Question

part a – multiple choice

1. the main purpose of sds in sds - page is to:

a. stain proteins b. break peptide bonds c. denature proteins and give them a uniform negative charge d. transfer proteins to a membrane

1. in sds - page, proteins are separated primarily by:

a. charge b. shape c. size (molecular weight) d. color

1. after electrophoresis, proteins are transferred to a ______ membrane for western blotting.

a. agarose b. nitrocellulose c. glass d. plastic

1. the molecule that specifically detects the protein of interest in a western blot is a:

a. substrate b. enzyme c. antibody d. dye

Explanation:

Question 1

SDS (sodium dodecyl sulfate) in SDS - PAGE is used to denature proteins (break their non - covalent bonds like hydrogen bonds, hydrophobic interactions etc.) and also gives them a uniform negative charge. This eliminates the effect of the protein's native charge and shape on migration during electrophoresis, so that separation is based on molecular weight. Option A is wrong as staining is done after electrophoresis. Option B is wrong as SDS does not break peptide bonds (that's done by proteases). Option D is wrong as the transfer to membrane is part of Western blotting, not the role of SDS in SDS - PAGE.

In SDS - PAGE, after SDS denatures the proteins and gives them a uniform negative charge, the proteins migrate through the polyacrylamide gel based on their size (molecular weight). The smaller the molecular weight, the faster the protein migrates. Charge is made uniform by SDS, so option A is wrong. Shape is also standardized by the denaturing effect of SDS, so option B is wrong. Color has nothing to do with the separation in SDS - PAGE (staining for color comes after), so option D is wrong.

After electrophoresis in Western blotting, proteins are transferred to a nitrocellulose membrane (or sometimes PVDF membrane). Agarose is used in agarose gel electrophoresis (for nucleic acids or sometimes proteins in native conditions but not for Western blot transfer). Glass and plastic are not used as membranes for Western blotting protein transfer.

Answer:

C. Denature proteins and give them a uniform negative charge

Question 2