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Elucidation of structure-function relationships of soybean glycinin by protein engineering

通过蛋白质工程阐明大豆甘氨酸的结构与功能关系

基本信息

批准号：
04660144
负责人：
UTSUMI Shigeru
金额：
$ 1.34万
依托单位：
KYOTO UNIVERSITY
依托单位国家：
日本
项目类别：
Grant-in-Aid for General Scientific Research (C)
财政年份：
1992
资助国家：
日本
起止时间：
1992 至 1993
项目状态：
已结题

项目摘要

Glycinin is one of the dominant storage proteins of soybean seeds. The relationships between the structure and the food functional properties of glycinin were investigated at the molecular level by means of protein engineering. Glycinin was modified on the basis of its structural characteristics and the relationships between the structure and the functional properties elucidated at the subunit level. The functional properties of the modified proglycinins expressed in E.coli were measured. The results suggest that the hydrophobicity of the C-terminal region is closely related to emulsification, that the disulfide bond 12-45 plays an important role in the initiation of the disulfide exchange reaction required for gelation, that the number and the topology of free sulfhydryl residues are closely related to gelation, and that the factors of the structural instability desired for gelation and emulsification are different each other.Crystals of the normal and modified proglycinins (DELTAI, DELTAV8, IV+4Met, V+4Met, Gly12, Ser88) expressed in E.coli have been grown. The crystals of normal, DELTAI, V+4Met Gly12 and Ser88 diffracted X-rays sufficiently for crystallographic analysis. Normal, DELTAI, Clyl2 and Ser88 crystals belong to the tetragonal system, space group P4_1 or P4_3 with unit cell dimensions a=b=114.3-115.9 A and c=145.1-147.1 A.V+ 4Met crystals are monoclinic, belonging to space group P2 with unit cell parameters a=118.7 A, b=78.1 A, c=109.9 A and beta=119ﾟ. The number of protomers/asymmetric unit of these crystals is calculated to be around 3. This value is consistent with the fact that proglycinins are trimers. These data indicate that these crystals examined here are suitable for X-ray crystallography to elucidate the relationships between the structure and the functional properties of glycinin at molecular level.

大豆球蛋白是大豆种子的主要储存蛋白之一。通过蛋白质工程在分子水平上研究了甘氨酸的结构与食品功能特性之间的关系。根据其结构特征以及在亚基水平上阐明的结构与功能特性之间的关系，对大豆球蛋白进行了修饰。测量了在大肠杆菌中表达的修饰的大豆球蛋白原的功能特性。结果表明，C端区域的疏水性与乳化作用密切相关，二硫键12-45在凝胶化所需的二硫键交换反应的引发中起重要作用，游离巯基残基的数量和拓扑结构与凝胶化密切相关，凝胶化和乳化所需的结构不稳定的因素是不同的。在大肠杆菌中表达的修饰的大豆球蛋白原（DELTAI、DELTAV8、IV+4Met、V+4Met、Gly12、Ser88）已经生长。普通、DELTAI、V+4Met Gly12 和 Ser88 的晶体足以衍射 X 射线以进行晶体学分析。 Normal、DELTAI、Clyl2 和 Ser88 晶体属于四方晶系，空间群 P4_1 或 P4_3，晶胞尺寸 a=b=114.3-115.9 A 和 c=145.1-147.1 A。V+ 4Met 晶体是单斜晶系，属于空间群 P2，晶胞参数 a=118.7 A，b=78.1 A， c=109.9 A 且贝塔=119ﾟ。这些晶体的原聚体/不对称单元的数量经计算约为3。该值与大豆球蛋白原是三聚体的事实一致。这些数据表明，这里检查的这些晶体适用于 X 射线晶体学，以在分子水平上阐明大豆球蛋白的结构和功能特性之间的关系。