Changes in the ovalbumin proteolysis profile by high pressure and its effect on IgG and IgE binding.
Egg proteins are responsible for one of the most common forms of food allergy, especially in children, and one of the major allergens is ovalbumin (OVA). With the aim to examine the potential of high pressure to enhance the enzymatic hydrolysis of OVA and modify its immunoreactivity, the protein was proteolyzed with pepsin under high-pressure conditions (400 MPa). Characterization of the hydrolysates and peptide identification was performed by reversed-phase high-performance liquid chromatography-tandem mass spectrometry (RP-HPLC-MS/MS). The antigenicity (binding to IgG) and binding to IgE, using the sera of patients with specific IgE to OVA, were also assessed. The results showed that, upon treatment with pepsin at 400 MPa, all of the intact protein was removed in minutes, leading to the production of hydrolysates with lower antigenicity than those produced in hours at atmospheric pressure. However, the exposure of new target residues only partially facilitated the removal of allergenic epitopes, because the hydrolysates retained residual IgG- and IgE-binding properties as a result of the accumulation of large and hydrophobic peptides during the initial stages of hydrolysis. These peptides disappeared at later stages of proteolysis, although reactivity toward IgG and IgE was not completely abolished. Some fragments identified in the hydrolysates (such as Leu124-Phe134, Ile178-Ala187, Leu242-Leu252, Gly251-Ile259, Lys322-Gly343, Phe358-Phe366, and Phe378-Pro385) carried previously identified IgE-binding epitopes. Because some of the peptides found, such as Phe358-Phe366, probably contain only one binding site for IgE, the possibility to use high pressure to tailor hydrolysates that contain mostly peptides with only one IgE-binding site, which may help the immune system to tolerate egg proteins, is suggested.
López-Expósito
Ivan
I
Instituto de Fermentaciones Industriales, Consejo Superior de Investigaciones Científicas, Juan de la Cierva 3, Madrid 28006, Spain.
Chicón
Rosa
R
Belloque
Josefina
J
Recio
Isidra
I
Alonso
Elena
E
López-Fandiño
Rosina
R
D000818:Animals; D002648:Child; D002851:Chromatography, High Pressure Liquid; D005260:Female; D005512:Food Hypersensitivity / Q000276:immunology*; D006801:Humans; D006868:Hydrolysis; D007073:Immunoglobulin E / Q000276:immunology*; D007074:Immunoglobulin G / Q000276:immunology*; D008297:Male; D010047:Ovalbumin / Q000737:chemistry* / Q000276:immunology*; D010446:Peptide Fragments / Q000737:chemistry / Q000276:immunology; D011312:Pressure; D011485:Protein Binding; D053719:Tandem Mass Spectrometry
D007074:Immunoglobulin G; D010446:Peptide Fragments; D007073:Immunoglobulin E; D010047:Ovalbumin