用有限酶切法分析蛋白质的氨基酸序列

报道了一个通过有限酶切蛋白质产生多肽片段的方法.蛋白质经单向SDS-聚丙烯酰胺凝胶电泳(SDS-PAGE)分离和用考马斯亮蓝短暂染色后，切下所需的蛋白质带，将其放入另一个SDS-PAGE凝胶的样品槽内，在电泳过程中该蛋白质被蛋白酶如蛋白酶V8降解，所产生的多肽片段随之被分离.电泳结束后，将多肽片段电印迹至聚偏二氟乙烯(polyvinylidene difluoride,PVDF)膜上.这些多肽片段从PVDF膜上切下后可以直接被用于分析氨基酸序列.该方法能广泛适用于分析一般蛋白质和N端被修饰蛋白质的氨基酸序列.

Internal Amino Acid Sequence Analysis of Proteins After Limited Proteolytic Cleavage

A procedure was used to obtain peptide fragments for sequence analysis from proteins separated by gel electrophoresis.After separation by SDS-polyacrylamide gel electrophoresis (SDS-PAGE),proteins were briefly stained with Coomassie blue,interested protein-containing bands were cut and loaded onto the wells of a new SDS-PAGE gel.The protein was digested by protease V8 during electrophoresis,the resulting peptide fragments were electroblotted onto polyvinylidene difluoride membrane,and then sequenced with protein sequencer.The method can be used not only to obtain amino acid sequences from N-terminal blocked proteins,but also to produce multiple and independent amino acid sequence information from normal proteins.