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|Title:||Defining the wheat gluten peptide fingerprint via a discovery and targeted proteomics approach|
|Authors:||MARTINEZ ESTESO MARIA JOSÉ; NORGAARD Jorgen; BROHEE Marcel; HARASZI RÈKA; MAQUET Alain; O'CONNOR GAVIN|
|Citation:||JOURNAL OF PROTEOMICS vol. 147 p. 156-168|
|Publisher:||ELSEVIER SCIENCE BV|
|Type:||Articles in periodicals and books|
|Abstract:||Accurate, reliable and sensitive detection methods for gluten are required to support current EU regulations. The enforcement of legislative levels requires that measurement results are comparable over time and between methods. This is not a trivial task for gluten which comprises a large number of protein targets. This paper describes a strategy for defining a set of specific analytical targets for wheat gluten. A comprehensive proteomic approach was applied by fractionating wheat gluten using RP-HPLC (reversed phase high performance liquid chromatography) followed by a multi-enzymatic digestion (LysC, trypsin and chymotrypsin) with subsequent mass spectrometric analysis. This approach identified 434 peptide sequences from gluten. Peptides were grouped based on two criteria: unique to a single gluten protein sequence; contained known immunogenic and toxic sequences in the context of coeliac disease. An LC-MS/MS method based on a selected reaction monitoring method (SRM) was developed on a triple quadrupole mass spectrometer for the specific detection of the target peptides. The SRM based screening approach was applied to gluten containing cereals (wheat, rye, barley and oats) and non-gluten containing flours (corn, soy and rice). A unique set of wheat gluten marker peptides were identified and are proposed as wheat specific markers. They are representative of all the sub-fractions of the wheat gluten proteome and include peptide targets known to elicit an immune response in celiac disease patients. In order to make comparable gluten measurements a reality, the analytical measurement community requires: a set of agreed targets; known conversions from these targets to total gluten content; and appropriately characterised and commutable certified reference materials.|
|JRC Directorate:||Health, Consumers and Reference Materials|
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