大豆球蛋白酸、碱性链分离及其酶交联特性的研究

Isolation of Acidic and Basic Polypeptides in Soybean Glycinin and Their Characteristics in Enzymatic Cross-linking

摘要:

本文以大豆为原料，分离大豆球蛋白中酸、碱性链，并探讨酸、碱性链在微生物转谷氨酰胺酶(MTG)介导下参与交联反应的差异性。利用等电点沉淀、凝胶过滤层析以及亲和层析分离得到纯度大于94%的大豆球蛋白，再通过热变性、β-巯基乙醇(β-ME)还原和等电点沉淀分离大豆球蛋白酸和碱性链，并对其进行了电泳鉴定和氨基酸分析；进一步以MTG催化大豆球蛋白及其酸、碱性链交联，利用SDS-PAGE和溶解性试验探讨大豆球蛋白酸、碱性链交联特性和交联产物的溶解性。结果表明大豆球蛋白酸、碱性链都是MTG的有效底物，均可参与交联反应并形成高聚物，但碱性链反应率较酸性链低，两者参与交联反应差异受其谷氨酰胺和赖氨酸残基含量的影响较小，而受其疏水性氨基酸含量影响较大。

Abstract:

Using soybean as the raw material, the acidic and basic polypeptides of glycinin were isolated, and the differences between their effects in microbial transglutaminase (MTG)-mediated cross-linking were explored. Glycinin was obtained with a purity of more than 94% from soybean protein by isoelectric precipitation, gel filtration chromatography, and affinity chromatography. The acidic and basic polypeptides were isolated by isoelectric precipitation after heat denaturation and reduction with β-mercaptoethanol, and then characterized by electrophoresis and amino acid analysis. Furthermore, sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and dissolution experiments were performed to determine the effects of acidic and basic polypeptides on enzymatic cross-linking and the solubility of the cross-linked products. The results suggested that both the acidic and basic polypeptides were effective substrates for MTG and could participate in the cross-linking reaction to form polymers with high molecular weight; however, the reaction rate of basic polypeptides was lower than that of acidic polypeptides. The differences between the acidic and basic polypeptides in the cross-linking reaction were slightly affected by the glutamine and lysine residue content, and were significantly affected by the hydrophobic amino acid content.