This set of Biophysics Multiple Choice Questions & Answers (MCQs) focuses on “Affinity Chromatography”.
1. What type of interaction happens in Affinity chromatography between a protein and a ligand?
a) Covalent
b) Noncovalent
c) Ionic
d) Ionic + covalent interaction
View Answer
Explanation: The type of interaction happens in Affinity chromatography between a protein and a ligand is a non-covalent interaction. But the interaction happens between a ligand and the matrix is a covalent type of interaction.
2. Affinity chromatography is a __________________
a) Liquid – Liquid chromatography
b) Gas-Liquid chromatography
c) Solid – Gas chromatography
d) Solid-liquid chromatography
View Answer
Explanation: Affinity chromatography is basically a Liquid-Liquid chromatography because the here the interacting station phase is a solid which is a matrix-bound ligand. On the other hand, the mobile phase is solid that is protein solution.
3. Where ligands are used in an affinity chromatography?
a) Ligands are immobilized onto the column
b) Ligands are immobilized onto the protein
c) Ligands are not used at all
d) Ligands used as an eluents
View Answer
Explanation: In affinity chromatography, the ligands are immobilized onto the column to serve as a stationary phase for the protein separation. The other options are invalid as proteins never used as a stationary phase.
4. What is the chemical property of the matrix used in an affinity chromatography?
a) Ionic
b) Inert
c) Acidic
d) Basic
View Answer
Explanation: The matrix should be inert so that it does not interact with the eluting proteins. Another reason is that the matrix should not chemically interact with the bound ligands so that the ligand remains unchanged.
5. Which of the following chromatography technique is extremely specific in terms of separation?
a) Ion exchange
b) Thin layer chromatography
c) Affinity chromatography
d) Gel filtration
View Answer
Explanation: In an Affinity chromatography, specific antigen-antibody type interaction happens between the ligand molecule and the protein molecule. Therefore, Affinity chromatography is highly specific.
6. In which of the following matrix, a ligand can be attached?
a) Agarose not treated with Cyanogen Bromide
b) Agarose after reacting with Cyanogen Bromide
c) Cellulose after reacting with Cyanogen Bromide
d) Cellulose not treated with Cyanogen Bromide
View Answer
Explanation: To attach ligand onto an agar matrix, the agar has to be treated with CNBr first. After treating with CNBr, the agar has now side chains to react with a ligand.
7. What type of interaction found between matrix (can be agarose) and ligand in an affinity chromatography?
a) Mixed interaction
b) Ionic
c) Noncovalent
d) Covalent
View Answer
Explanation: The type of interaction found between matrix and ligand in affinity chromatography is covalent interaction. The ligand should be firmly attached to the matrix, therefore, the interaction should be very tight to hold the protein bound ligands. So the interaction is a covalent type of interaction.
8. What is the main problem of CNBr (Cyanogen Bromide) coupled ligand in an affinity chromatography?
a) Its covalent interaction
b) The interactions very loose
c) Steric hindrance
d) The interaction is not stable
View Answer
Explanation: In a CNBr (Cyanogen Bromide) coupled ligand in affinity chromatography, there is no spacer arms present. Therefore, ligand gets attached randomly and congested way which creates steric hindrance which prevents them from bonding with proteins.
9. The main problem of CNBr (Cyanogen Bromide) coupled ligand in affinity chromatography, is removed by following which of the method?
a) Further reacting the agarose matrix with CNBr
b) Introducing a spacer molecule
c) Increasing the matrix volume
d) Acid treatment of Matrix
View Answer
Explanation: In a CNBr (Cyanogen Bromide) coupled ligand in affinity chromatography, ligand gets attached randomly and congested way which creates steric hindrance and prevents them from binding with proteins. Therefore, the introduction of a spacer arms molecule helps to reduce this steric hindrance.
10. Which of the following is very useful to introduce a spacer molecule to the matric in an affinity chromatography?
a) CNBr group
b) Methyl group
c) Ethyl group
d) Epoxy group
View Answer
Explanation: An Epoxy group is used to introduce a spacer molecule to the matric in an affinity chromatography. The epoxy group reacts with spacer arms which helps to introduce it to the matrix molecules.
11. To elute the bound protein from an affinity chromatography column is to add more powerful attractant of the ligand in the second elution buffer.
a) True
b) False
View Answer
Explanation: Yes, this statement is true. A more powerful attractant of the ligand in the second elution buffer is added to replace the already bound protein.
12. In which of the following case, an affinity chromatography cannot be used for a one-step separation?
a) Isolation of an enzyme
b) Isolation of a transporter protein
c) Hormone receptor
d) Separation of a mixture of 3 proteins at a time
View Answer
Explanation: In an Affinity chromatography, a very specific chemical reaction happen which is more like an antigen-antibody reaction. Therefore, only one protein can react with the ligand so the mixture of two proteins can be separated in a single separation process.
13. Which of the following is a correct method to elute a particular protein from the ligand in an affinity chromatography?
a) Elute with a compound which has lower affinity towards the ligand than that protein molecule
b) Elute with a compound which has higher affinity towards the proteins than that protein molecule
c) Elute with a compound which has lower affinity towards the proteins than that protein molecule
d) Elute with a compound which has higher affinity towards the ligand than that protein molecule
View Answer
Explanation: The correct method to elute a protein is to add higher affinity compound (towards ligand) in the elution buffer. This compound will replace protein from the ligand and protein will come to the solution.
14. To isolate insulin receptor, which of the following material can be used as a ligand molecule in an affinity Chromatography?
a) Proinsulin hormone
b) Oxytocin hormone
c) Insulin hormone
d) Preproinsulin hormone
View Answer
Explanation: Affinity chromatography is a specific chromatography where antigen-antibody interaction is followed. Therefore, to isolate insulin receptor, insulin hormone has to be used as a ligand.
15. What is the name of the questioned marked section in the following affinity chromatography picture?
a) Chemical bond
b) Protein molecule
c) Ligand molecule
d) Spacer arm
View Answer
Explanation: The above molecule is called spacer arm. This is introduced to reduce the steric hindrance caused by the overpopulation of the ligand molecules.
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