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#protein-protein interaction

2 public questions tagged with this topic.

Electrophoresis

Which method is used for detecting protein-protein interaction?

Protein-protein interactions are commonly examined using co-immunoprecipitation followed by electrophoretic detection. After precipitating bait protein with specific antibody bound to beads, associated partners co-precipitate, are eluted, resolved by SDS-PAGE, transferred to membrane and identified by Western blotting with antibodies against suspected interactors. Polymerase chain reaction amplifies DNA, Northern blot detects RNA transcripts, and electrophoretic mobility shift assay detects DNA-protein complexes, not protein-protein binding. Coupled with far-Western, pull-down or crosslinking

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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DNA-Protein Interaction analysis

In a protein-protein interaction assay, Co-IP is used to:

Co-immunoprecipitation is standard biochemical technique to detect protein-protein interactions under native conditions. Cellular lysates prepared in non-denaturing buffer are incubated with antibody specific to bait protein immobilized on Protein A or G agarose beads. Interacting prey proteins co-precipitate with bait, are eluted and detected by Western blotting. Successful co-precipitation indicates physiological association within complex, signalosome or regulatory scaffold. It does not detect direct DNA binding or amplify RNA. Therefore it specifically precipitates interacting proteins, di

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.

View Details