Rapid Gastric Fluid Digestion and Biochemical Characterization of Engineered Proteins Enriched in Essential Amino Acids

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• 作者：Keith R. Roesler and A. Gururaj Rao
• 刊名：Journal of Agricultural and Food Chemistry
• 出版年：2001
• 出版时间：July 2001
• 年：2001
• 卷：49
• 期：7
• 页码：3443 - 3451
• 全文大小：198K
• 年卷期：v.49,no.7(July 2001)
• ISSN：1520-5118

文摘

The barley high lysine (BHL) proteins are nutritionally enhanced derivatives of barley chymotrypsininhibitor-2 (CI-2). A compactly folded new CI-2 derivative, BHL9, was engineered with the highestcontent of threonine, tryptophan, and isoleucine yet achieved in this protein family (15.1, 9.4, and12.1 wt %, respectively). BHL9 had an unfolding midpoint of 5.5 M guanidinium chloride,significantly greater than values for wild type (3.9 M) or for the previously most stable BHL protein,BHL8 (3.6 M). BHL9 and all other derivatives were digested within 15 s in simulated gastric fluid(SGF), suggesting nutritional availability upon ingestion. Denaturation of the proteins in SGF minuspepsin was revealed by changes in their fluorescence emission spectra and/or far UV circulardichroism spectra. The proteins lack homology to known allergens. Significantly, the BHL8 andBHL9 proteins were stable to proteases at pH 7.5 or 8.0, attesting to their potential for highexpression in plants.Keywords: Gastric fluid; intestinal fluid; protein digestion; nutritional proteins; protein engineering;protein stability