Proteins with mechanical functionality: a physico-chemical study on the viscoelasticity of gliadin, a storage protein from wheat gluten / Proteínas com funcionalidade mecânica: um estudo físico-químico sobre a viscoelasticidade da gliadina, uma proteína de reserva do glúten do trigo

AUTOR(ES)
DATA DE PUBLICAÇÃO

2004

RESUMO

The thesis consisted in a rheological study of the dynamics of gels and dispersions of gliadin from wheat gluten in pure dimethylsulfoxide, dimethylformamide or formamide and in their mixtures with water. The effect of temperature and that of the concentration of the component species (protein and solvent constituents) on flow behavior and deformation were evaluated. Steady-state, transient and dynamical (oscillatory) tests were performed. Protein characterization in the various media was performed through dynamic and static light scattering and small angle X ray scattering techniques. The application of techniques allowed to describe the distribution of protein populations in the gels and dispersions, in their free and associated forms, as well as to estimate their size and mobility, through the determination of the hydrodynamical radii and diffusion coefficients. The amino acid compositions of gliadin, both in the crude protein and in an isolated fraction in ethanol, were determined by HPLC technique. A preliminary isolation procedure of gliadin sub-fractions was achieved by another chromatographic technique (FPLC). The effect of gliadin on the phospholipid bilayer of giant vesicles was studied through the technique of micropipet aspiration coupled to videomicroscopy. The aim was to verify the possible effect of protein on the elasticity constant, kC of the bilayer. Results obtained can be summarized as follows: a) the viscoelastic behavior presented by gliadin was found to be dependent on the nature and composition of the dispersing media. Temperature-dependent tixotropic or rheopexic behavior were observed, as well as a cyclic behaviour for viscosity as a function of time. b) it was possible to correlate some aspects of solvent molecular structure and polarity with its greater or lesser capacity of gel formation: solvents with more expressive apolar regions, such as DMSO and DMF (ε = 47,24 and 38,25, respectively) induce formation of rigid gels, whereas a compound such as formamide, devoid of a minimally significant apolar domain and ε greater than that of water (ε = 84), is significantly less able to promote gel formation with the same characteristics of rigidity. c) gels are formed by units with gyration radius around 15Å and also by aggregates of greater size. Considering that the average molar mass in our sample for gliadin is 32350, one can assume that the protein is very densely packed in the gel (consider for instance that lysozyme, with a much lower molar mass, 14300 g/mol, has a gyration radius of this same size). A compact, interpenetrating folding model for gliadin, developed in our group through molecular dynamics, is compatible with such experimental observations. d) ω gliadin, selectively extracted in ethanol, was found to be able to reduce membrane elasticity of phospholipid microvesicles, by increasing its elastic curvature constant kC. A saturation effect was observed, for a mass ratio as low as gliadin/DOPC ~2-3%. e) diffusion coefficients for gliadin dispersions are circa 10-10 cm2.s-1, with hydrodynamical radii in the nm-µm range. Two dynamically distinct populations are identified in the gliadin gels. f) gel polydispersity and aggregate size depend on protein concentration: smaller aggregates (lower average molar mass) of more uniform sizes are produced for increasing gliadin concentration. Variation with time followed the same pattern, indicating that structural rearrangements take place during gel aging.

ASSUNTO(S)

rheology química coloidal gliadina gliadin protein reologia supramolecular chemistry proteína química supramolecular

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