Dr. Sérgio S. Funari - Scientist at HASYLAB

*. Structure-Dynamics Relationship in Crystallizing Poly(ethylene terephthalate) as Revealed by Time-resolved X-ray and Dielectric Methods.  PDF
C. Alvarez, I. Šics,  A. Nogales,  Z. Denchev,  S. S. Funari, T.A. Ezquerra. Polymer, 45,  3953-3959  (2004).

The isothermal cold crystallization of poly(ethylene terephthalate) was investigated by simultaneous small and wide angle X-ray scattering (SAXS and WAXS) and dielectric spectroscopy (DS). By this experimental approach (SWD), simultaneously collected information was obtained about the specific changes occurring in both crystalline and amorphous phases during crystallization. The  main features which are directly derived from our experiments can be explained assuming the  formation of a  heterogeneous multiple lamellar population. arrangement. The rigid amorphous phase can be associated with the intra-lamellar stack amorphous phase. The  restriction of  the  amorphous phase mobility mainly occurs in the inter-lamellar stacks regions probably due to the formation of secondary lamellae.
 

*. Small Angle X-Rays Scattering Studies of Biomolecules.  PDF format
Sérgio S. Funari,  Acta Phys. Pol., 101, 647-658, 2002.

The building units of biological systems, the biomolecules, cannot easily be organized or even classified into defined categories. They can be as simple as water or complex as tintin, a muscle protein extremely large with several thousand atoms. To understand their function, one must know their characteristics, where they occur and what they do. One approach to reach such an ambitious task is to determine their structure, as single molecules or assembled into aggregates. Small angle X-ray scattering (SAXS) is the most important method for this purpose.
 We present studies carried out on several systems, and aiming at different questions about them. We start with lipids, the main components of the cell membranes. These membranes form the cell boundaries, the moiety required for the so-called membrane proteins, but also influence significantly several aspects of biological activity. More complex systems like a muscle fibre is also presented, showing that changes in the structure are related to the movement mechanism.
 It becomes easy to conclude that knowing the structures and the changes occurring in them is an important way to understand the function of biomolecules and therefore their role in the life cycle.
 

*. Detergent-phospholipid mixed micelles with a crystalline phospholipid core.  PDF format
Sérgio S. Funari, Brigitte Nuscher, Gert Rapp, and Klaus Beyer. Proc. Natl. Acad. Sci. USA,  98(16), 8938-8943, (2001).

An unusual micelle was discovered in mixtures of the nonionic detergent octaethyleneglycol-mono-n-dodecylether with disaturated phospholipids such as 1,2-dimyristoyl-sn-glycero-3-phosphocholine or 1,2-dipalmitoyl-sn-glycero-3-phosphocholine in water. These mixtures undergo a structural transition upon cooling through the chain-melting temperatures of the respective phospholipids, resulting in the formation of mixed micelles. Structural features of the micellar particles were studied here by synchrotron x-ray scattering. The translucent micellar solutions showed characteristic wide-angle reflections that were attributed to ordered hydrocarbon chains, whereas the absence of small-angle x-ray reflections indicated that there is no long-range order in these mixtures. The presence of ordered phospholipid acyl chains was confirmed by differential scanning calorimetry and isothermal titration calorimetry. The endothermic differential scanning calorimetry signals observed in the up-scan mode were tentatively ascribed to chain melting and mixing of the components. Isothermal titration of the mixed-micellar solutions into an excess of the detergent octaethyleneglycol-mono-n-dodecylether resulted in sudden uptake of the latent heat by the gel-state phospholipids.
The heat uptake per mol of phospholipid decreased with increasing detergent/phospholipid molar ratio. A simple geometric model is presented assuming that the dominating particle species in the mixtures is a discoidal phospholipid aggregate with ordered acyl chains, surrounded by a toroidal detergent hoop. The model implies that the fraction of ordered phospholipid chains decreases with increasing detergent/phospholipid molar ratio, in agreement with the calorimetric results and high-resolution NMR spectroscopy.
 

*. Structure of free Thermus flavus 5S rRNA at 1.3 nm resolution from synchrotron X-ray solution scattering
Sérgio S. Funari, Gert Rapp, Markus Perbrandt, Karsten Dierks, Marco Vallazza, Christian Betzel, Volker A. Erdmann and Dmitri I. Svergun -  J. Biol. Chem.,  275( 40), 31283-31288, (2000).

