Virtual Labs

Protein purification by affinity chromatography

1. The full form of Ni-NTA is

a: Nickel-nitrilotrioacetic acid (NTA) Explanation

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b: Nickel-nitrilotriacetic acid (NTA) Explanation

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c: Nickel-triacetic acid (NTA) Explanation

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d: None of the above Explanation

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2. Ni-NTA Agarose is used for the purification of recombinant proteins expressed in

a: Bacteria Explanation

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b: Insect Explanation

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c: Mammalian cells Explanation

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d: All of the above Explanation

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3. Which amino acid in the recombinant protein binds to Nickel, in Ni-NTA affinity chromatography

a: Histidine Explanation

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b: Tryptophan Explanation

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c: Alanine Explanation

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d: None of the above Explanation

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4. The Ni-NTA resins exhibit high affinity and selectivity for 6xHis-tagged recombinant proteins where 6xHis-tag can be fused to its

a: N-terminus Explanation

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b: C-terminus Explanation

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c: Both a and b Explanation

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d: None of the above Explanation

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5. Why does loading/wash buffer contain a low concentration of imidazole

a: To remove 6xHis-tag recombinant protein Explanation

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b: To elute weak binding contaminant proteins Explanation

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c: To increase the binding between Nickel and 6xHis-tag recombinant protein Explanation

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d: None of the above Explanation

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6. Why do we add a higher concentration of imidazole in the elution buffer

a: Because it competes with the recombinant proteins for nickel binding Explanation

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b: Because it acts as an inhibitor for recombinant proteins and nickel binding Explanation

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c: To stabilize the recombinant proteins and nickel binding Explanation

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d: None of the above Explanation

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7. Nickel in Ni-NTA affinity chromatography interacts with how many histidines of a His-tag

a: One Explanation

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b: Six Explanation

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c: Four Explanation

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d: Two Explanation

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8. Nickel makes coordinate bond with the ……………… of histidine

a: Side chain Explanation

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b: Amino group Explanation

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c: Carboxyl group Explanation

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d: All of the above Explanation

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9. Why protein does not bind to the Ni-NTA matrix in an acidic pH range

a: Because it can cause protonation of the histidine side chains and thus prevent the histidine residues from coordinating with nickel Explanation

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b: Because it can cause deprotonation of the histidine side chains and thus prevent the histidine residues from coordinating with nickel Explanation

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c: Because protein aggregates Explanation

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d: None of the above Explanation

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10. What are different methods of elution of recombinant protein from Ni-NTA column other than imidazole

a: Low pH buffe Explanation

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b: By competition with histidine Explanation

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c: Both a and b Explanation

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d: None of the above Explanation

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