Using LC-MS Based Methods for Testing the Digestibility of a Nonpurified Transgenic Membrane Protein in Simulated Gastric Fluid

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• 作者：Wayne S. Skinner ; Brett S. Phinney ; Anthony Herren ; Floyd J. Goodstal ; Isabel Dicely ; Daniel Facciotti
• 刊名：Journal of Agricultural and Food Chemistry
• 出版年：2016
• 出版时间：June 29, 2016
• 年：2016
• 卷：64
• 期：25
• 页码：5251-5259
• 全文大小：661K
• 年卷期：0
• ISSN：1520-5118

文摘

The digestibility of a nonpurified transgenic membrane protein was determined in pepsin, as part of the food safety evaluation of its resistance to digestion and allergenic potential. Delta-6-desaturase from Saprolegnia diclina, a transmembrane protein expressed in safflower for the production of gamma linolenic acid in the seed, could not be obtained in a pure, native form as normally required for this assay. As a novel approach, the endoplasmic reticulum isolated from immature seeds was digested in simulated gastric fluid (SGF) and the degradation of delta-6-desaturase was selectively followed by SDS–PAGE and targeted LC-MS/MS quantification using stable isotope-labeled peptides as internal standards. The digestion of delta-6-desaturase by SGF was shown to be both rapid and complete. Less than 10% of the initial amount of D6D remained intact after 30 s, and no fragments large enough (>3 kDa) to elicit a type I allergenic response remained after 60 min.