Protein Purification - Basic 10 Flashcards
(10 cards)
What is the primary goal of protein purification?
a) To identify protein structure
b) To synthesize new proteins
c) To isolate a specific protein from a complex mixture
d) To denature all proteins in a sample
Answer: c) To isolate a specific protein from a complex mixture
Explanation: Protein purification focuses on separating and isolating a target protein from a mixture of many proteins and biomolecules. This allows further study of the protein’s properties, function, and structure.
Which of the following techniques separates proteins based on size?
a) Ion-exchange chromatography
b) Gel-filtration chromatography
c) Affinity chromatography
d) Isoelectric focusing
Answer: b) Gel-filtration chromatography
Explanation: Gel-filtration (size exclusion) chromatography separates proteins based on size. Larger proteins elute first because they don’t enter the pores of the gel matrix, while smaller proteins are delayed.
Which method separates proteins based on charge?
a) Size-exclusion chromatography
b) Ion-exchange chromatography
c) SDS-PAGE
d) Gel-filtration chromatography
Answer: b) Ion-exchange chromatography
Explanation: Ion-exchange chromatography separates proteins based on their net charge by using charged resin beads. Proteins with opposite charges to the resin bind, while others elute more quickly.
What is the role of SDS in SDS-PAGE?
a) To give proteins a uniform negative charge
b) To dye the protein for visualization
c) To preserve protein structure
d) To reduce protein aggregation
Answer: a) To give proteins a uniform negative charge
Explanation: SDS denatures proteins and coats them with a uniform negative charge. This ensures separation by size alone during electrophoresis.
Which technique uses a specific binding interaction to isolate a target protein?
a) Gel filtration
b) Affinity chromatography
c) Isoelectric focusing
d) Western blotting
Answer: b) Affinity chromatography
Explanation: Affinity chromatography relies on the specific interaction between the protein of interest and a ligand attached to the resin, allowing high specificity in purification.
Which of the following will elute LAST in gel-filtration chromatography?
a) Large proteins
b) Medium-sized proteins
c) Proteins with high charge
d) Small proteins
Answer: d) Small proteins
Explanation: Small proteins enter the pores of the beads in gel-filtration and thus travel a longer path, eluting later than large proteins, which are excluded from the pores.
In ion-exchange chromatography, what condition typically elutes bound proteins?
a) Decreasing pH
b) Adding SDS
c) Increasing salt concentration
d) Lowering temperature
Answer: c) Increasing salt concentration
Explanation: Increasing salt concentration disrupts electrostatic interactions between proteins and the charged resin, allowing bound proteins to elute.
What does the specific activity of a protein measure?
a) Enzyme purity
b) Enzyme catalytic rate
c) Protein concentration
d) Molecular weight
Answer: a) Enzyme purity
Explanation: Specific activity is the ratio of enzyme activity to the amount of protein present, and it increases as the protein becomes purer.
Which of the following is a common first step in protein purification?
a) Gel-filtration chromatography
b) SDS-PAGE
c) Cell lysis
d) Western blotting
Answer: c) Cell lysis
Explanation: Protein purification typically begins with lysis to release the proteins from inside.
Which method separates proteins based on their isoelectric point?
a) Gel-filtration chromatography
b) Affinity chromatography
c) SDS-PAGE
d) Isoelectric focusing
Answer: d) Isoelectric focusing
Explanation: Isoelectric focusing separates proteins based on their pI—the pH at which they carry no net charge—causing each protein to migrate to and focus at its pI
