on aqueous solutions of poly(Val-Pro-Gly-Gly), a representative synthetic bioelastomer that differs from the previously studied poly(Val-Pro-Gly-Val-Gly) by the deletion of the hydrophobic Val in position four. When the spinodal line was approached from the region of thermodynamic stability, the intensity of light scattered by fluctuations, and the related lifetime and correlation length, were observed to diverge with mean-field critical exponents for both systems. Fitting of the experimental data allowed determining the spinodal and binodal (coexistence) lines that characterize the phase diagrams of the two systems, and it also allowed a quantitative sorting out of the enthalpic and entropic contributions to the Flory-Huggins interaction parameters. The contribution of valine is derived by comparison of the two cases. This can be viewed as sorting out the effect of a modulation of the solute. The same approach may allow sorting out the entropic and enthalpic effect of modulations of the solvent by cosolutes (or by cosolvents). This could be of particular interest in the case of small osmolytes, affording important adaptive roles in nature, at the cost of very limited changes in genetic information. Finally, the suggestion is further supported that statistical fluctuations of anomalous amplitude, such as those occurring in proximity of the spinodal line, have a role in promoting the process of self-assembly of extended supramolecular structures. On the practical side, the present approach appears useful in the design of novel synthetic model systems for bioelastomers.

Amino Acid Sequence↗
PubMed1991

Spinodal lines and Flory-Huggins free-energies for solutions of human hemoglobins HbS and HbA.

Gelation of deoxygenated solutions of sickle-cell human Hemoglobin (HbS) is of high theoretical interest and it has serious pathological consequences. For this reason HbS is probably the most studied protein capable of self-organization. This notwithstanding, the location in the T, c plane of the region of thermodynamic instability of solutions of deoxy-HbS (as bounded by the spinodal line and as distinct from the gelation region) has remained unknown, along with related values of Flory-Huggins enthalpies and entropies. In the present work this information is derived from experiments for the two cases of (deoxy) HbS and of human adult hemoglobin (HbA). Experiments also show critical exponents having mean-field values, which validates a Flory-Huggins approach. Altogether, the present work offers a quantitative understanding of the thermodynamic effects of the genetic HbA----HbS mutation and it opens the way to similar quantitative evaluations of contributions of pH, salts, cosolutes, and single peptides (even for nongelling hemoglobins), and of potential therapeutic strategies.

Biophysical Phenomena↗
PubMed1988

Nucleation and accretion of bioelastomeric fibers at biological temperatures and low concentrations.

Quasi-elastic light scattering (QELS) studies are reported, which address the early stages of aggregation of the polypentamer poly(VPGVG). This reflects the major primary structural feature of native elastin. The study is focused on the region of the phase diagram which in both its temperature and concentration range is closest to the state of affairs occurring in the course of bioelastogenesis by progressive synthesis of the precursor protein. Results here reported allow for the first time a self-consistent view of the physics of elastogenesis, and specify the role of the region of metastability and of that of instability of the phase diagram in the non-chaotic, orderly formation of elastomeric fibers.

Biopolymers↗
PubMed1983

Biomolecular-solvent stereodynamic coupling probed by deuteration.

Thermodynamic interpretation of experiments with isotopically perturbed solvent supports the view that solvent stereodynamics is directly relevant to thermodynamic stability of biomolecules. According with the current understanding of the structure of the aqueous solvent, in any stereodynamic configuration of the latter, connectivity pathways are identifiable for their topologic and order properties. Perturbing the solvent by isotopic substitution or, e.g., by addition of co-solvents, can therefore be viewed as reinforcing or otherwise perturbing these topologic structures. This microscopic model readily visualizes thermodynamic interpretation. In conclusion, the topologic stereodynamic structures of connectivity pathways in the solvent, as modified by interaction with solutes, acquire a specific thermodynamic and biological significance, and the problem of thermodynamic and functional stability of biomolecules is seen in its full pertinent phase space.

Deuterium↗
PubMed1980

Thermal stability of poly(A) and poly(U) complexes in H2O and D2O: isotopic effects on critical temperatures and transition widths.

This is a study of the effect of total and partial deuteration of solvent on critical temperatures and profiles of all four reactions occurring in poly(A) x n poly(U) (n = 1 or 2) aqueous systems. The study was done at observational times not longer than hydrogen exchange times at base pairs in helically ordered structures, and it was extended to a wide range of salt concentrations at neutral pH. The dependence of stability of polymer helical order on hydrogen mass does not appear to be merely attributable to the stronger intrahelical deuterium bonding. Substituting Deuterium for Hydrogen implies a probably predominant modulation of the entrophy term of polymer-solvent interactions. Effects of deuteration on the width of the 2(poly(A) x poly (U)) leads to poly(A) x 2poly(U)+poly(A) interconversion reaction were also observed. They bear on the role of polymer-solvent interaction on pattern recognition leading to formation of ordered structures. They also bear on the role of the same interaction on the "breathing" of ordered structures of this type.

Deuterium↗
PubMedDate not supplied

Self-assembly of a bioelastomeric structure: solution dynamics and the spinodal and coacervation lines.

The stability, metastability, and instability regions of aqueous solutions of a representative synthetic bioelastomeric polymer, poly (Val-Pro-Gly-Val-Gly), were determined by a combined use of elastic and quasi-elastic light scattering experiments. The approach followed here offers the attractive advantage of singling out the relevant contributions to the total scattering even in the presence of traces of noninteracting larger sized impurities. Conclusions so reached were checked by means of independent experiments. The present results provide descriptions of the very early events in the physics of bioelastogenesis in terms of general polymer science and phase transitions, and in terms of an unexpected possible functional role of density fluctuations.

Amino Acid Sequence↗
PubMedDate not supplied

Photon scattering as a probe of microviscosity and channel size in gels such as sickle haemoglobin.

The aggregation of sickle-cell haemoglobin (HbS) is one of the most physiologically important and widely studied macromolecular gelation processes. Both the thermodynamics and kinetics of the process are important in determining the pathological consequences of deoxygenation of the red cells (and both must be understood if a rational strategy is to be developed for pharmacological intervention). We describe here a new and versatile technique for the study of the structure and formation of the HbS aggregates, that should be widely applicable to gel systems generally. We use laser autocorrelation spectroscopy to observe the diffusion of monodisperse polystyrene latex spheres in the interstices of the gel.

Chemical Phenomena↗