What Determines the Migration Direction of Proteins in an Electric Field
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In native PAGE, proteins maintain their natural state and are not denatured.
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The migration direction of proteins depends on their charge state at the pH of the electrophoresis buffer.
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When pH < pI, the protein carries a positive charge and migrates towards the negative electrode. When pH > pI, the protein carries a negative charge and migrates towards the positive electrode.
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In SDS-PAGE, proteins bind to SDS and become denatured, acquiring a uniform negative charge.
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As a result, all proteins migrate towards the positive electrode. In this case, migration speed mainly depends on protein size rather than charge state.
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IEF is a special type of electrophoresis that uses a gel with a pH gradient. Under an applied electric field, proteins migrate to their isoelectric points.
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At this position, the protein's net charge is zero, and it stops migrating.
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This technique combines IEF and SDS-PAGE.
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First, IEF is performed to separate proteins based on their isoelectric points.
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Then, SDS-PAGE is conducted to separate proteins by size. In this step, all proteins migrate towards the positive electrode.
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In capillary electrophoresis, samples migrate within a narrow capillary tube.
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The migration direction of proteins still depends on their charge state at the buffer pH and the denaturants used.
In different types of electrophoresis, the migration direction of proteins is determined by their charge state as well as the electrophoretic conditions and medium used. Below are several common types of electrophoresis and the factors determining protein migration direction:
1. Native PAGE (Non-denaturing Electrophoresis)
2. SDS-PAGE (Sodium Dodecyl Sulfate-Polyacrylamide Gel Electrophoresis)
3. Isoelectric Focusing (IEF)
4. Two-Dimensional Electrophoresis (2D-Electrophoresis)
5. Capillary Electrophoresis (CE)
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