The following statements describe SDS-PAGE, a method used to separate proteins.

Question

The following statements describe SDS-PAGE, a method used to separate proteins. Select the true statements. Assume that SDS-PAGE is performed under reducing conditions.

A.A protein binds roughly 1.4 times its mass of SDS resulting in a large overall negative charge.

B.SDS-PAGE utilizes agarose gel to separate proteins.

C.Smaller proteins migrate faster through the polyacrylamide gel.

D.Sodium dodecyl sulfate binds proteins, resulting in protein-SDS complexes that are similar in size.

E.Protein-SDS complexes migrate toward the positive electrode.

F. Proteins are visualized using a dye that binds to the gel matrix, but not to proteins.

Explanation / Answer

A. False.

B. False

C. True

D. False

E. True

F. False

SDS-PAGE(sodium dodecyl sulfate polyacrylamide gel electrophoresis) is an analytical method used to separate charged molecules in mixture on the basis of their molecular masses in an electric field. SDS-PAGE is commonly used to separate proteins on the basis of their weight by making them pass through a polyacrylamide-based discontinuous gel that act as separation medium(known as matrix) under electric field, SDS is an anionic surfactant. About 1.4gms of SDS binds to 1gm of protein. This masks the intrinsic charge of the protein and confers uniform charge distribution to the proteins. The uniform negative charge causes mutual repulsion between protein molecules, this combined with linearization of folded protein under denaturing conditions(Boiling) allows for separation of proteins by chain length, proportional to the molecular mass, as the longer proteins are more strongly retained in the gel while smaller chain proteins migrate faster. Having uniform negative charge all protein molecules will migrate towards positive electrode when an electric field is applied. After separation the proteins can visualized by staining with dyes coomassie brilliant blue. The dye interacts electrostatically but noncovalently with the amino and carboxyl groups of proteins to form a protein dye complex. This stabilises negatively charged anionic form of the dye producing the blue colour

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