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#Y2H

2 public questions tagged with this topic.

Protein-Protein Interaction

Which of the following is not a protein interaction method?

Techniques to map physical protein associations include co-immunoprecipitation that isolates native complexes via specific antibodies, fluorescence resonance energy transfer that detects proximity under 10 nm in live cells, phage display linking phenotype to genotype, yeast two-hybrid, surface plasmon resonance, and affinity pull-downs. SDS-PAGE is a fundamental analytical separation method that denatures proteins with sodium dodecyl sulfate and resolves them by molecular weight under electric field. It provides size information but alone does not report interaction without further methods like far-western or immunoblotting for detection.

Ref: NCERT Biology Class XII Principles on Klenow fill-in labeling, Lehninger Chapter 9 DNA cloning techniques, and Molecular Cloning by Sambrook Chapter 10 documenting end-labeling of cohesive termini.

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Protein-Protein Interaction

In a research problem, a student finds proteins A-D with gel filtration, A-C with Co-IP, and B with Y2H. What is the bes

Gel filtration separates native protein complexes by hydrodynamic size; co-elution suggests residence in same large complex but does not distinguish direct versus bridged association. Co-immunoprecipitation from lysates preserves multiprotein assemblies, confirming co-complex membership. Yeast two-hybrid tests binary, direct physical interaction inside nucleus because reporter activation requires intimate contact between bait and prey fusions independent of other yeast proteins. When protein B uniquely appears positive in two-hybrid while others co-purify only, it indicates B is the direct interactor contacting bait, while A, C, D likely associate via B.

Ref: NCERT Biology Class XII Principles on Klenow fill-in labeling, Lehninger Chapter 9 DNA cloning techniques, and Molecular Cloning by Sambrook Chapter 10 documenting end-labeling of cohesive termini.

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