This set of Protein Engineering Multiple Choice Questions & Answers (MCQs) focuses on “Applications – Affinity Purification – 1”.
1. Which of the following is not associated with obtaining a pure protein?
a) Precipitation
b) Adsorption/desorption
c) Column chromatography
d) Solubilization
View Answer
Explanation: Solubilization is not associated with obtaining a pure protein. Traditionally, a long series of precipitation, adsorption/desorption, and column chromatography steps have to be carried out in order to obtain a pure protein.
2. Which of the following affinity tag is an 11 amino acid residue peptide that can be a target of monoclonal antibody?
a) Flag-peptide
b) S-tag
c) His-tag
d) Myc tag
View Answer
Explanation: Myc tag is an 11 amino acid residue peptide that can be a target of monoclonal antibody. This tag requires physiological conditions and low pH for washing/elution. Flag-peptide, S-tag, or His-tag are not affinity tags that are 11 amino acid residue peptides which can be a target of monoclonal antibody.
3. Which of the following is a short sequence of amino acids that is recognized as an epitope by a monoclonal antibody and is appended to a recombinant protein on the genetic level?
a) Antigenic peptide
b) Peptide tag
c) Sequence tag
d) Affinity tag
View Answer
Explanation: Affinity tag is a short sequence of amino acids that is recognized as an epitope by a monoclonal antibody and is appended to a recombinant protein on the genetic level. Thus, the molecular recognition property of the peptide is transferred to the new protein context, and, given the proper choice of the amino acid sequence, it is likely not to interfere with the protein’s biological activity.
4. Which of the following affinity tag does not require harsh conditions for elution of the tagged protein from a column with the immobilized antibody?
a) Flag-peptide
b) His-tag
c) Calmodulin-binding peptide (CBP)
d) Strep-tag
View Answer
Explanation: Strep-tag is an affinity tag that does not require harsh conditions for elution of the tagged protein from a column with the immobilized antibody. Flag-peptide, His-tag, and Calmodulin-binding peptide (CBP) require harsh conditions for elution of the tagged protein from a column with the immobilized antibody. Therefore, these affinity tags cannot be readily employed for purification of the protein.
5. Which of the following is a ligand of streptavidin?
a) Oxalic acid
b) Pantothenic acid
c) Zinc ion
d) Biotin
View Answer
Explanation: Biotin is a ligand of streptavidin/avidin. This ligand is also known as vitamin H/vitamin B7. The binding of biotin to streptavidin is one of the strongest non-covalent interactions known in nature. Oxalic acid, pantothenic acid, or zinc ion are not ligands of streptavidin.
6. Which of the following affinity tag is an 8 amino acid residue peptide that can be a target of monoclonal antibody?
a) S-tag
b) Myc tag
c) His-tag
d) Flag-peptide
View Answer
Explanation: Flag-peptide is an 8 amino acid residue peptide that can be a target of monoclonal antibody. This tag requires physiological conditions and chelator compounds for washing/elution. S-tag, Myc tag, or His-tag is not affinity tags with 8 amino acid residues, that can be a target of monoclonal antibody.
7. Which of the following technique made it possible to shorten the laborious procedure of protein purification?
a) Precipitation
b) Column chromatography
c) Adsorption/desorption
d) Affinity chromatography
View Answer
Explanation: Affinity chromatography made it possible to shorten the laborious procedure of protein purification. Precipitation, column chromatography, and adsorption/desorption have no role in the shortening of the laborious procedure of protein purification. A suitable combination of ligand, matrix, and elution conditions has first to be optimized in each case.
8. Which of the following affinity tag is a 6 amino acid residue peptide which can be immobilized by transition metal ions?
a) S-tag
b) Myc tag
c) Flag peptide
d) His-tag
View Answer
Explanation: His-tag is a 6 amino acid residue peptide that can be immobilized by transition metal ions. It typically consists of at least six histidine residues. This tag requires high salt/low pH conditions, chelator compounds, and imidazole for washing/elution. Myc tag, flag peptide, and S-tag are not 6 amino acid residue peptide that can be immobilized by transition metal ions.
9. Which of the following is not true for streptavidin?
a) It is extremely stable against proteases
b) It is extremely stable against elevated temperatures
c) It is devoid of sulfur-containing amino acids
d) It is highly glycosylated
View Answer
Explanation: Streptavidin is extremely stable against proteases, elevated temperatures, and detergents. It is devoid of sulfur-containing amino acids, glycosylation, and slightly acidic. Hence the statement ‘It is highly glycosylated’ is not true for streptavidin.
10. Which of the following conserved consensus sequence is considered crucial for streptavidin-biotin binding?
a) Ala-Gln-Gly
b) Pro-His-Gln
c) Gly-His-Gln
d) His-Pro-Gln
View Answer
Explanation: His-Pro-Gln is a conserved consensus sequence on streptavidin that is considered crucial for streptavidin-biotin binding. Ala-Gln-Gly, Pro-His-Gln, and Gly-His-Gln are not crucial for streptavidin-biotin binding. The avidin-biotin complex is the strongest known non-covalent interaction between a protein and a ligand.
11. Strep-tag is only used for the purification of recombinant proteins.
a) True
b) False
View Answer
Explanation: The above statement is false. The strep-tag has been successfully used for the detection and the purification of recombinant proteins in a variety of examples. The fact that the streptavidin affinity chromatography can be performed under gentle physiological conditions is a particular advantage of this method.
12. Which of the following affinity tag is a 15 amino acid residue peptide which can bind to S-fragment of RNaseA?
a) His-tag
b) Myc-tag
c) Calmodulin-binding peptide (CBP)
d) S-tag
View Answer
Explanation: S-tag is a 15 amino acid residue peptide that can bind to S-fragment of RNaseA. This tag requires physiological/thrombin cleavage conditions for washing/elution. His-tag, Myc-tag, and Calmodulin-binding peptide (CBP) are not 15 amino acid residue peptides that can bind to S-fragment of RNaseA.
13. Strep-tag affinity chromatography requires high salt concentrations, extreme pH, chelator compounds for washing or elution.
a) True
b) False
View Answer
Explanation: The above statement is false. Strep-tag affinity chromatography does not require high salt concentrations, extreme pH, elution gradients, or chelator compounds for washing or elution. It requires gentle conditions.
14. Most of the interactions between the strep-tag and streptavidin are of non-polar nature.
a) True
b) False
View Answer
Explanation: The above statement is false. Most of the interactions between the strep-tag and streptavidin are of polar nature. These electrostatic interactions are possible because of the characteristic conformational flexibility of the two Gly residues at the end of the strep-tag.
15. Which of the following affinity tag is a 26 amino acid residue peptide which can be bound to calmodulin?
a) Myc-tag
b) His-tag
c) S-tag
d) Calmodulin-binding peptide (CBP)
View Answer
Explanation: Calmodulin-binding peptide (CBP) is a 26 amino acid residue peptide that can be bound to calmodulin. This tag requires high salt/chelator compounds for washing/elution. His-tag, Myc-tag, and S-tag are not 26 amino acid residue peptide that can be bound to calmodulin.
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