Q: A 1.0 mL protein solution in 0.500 M Tris, pH 7 with 0.250 M NaCl is dialyzed against 500 mL of 0.500 M Tris, pH 7. What is the final concentration of NaCl in the protein solution at equilibrium? Use the formula $$C_A \times V_A = C_B \times V_B.$$

$$C_B = \frac{0.250 \times 1.0}{500} = 0.0005\ \mathrm{M}$$

Question 1

Which laboratory method is commonly used to remove small molecules and ions from protein solutions, and what principle does it rely on?

Accepted Answer

Dialysis, which relies on the principle of diffusion across a semi-permeable membrane

Question 2

A 1.0 mL protein solution in 0.500 M Tris, pH 7 with 0.250 M NaCl is dialyzed against 500 mL of 0.500 M Tris, pH 7. What is the final concentration of NaCl in the protein solution at equilibrium? Use the formula $$C_A 	imes V_A = C_B 	imes V_B.$$

Accepted Answer

$$C_B = \frac{0.250 	imes 1.0}{500} = 0.0005\ \mathrm{M}$$

Question 3

In ion exchange chromatography, what type of resin would you use to separate proteins with a net negative charge at pH 7?

Accepted Answer

Anion exchanger, such as DEAE-cellulose

Question 4

Which of the following best describes the principle behind size exclusion (gel filtration) chromatography?

Accepted Answer

Molecules are separated based on their size, with larger molecules eluting first

Question 5

You have a mixture of three proteins with molecular weights: A) 13.9 kDa, B) 45.6 kDa, and C) 120 kDa. What is the order of elution from a size exclusion chromatography column?

Accepted Answer

C (120 kDa) first, then B (45.6 kDa), then A (13.9 kDa)

Question 6

In SDS-PAGE, what property of proteins determines their rate of migration through the gel?

Accepted Answer

Molecular weight, with smaller proteins migrating faster

Question 7

A protein subjected to SDS-PAGE under non-reducing conditions shows two bands at 200 kDa and 120 kDa. Under reducing conditions, the bands are at 120 kDa and 100 kDa. What does this suggest about the protein's structure?

Accepted Answer

The protein is composed of subunits held together by disulfide bonds

Question 8

Which method would you use to separate proteins based on their isoelectric point (pI)?

Accepted Answer

Isoelectric focusing

Question 9

In ion exchange chromatography, which protein will elute first from a DEAE-cellulose column at pH 7: a protein with pI 5.0, pI 7.0, or pI 9.0?

Accepted Answer

Protein with pI 9.0

Question 10

Which of the following is TRUE regarding affinity chromatography?

Accepted Answer

It separates proteins based on specific interactions with a ligand bound to the column

Question 11

After purifying a peptide, which method would you use to determine its amino acid composition?

Accepted Answer

Complete hydrolysis followed by ion exchange chromatography

Question 12

Which dye is commonly used to identify amino acids after ion exchange chromatography?

Accepted Answer

Ninhydrin

Question 13

What is the main advantage of 2D electrophoresis (IEF + SDS-PAGE) over single-dimension electrophoresis?

Accepted Answer

It allows separation of proteins based on both isoelectric point and molecular weight

Question 14

Which of the following statements about Bruce Merrifield's contribution to biochemistry is correct?

Accepted Answer

He developed the automated solid-phase synthesis of proteins

Question 15

Which equation would you use to calculate the concentration of a solute after dialysis, given initial and final volumes and concentrations?

Accepted Answer

$$C_A 	imes V_A = C_B 	imes V_B$$

Question 16

Which protein purification method would be most suitable for isolating a protein that binds specifically to ATP?

Accepted Answer

Affinity chromatography with ATP immobilized on the column

Question 17

During agarose gel electrophoresis, which direction do cations migrate?

Accepted Answer

Toward the cathode (negative electrode)

Question 18

Which of the following is a common misconception about SDS-PAGE?

Accepted Answer

Proteins are separated by their molecular weight

Question 19

What is the main reason smaller proteins migrate faster in SDS-PAGE?

Accepted Answer

They experience less resistance moving through the gel matrix

Laboratory Methods for Protein Isolation, Purification, and Characterization

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