Navegando por Autor "Tavares, Guilherme M."

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Binding of folic acid induces specific self-aggregation of lactoferrin: thermodynamic characterization
(Langmuir, 2015-10-21) Tavares, Guilherme M.; Croguennec, Thomas; Le, Sébastien; Lerideau, Olivia; Hamon, Pascaline; Carvalho, Antônio F.; Bouhallab, Saïd
In the study presented here, we investigated the interaction at pH 5.5 between folic acid (FA) and lactoferrin (LF), a positively charged protein. We found a binding constant Ka of 10(5) M(-1) and a high stoichiometry of 10 mol of FA/mol of LF. The size and charge of the complexes formed evolved during titration experiments. Increasing the ionic strength to 50 mM completely abolished the isothermal titration calorimetry (ITC) signal, suggesting the predominance of electrostatic interactions in the exothermic binding obtained. We developed a theoretical model that explains the complex triphasic ITC profile. Our results revealed a two-step mechanism: FA/LF interaction followed by self-association of the complexes thus formed. We suggest that 10 FA molecules bind to LF to form saturated reactive complexes (FA10/LF) that further self-associate into aggregates with a finite size of around 15 nm. There is thus a critical saturation degree of the protein, above which the self-association can take place. We present here the first results that provide comprehensive details of the thermodynamics of FA/LF complexation-association. Given the high stoichiometry, allowing a load of 55 mg of FA/g of LF, we suggest that FA/LF aggregates would be an effective vehicle for FA in fortified drinks.
Heteroprotein complex coacervation: a generic process
(Advances in Colloid and Interface Science, 2017-06-17) Croguennec, Thomas; Tavares, Guilherme M.; Bouhallab, Saïd
Proteins exhibit a rich diversity of functional, physico-chemical and biodegradable properties which makes them appealing for various applications in the food and non-food sectors. Such properties are attributed to their ability to interact and assemble into a diversity of supramolecular structures. The present review addresses the updated research progress in the recent field of complex coacervation made from mixtures of oppositely charged proteins (i.e. heteroprotein systems). First, we describe briefly the main proteins used for heteroprotein coacervation. Then, through some selected examples, we illustrate the particularity and specificity of each heteroprotein system and the requirements that drive optimal assembly into coacervates. Finally, possible and promising applications of heteroprotein coacervates are mentioned.
How the presence of a small molecule affects the complex coacervation between lactoferrin and β-lactoglobulin
(International Journal of Biological Macromolecules, 2017-09) Tavares, Guilherme M.; Croguennec, Thomas; Hamon, Pascaline; Carvalho, Antônio F.; Saïd Bouhallab
Heteroprotein complex coacervation corresponds to the formation of two liquid phases in equilibrium induced by the interaction of two oppositely charged proteins. The more concentrated phase known as coacervate phase, has attracted interest from several fields of science due to its potential applications for example for encapsulation and delivery of bioactives. Prior such application, it is necessary to understand how the presence of small ligands affects the complex coacervation. In this work, we report on the interaction of small ligand with individual proteins β-lactoglobulin (β-LG) and lactoferrin (LF) and consequences on their complex coacervation. ANS (8-Anilinonaphthalene-1-sulfonic acid), a fluorescent probe, was used as model ligand. While ANS did not interact with β-LG, it presented two sets of binding sites with LF inducing its self-aggregation. Depending on its concentration, ANS modulated the shape of β-LG-LF macromolecular assembly. Coacervates were observed for ANS/LF molar ratio <25 against amorphous aggregates for higher ANS/LF molar ratios. A maximum loading capacity of around 40 mg of ANS per gram of LF in the formed heteroprotein coacervates was reached.
Selective coacervation between lactoferrin and the two isoforms of β-lactoglobulin
(Food Hydrocolloids, 2015-06) Tavares, Guilherme M.; Croguennec, Thomas; Hamon, Pascaline; Carvalho, Antônio F.; Bouhallab, Saïd
This work reports on the impact of subtle change of protein charge on coacervation and subsequent liquid–liquid phase separation between two oppositely charged globular proteins. For this purpose, a comparative study was conducted on the coacervation of lactoferrin (LF) with the two β-lactoglobulin (β-LG) isoforms. Upon mixing LF with an excess of β-LG, microspheres were formed throughout coacervation under narrow pH range (5.4–6.0). At the optimal pH of coacervation, LF being the limiting partner under tested concentration ranges. The β-LG/LF molar ratio recovered in the formed coacervates varied from 4 to 8 depending on the total protein concentration. Remarkably, LF showed a selective coacervation with isoform A of β-LG as judged by a larger concentration domain for coacervation and a high yield of LF recovered once mixed with β-LG A i.e. 80% against a maximum of 42% with β-LG B. At thermodynamic level, the interaction of LF with both β-LG isoforms exhibited complex exothermic binding isotherms with both enthalpic and entropic contributions.
Structure and dynamics of heteroprotein coacervates
(Langmuir, 2016-06-29) Peixoto, Paulo D. S.; Tavares, Guilherme M.; Croguennec, Thomas; Nicolas, Aurélie; Hamon, Pascaline; Roiland, Claire; Bouhallab, Saïd
Under specific conditions, mixing two oppo-sitely charged proteins induces liquid−liquid phase separation. The denser phase, or coacervate phase, can be potentially applied as a system to protect or encapsulate different bioactive molecules with a broad range of food and/or medical applications. The optimization of the design and efficiency of such systems requires a precise understanding of the structure and the equilibrium of the nanocomplexes formed within the coacervate. Here, we report on the nanocomplexes and the dynamics of the coacervates formed by two well-known, oppositely charged proteins β-lactoglobulin (β-LG, pI ≈ 5.2) and lactoferrin (LF, pI ≈ 8.5). Fluorescence recovery after photobleaching (FRAP) and solid-state nuclear magnetic resonance (NMR) experiments indicate the coexistence of several nanocomplexes as the primary units for the coacervation. To our knowledge, this is the first evidence of the occurrence of an equilibrium between quite unstable nanocomplexes in the coacervate phase. Combined with in silico docking experiments, these data support the fact that coacervation in the present heteroprotein system depends not only on the structural composition of the coacervates but also on the association rates of the proteins forming the nanocomplexes.
Technological aspects of lactose-hydrolyzed milk powder
(Food Research International, 2017-08-24) Torres, Jansen Kelis Ferreira; Stephani, Rodrigo; Tavares, Guilherme M.; Carvalho, Antônio Fernandes de; Costa, Renata Golin Bueno; Almeida, Carlos Eduardo Rocha de; Almeida, Mariana Ramos; Oliveira, Luiz Fernando Cappa de; Schuck, Pierre; Perrone, Ítalo Tuler
Few reports describe the effect of lactose hydrolysis on the properties of milk powder during production and storage. Hence, the aim of this study was to evaluate the effects of five different levels of enzymatic lactose hydrolysis during the production and storage of milk powder. As the lactose hydrolysis rate increased, adhesion to the drying chamber also increased, due to higher levels of particle agglomeration. Additionally, more brown powder was obtained when the lactose hydrolysis rate was increased, which in turn negatively affected rehydration ability. Using Raman spectroscopy, crystallization of the lactose residues in various samples was assessed over 6 weeks of accelerated aging at a room temperature environment with 75.5% of air moisture. Products with 25% or greater lactose hydrolysis showed no signs of crystallization, in contrast to the non-hydrolyzed sample.