Influence of denaturation and aggregation of β-lactoglobulin on its tryptic hydrolysis and the release of functional peptides

Leeb, Elena; Götz, Alexander; Letzel, Thomas; Cheison, Seronei Chelulei; Kulozik, Ulrich

doi:10.1016/j.foodchem.2015.04.034

Cited by 58 publications

(28 citation statements)

References 33 publications

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“…At pH 4, which is relatively close to the pI of β-Lg, the net charge of protein is close to zero, and so the repulsive electrostatic forces weaken and the protein tends to aggregate, thus resulting in higher particle size values (Salgin, Salgin, & Bahadir, 2012). These observations agree with those reported by Leeb, Gotz, Letzel, Cheison, & Kulozik, 2015, which showed that β-Lg (5 mg mL −1 ) prepared at pH 5.1 formed structures with particle sizes of 1332.1 ± 56.3 nm during thermal heating at 80°C…”

Section: Development Of Micro-and Nanostructuressupporting

confidence: 93%

Design of β-lactoglobulin micro- and nanostructures by controlling gelation through physical variables

et al. 2020

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A B S T R A C Tβ-lactoglobulin (β-Lg) is the major protein fraction of bovine whey serum and its principal gelling agent. Its gelation capacity enables conformational changes associated with protein-protein interactions that allow the design of structures with different properties and morphologies. Thus, the aim of this work was to successfully use β-Lg, purified from a commercial whey protein isolate, to develop food-grade micro-(with diameters between 200 and 300 nm) and nano-(with diameters ≤ 100 nm) structures. For this purpose, the phenomena involved in β-Lg gelation were studied under combined effects of concentrations (from 5 to 15 mg mL −1 ), heating temperature (from 60 to 80°C) and heating time (from 5 to 25 min) for pH values of 3, 4, 6 and 7. The effects of such conditions on β-Lg structures were evaluated and the protein was fully characterized in terms of size, polydispersity index (PDI) and surface charge (by dynamic light scattering -DLS), morphology (by transmission electron microscopy -TEM) and conformational structure (circular dichroism, intrinsic and extrinsic fluorescence). Results have shown that β-Lg nanostructures were formed at pH 3 (with diameters between 12.1 and 22.3 nm) and at 7 (with diameters between 8.9 and 35.3 nm). At pH 4 structures were obtained at macroscale (i.e., ≥ 6 μm) for all β-Lg concentrations when heated at 70 and 80°C, independent of the time of heating. For pH 6, it was possible to obtain β-Lg structures either at micro-(245.0 -266.4 nm) or nanoscale (≤ 100 nm) with the lowest polydispersity (PDI) values (≤ 0.25), in accordance with TEM analyses, for heating at 80°C for 15 min. Intrinsic and extrinsic fluorescence data and far-UV circular dichroism spectra measurements revealed conformational changes on β-Lg structure that support these evidences. A strict control of the physical and environmental conditions is crucial for developing β-Lg structures with the desired characteristics, thus calling for the understanding of the mechanisms of protein aggregation and intermolecular interaction when designing β-Lg structures with novel functionalities.

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Section: Development Of Micro-and Nanostructuressupporting

confidence: 93%

Design of β-lactoglobulin micro- and nanostructures by controlling gelation through physical variables

et al. 2020

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“…Along the hydrolysis process, the reaction velocity continuously decreased, especially using free trypsin. The final DH within 4 h by 1.5 mg free trypsin was only around 6%, while its theoretical DH max is 11.18% . The effects of autodigestion, reduction of substrate and potential inhibition of products can account for this result.…”

Section: Resultsmentioning

confidence: 92%

“…Enzymatic hydrolysis of this protein is a common approach to reduce allergenicity. The use of the serine protease Trypsin (EC 3.4.21.4), which preferably cleaves the C‐terminal peptide bonds of Arginine (Arg/R) and Lysine (Lys/K), not only can significantly reduce its allergenicity , but also leads to the release of five biofunctional peptides . In addition, Schmidt and Poll showed that native α‐Lactalbumin (α‐La) was highly resistant to tryptic digestion while native β‐Lg was not.…”