The shape of free Thermus flavus 5S rRNA in solution at 1.3 nm resolution is restored from synchrotron X-ray scattering data using an ab initio simulated annealing algorithm. The free 5S rRNA is a bent elongated molecule displaying a compact central region and two projecting arms, similar to those of the tRNA. The atomic models of the 5S rRNA domains A-D-E and B-C in the form of elongated helices can be well accommodated within the shape yielding a tentative model of the structure of the free 5S rRNA in solution. Its comparison with the recent protein-RNA map in the ribosome (Svergun, D.I. and Nierhaus, K.H. 2000) J. Biol. Chem. 275, 14432-14439) indicates that the 5S rRNA becomes essentially more compact upon complex ormation with specific ribosomal proteins. A conceivable conformational change involves rotation of the B-C domain towards the A-D-E domain. The model of free 5S rRNA displays no interactions between the domains E and C, but such interactions are possible in the bound molecule.
 

*. Nonaqueous Synthesis of Mesostructured Aluminophosphate / Surfactant Composites: Synthesis, Characterization, and In-situ SAXS Studies
Michael Tiemann, Michael Fröba, Gert Rapp, Sérgio S. Funari -  Chem. Mater  12, 1342-1348 (2000).

Nanostructured aluminophosphate / dodecyl phosphate composite materials were synthesized under aqueous and alcoholic conditions. The syntheses were monitored by temperature- and time-resolved in-situ small angle X-ray scattering (SAXS). In the aqueous synthesis a lamellar mesostructure is formed within the first few minutes of the reaction; this structure maintains a constant d spacing independent of the reaction time and temperature. The alcoholic synthesis at low temperature yields a mixture of a lamellar and an inverted hexagonal mesostructure. SAXS investigations show that these two phases evolve competitively. The lamellar structure is favored by higher temperatures and/or longer synthesis times; above ~ 70 °C it is formed exclusively. Mixtures of both phases can be isolated as solid materials, but thermal analysis shows that the inverted hexagonal product transforms into the lamellar phase at ~ 35 - 43 °C. The alcoholic synthesis is a highly cooperative reaction; the pure surfactant/alcohol systems are not lyotropic as long as the inorganic reactants are absent. In comparison, the surfactant/water system with the same surfactant concentrations as employed for the aqueous syntheses is lyotropic with a lamellar structure.
 

*. A Continuous Topological Change during Phase Transitions in Amphiphile/Water Systems
Sérgio S. Funari and Gert Rapp -  PNAS ,  96, 7756-7759 (1999).

Amphiphiles are molecules such as surfactants or lipids which have a polar head group (hydrophilic) attached to non-polar hydrophobic alkyl chains. Due to this characteristic they self-assemble in water and give rise to a wide range of phases with different structures and properties. Aqueous dispersions of amphiphiles are present in every aspect of day-to-day life, e.g. forming biological cell membranes, stabilizing emulsified food or being used as soap. Time-resolved X-ray diffraction has been used to study the C₁₆EO₆/water system that shows an intermediate phase whose structure depends on the thermal path between lamellar and hexagonal structures. Heating the hexagonal phase from room temperature leads to a lamellar phase via an Ia3d cubic structure. Cooling from the lamellar phase initially leads epitaxially to an intermediate R m before the hexagonal phase is reached. Both cubic and R m phases are formed by very similar rod units but the overall structures differ due to their spatial distribution and they both bridge morphologically the hexagonal and lamellar phases. The Ia3d does so on heating while the R m does on cooling. The structural path during the phase transitions is determined by topological similarities between the forming phase and the one it originates from. Although the estimated curvature energy for these two phases are similar, on cooling, kinetic and topology are initial factors determining the path for the phase transitions, while on heating energy is the dominant factor.
 

*. Induction of a Hexagonal Phase in Phospholipid-Surfactant Bilayers
Sérgio S. Funari -  Eur. Biophys. J. 27, 590-594 (1998).

