SDS-PAGE法分离和表征明胶中不同构象

The Full Analysis of Different Conformations in Photo-Gelatins by SDS-PAGE Mothod

仔细分析已发表的文献表明,迄今为止用十二烷基硫酸钠-聚丙烯酰胺凝胶电泳(SDS-PAGE)测定明胶分子量分布仅限于分出<α、α_1、α_2、和β组份,而γ及>γ的组份未能检出,也就是说,分子量大约30万以上的组份未能进入凝胶板内(以下简称“入板”)。这对明胶构象的全分析和对明胶质量的判断都是一个严重的缺陷。本文主要探讨和研究了γ及>γ组份的入板问题。所采取的办法有:有效地控制丙烯酰胺和甲叉双丙烯酰胺(交联剂)的相对含量,选择合适的交联度梯度和孔径大小,使之既适用于α和β组份迁移,也能让γ及>γ的组份入板。此外,在小交联度和大孔径情况下,还能符合凝胶板对凝胶强度的基本要求。实验结果表时,操作过程缩短,分辨率改善,成功地得到了从几万到30万以上的分子量分布的明胶全部构象的图谱。

A careful survey and investigation of the literature references indicates that up to now SDS-PAGE method can be used to characterize the <α、α_1、 α_2、β conformations con- tained in gelatins.However the characterization of γ and > γ conformations failed to be realized by this method.It is really a serious fault in full analysis of different conformations of photo-gelatin and in judgement of the gelatin quality based on conformation compo- nents. In general the trouble how to deal with the contradiction between thc crosslinkage dc- gree(hence the pore magnitude)and the physico-mechanical property of palyacrylamide gel plate is acknowledged to be the main problem in realzing the full con- formation analysis. In present study a series of factors such as the relative concentration of acrylamide and thc N、 N′-methylene-bis-acrylamide and the cross-linkage gradient formation were in- vestigated as a function of the migration of all conformations with an emphasis to observe whcther the γ and >γ comporents enter the polyacrylamide gel plate with good resolution together with the other conformation components. The experimental data demonstrated that the entire conformation components ranging from <α、α_1 、α_2、β、γ and>γ without exception can be separated and characterized with satisfactory resolution by the SDS-PAGE method improved and developed in present study. This improvement and development are of important sigrificance for gelatin routine analysis and controlling the process parameters so as to optimize the correctly relative con- tent of different conformations for a specialized type of gelatin products.