Protein complex or compound three-dimensional gel electrophoresis separating method

Abstract

The invention discloses a protein complex or compound three-dimensional gel electrophoresis separating method. The method comprises the following steps: 1) separating the protein complex by non-denatured polyacrylamide gel electrophoresis method and then dyeing the protein complex; 2) chopping the gel strips containing the target protein complex or compound obtained in step 1), and decoloring and dehydrating; and 3) soaking the gel strips processed in step 2) by protein denaturation solution, adequately pulverizing and directly performing separation by two-dimensional gel electrophoresis method. The method in the invention dispenses with protein electroelution and protein deposition operating procedures in traditional protein complex or compound three-dimensional gel electrophoresis method, thus avoiding protein loss and protein in vitro chemical decoration; and the method in the invention is simple to operate and needs no additional equipment. Proved by experiments, the method in the invention achieves good protein complex or compound electrophoresis separating effect.

Description

technical field [0001] The invention relates to a three-dimensional gel electrophoresis separation method for protein complexes or complexes. Background technique [0002] Protein complex or complex three-dimensional gel electrophoresis (three-dimensional gel electrophoresis, 3-DE) technology was first proposed by Wolf Werhahn et al. (Werhahn, W., Braun, H.P., Electrophoresis 2002, 23, 640-646), it Refers to the separation of the protein mixture by blue non-denaturing polyacrylamide gel electrophoresis (bluenative polyacrylamide gel electrophoresis, BN-PAGE) or non-denaturing polyacrylamide gel electrophoresis (native polyacrylamide gel electrophoresis, native-PAGE) method, and then The conventional two-dimensional gel electrophoresis (two-dimensional gel electrophoresis, 2-DE) method was used for separation. The specific method is to separate the protein mixture in the mitochondria by blue non-denaturing polyacrylamide gel electrophoresis, cut off the gel band containing t...

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Problems solved by technology

[0003] However, the main disadvantage of this method is that the operation process of protein electroelution and protein precipitation is cumbersome, and special protein electroelution equipment is required, and this process will inevitably cause protein loss, and protein chemical modification in vitro is prone to occur; at the same time Since there may be large differences in the solubility of protein complexes or subunits in complexes, the recovery of each subunit will be inconsistent, and ultimately lead to inaccurate stoichiometry of the analyzed protein complexes or subunits in complexes

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