The possibility of modifying the curvature of a lipid bilayer by mixing them with additives is shown and the evolution of geometrical parameters with composition is discussed. X-ray diffraction patterns of the POPC/C₁₂EO₂/²H₂O system were observed as a function of the relative humidity. The formation of an unexpected hexagonal phase evidences a peculiar behaviour of these mixtures. The cylinder radius of this phase is considerably smaller than previously observed in cubic phases. A discussion about the head groups' interactions is presented. Also, we have been able to show that the uptake of water by the L_b gel phase is higher as a single phase compared to the L_b+ H_II two phases region. The water content is important for the stabilization of the H_II phase and determination of its characteristic dimensions. However, it is argued that the interaction between the surfactant and lipid is the key factor for its formation and that the EO₂ head groups displace water from the inner parts of the polar region of the mesogenic units.

Keywords: gel phase, micelles, aggregation number, x-ray, hydration, geometrical parameters.
 

*. X-ray diffraction and NMR studies on mixtures of non-ionic surfactant (C₁₂EO₂) and phospholipids (POPC)
Sérgio S. Funari, Cláudio di Vitta and Gert Rapp -  Acta Phys. Pol.A91, 953-960 (1997).  PDF format

We have studied the temperature and concentration dependence of the POPC/C₁₂EO₂/water ternary system and identified and characterized the different phases occurring. The thermal behaviour is slightly dependent on the surfactant to lipid molar ratio R_s/l. At low surfactant content the behaviour is similar to the binary lipid/water system. Similarly at low lipid contents the system behaves like surfactant/water mixtures. However, at molar ratios R_s/l between 2 and 3 a new hexagonal phase forms which is peculiar to the mixture. ²H NMR measurements indicated the existence of a lamellar phase containing also curved surfaces additional to the flat ones. The surface area per surfactant molecule was calculated to 0.23 nm² which is in good agreement with the cross section of a paraffinic chain.

PACS no. 87.10.+e
 

*. X-ray Studies on the C₁₂EO₂/water System
Sérgio S. Funari and Gert Rapp -  J. Phys. Chem. B101, 732-739 (1997).

Binary mixtures of the poly(oxyethylene) surfactant C₁₂EO₂ and water have been investigated using optical microscopy and time-resolved X-ray diffraction during temperature scans. At concentrations in the range from 48 wt% to 70 wt% of surfactant a thermal sequence from lamellar L_a to cubic Ia3d to cubic Pn3m to L₂ was found upon heating. The geometrical parameters of the phases such as thickness of the hydrocarbon core of the lamellar phase and length and diameter of the rods forming the cubic structures were calculated for such conditions. In the lamellar L_a phase the surface area per molecule and the thickness of the hydrocarbon core showed little sensitivity to concentration. The thickness of the ethoxy groups was estimated to 1 nm. At high concentrations and temperatures near the cloud point, equilibrium conditions are difficult to be reached, leading to the formation of cubic phases with apparently the same structure but different thermal behaviour. One is insensitive to temperature changes while the other shrinks upon heating. The transition between the Ia3d and Pn3m cubic phases has the characteristics of a Bonnet transformation.
 

*. Cubic Topology in Surfactant and Lipid Mixtures
Sérgio S. Funari, Burkhard Mädler and Gert Rapp -  Eur. Biophys. J. 24, 293-299 (1996).

Ternary systems of palmitoyl-oleoyl-phosphatidylcholine (POPC) and the non-ionic surfactant C₁₂EO₂ (di-ethylene-oxide-mono-dodecyl-ether) in water have been studied with optical microscopy, NMR, DSC and X-rays from ambient temperatures to 45°C. Below 29°C the system is in the lamellar liquid crystalline state. Between 30 and 32°C it transforms into a cubic Ia3d structure which converts into the cubic Pn3m phase at 39°C. The transitions are fully reversible. An epitaxial relationship between all three phases was found, which is an elegant and convenient way to rearrange molecules from lamellar bilayers to a network of curved surfaces. The Ia3d (Q230) to Pn3m (Q224) transition occurs without measurable enthalpy change. This, together with the metric relation of 1.60 between the cubic lattice constants is strong evidence for a Bonnet transformation, where the structural changes occur without change in curvature. The potential significance of the cubic phases as intermediate structures for biological processes, e.g. transport across a bilayer or fusion of membranes, are discussed.

Keywords: Phase transition, NMR, DSC, X-ray diffraction, Bonnet Transformation.
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