Section: Introductionmentioning

confidence: 99%

Production of β‐Lactoglobulin hydrolysates by monolith based immobilized trypsin reactors

et al. 2017

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Tryptic hydrolysis of β-Lactoglobulin (β-Lg) is attracting more and more attention due to the reduced allergenicity and the functionality of resulting hydrolysates. To produce hydrolysates in an economically viable way, immobilized trypsin reactors (IMTRs), based on polymethacrylate monolith with pore size 2.1 μm (N1) and 6 μm (N2), were developed and used in a flow-through system. IMTRs were characterized in terms of permeability and enzymatic activity during extensive usage. N1 showed twice the activity compared with N2, correlating well with its almost two times higher amount of immobilized trypsin. N2 showed high stability over 18 cycles, as well as over more than 30 weeks during storage. The efficiency of IMTRs on hydrolyzing β-Lg was compared with free trypsin, and the resulting hydrolysates were analyzed by MALDI-TOF/MS. The final hydrolysis degree by N1 reached 9.68% (86.58% cleavage sites) within 4 h, while only around 6% (53.67% cleavage sites) by 1.5 mg of free trypsin. Peptides analysis showed the different preference between immobilized trypsin and free trypsin. Under the experimental conditions used in this study, the potential cleavage site Lys -Phe was resistant against the immobilized trypsin in N1.

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“…Therefore native-like secondary structure of BLG remains intact, but the tight packing of the native tertiary structure is undergoing conformational changes. Above 65 °C, larger secondary and tertiary structural change occurs and leads to thiol/disulphide exchange mechanism and corresponding unfolding and exposure of hydrophobic portions of the protein, promoting aggregation of the protein with a hydrodynamic radius much larger than that of the pore diameter of matrix support [ 48 , 49 , 50 , 51 , 52 ].…”

Section: Resultsmentioning

confidence: 99%

Combined Spectroscopic and Calorimetric Studies to Reveal Absorption Mechanisms and Conformational Changes of Protein on Nanoporous Biomaterials

Ahmadi

Farokhi

Padidar

et al. 2015

IJMS

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In this study the effect of surface modification of mesoporous silica nanoparticles (MSNs) on its adsorption capacities and protein stability after immobilization of beta-lactoglobulin B (BLG-B) was investigated. For this purpose, non-functionalized (KIT-6) and aminopropyl-functionalized cubic Ia3d mesoporous silica ([n-PrNH2-KIT-6]) nanoparticles were used as nanoporous supports. Aminopropyl-functionalized mesoporous nanoparticles exhibited more potential candidates for BLG-B adsorption and minimum BLG leaching than non-functionalized nanoparticles. It was observed that the amount of adsorbed BLG is dependent on the initial BLG concentration for both KIT-6 and [n-PrNH2-KIT-6] mesoporous nanoparticles. Also larger amounts of BLG-B on KIT-6 was immobilized upon raising the temperature of the medium from 4 to 55 °C while such increase was undetectable in the case of immobilization of BLG-B on the [n-PrNH2-KIT-6]. At temperatures above 55 °C the amounts of adsorbed BLG on both studied nanomaterials decreased significantly. By Differential scanning calorimetry or DSC analysis the heterogeneity of the protein solution and increase in Tm may indicate that immobilization of BLG-B onto the modified KIT-6 results in higher thermal stability compared to unmodified one. The obtained results provide several crucial factors in determining the mechanism(s) of protein adsorption and stability on the nanostructured solid supports and the development of engineered nano-biomaterials for controlled drug-delivery systems and biomimetic interfaces for the immobilization of living cells.

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Influence of denaturation and aggregation of β-lactoglobulin on its tryptic hydrolysis and the release of functional peptides

Cited by 58 publications

References 33 publications

Design of β-lactoglobulin micro- and nanostructures by controlling gelation through physical variables

Design of β-lactoglobulin micro- and nanostructures by controlling gelation through physical variables

Production of β‐Lactoglobulin hydrolysates by monolith based immobilized trypsin reactors

Combined Spectroscopic and Calorimetric Studies to Reveal Absorption Mechanisms and Conformational Changes of Protein on Nanoporous Biomaterials

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