Why Does Protein Electrophoresis Move Toward the Negative Electrode
In protein electrophoresis, whether a protein migrates toward the negative or positive electrode depends on its charge in the electrophoresis buffer. A protein’s charge is determined by the charges of its amino acid residues, which vary with pH. The charge of a protein is influenced by the solution's pH and its isoelectric point (pI).
The isoelectric point (pI) is the pH at which a protein has a net charge of zero. Below this pH, the protein carries a positive charge; above this pH, it carries a negative charge.
In traditional protein electrophoresis (e.g., Native-PAGE), if the buffer pH is lower than the protein’s pI, the protein carries a positive charge and migrates toward the negative electrode. Conversely, if the buffer pH is higher than the protein’s pI, the protein carries a negative charge and migrates toward the positive electrode.
However, in SDS-PAGE, things are different. SDS (Sodium Dodecyl Sulfate) is an anionic surfactant that binds to proteins, linearizes them, and imparts a nearly uniform negative charge. This negative charge causes proteins in SDS-PAGE to migrate almost entirely based on size rather than their original charge. Due to the presence of SDS, proteins always carry a negative charge and migrate toward the anode (positive electrode).
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