二维凝胶电泳分离的蛋白质之间氨基酸组成相似性的测定。

The determination of similarities in amino acid composition among proteins separated by two-dimensional gel electrophoresis.

作者信息

Cabral F, Gottesman M M

机构信息

Laboratory of Molecular Biology, National Cancer Institute, Bethesda, Maryland 20014, USA.

出版信息

Anal Biochem. 1978 Dec;91(2):548-56. doi: 10.1016/0003-2697(78)90542-0.

DOI:10.1016/0003-2697(78)90542-0

PMID:9762142

Abstract

A simple and rapid procedure has been developed to determine similarities in amino acid composition among cellular proteins separated by two-dimensional gel electrophoresis. Cells in tissue culture are simultaneously labeled with two different amino acids each tagged with a different radioisotope. The proteins are then separated on two-dimensional gels and their location on the gels determined by Coomassie-blue staining or autoradiography. Elution of the protein from the appropriate region of the gel followed by liquid scintillation counting yields an isotope ratio which reflects the ratio of the two amino acids in the protein. Examples of the use of this technique in analyzing mutant proteins, proteins altered by carbamylation, and cell proteins with similar amino acid composition (e.g., actin and tubulin) are given.

摘要

已开发出一种简单快速的方法来确定通过二维凝胶电泳分离的细胞蛋白质之间氨基酸组成的相似性。组织培养中的细胞同时用两种不同的氨基酸进行标记，每种氨基酸都带有不同的放射性同位素。然后将蛋白质在二维凝胶上分离，并通过考马斯亮蓝染色或放射自显影确定它们在凝胶上的位置。从凝胶的适当区域洗脱蛋白质，然后进行液体闪烁计数，得到一个同位素比率，该比率反映了蛋白质中两种氨基酸的比率。给出了该技术在分析突变蛋白、经氨甲酰化改变的蛋白以及具有相似氨基酸组成的细胞蛋白（如肌动蛋白和微管蛋白）中的应用实例。