Has anyone ever seen an increase in protein-protein interaction after using a peptide, which was ideally supposed to block or reduce interaction? The peptide mimics one protein's active site, so theoretically it should inhibit the binding between 2 proteins, but I have consistently been seeing increased interaction. Can someone suggest what could be the reason or mechanism?
Thank you
Hi,
Just to make sure I understand you correctly: the peptide mimicks the active site of the protein, or the domain targeted by the active site?
You expect to inhibit the interaction between protein A and its target protein B by adding the peptide?
Do you see more interaction between protein A and B after adding the peptide, or (also) more self-association of protein A (and/or self-association of protein B)?
Could it be that adding the peptide changes the conformation of protein A, resulting in non-specific protein interactions?
Hi Eef,
Thank you for replying.
You expect to inhibit the interaction between protein A and its target protein B by adding the peptide? Yes
Do you see more interaction between protein A and B after adding the peptide? Yes
Proteins A and B are known to interact with each other. We have designed a peptide that mimics the binding site of protein A where protein B binds and want to check if addition of peptide to this equation blocks the pulldown of protein B by A. We expected that there should have been reduced pulldown of B by A in the presence of peptide (because the site of interaction is blocked). However, the pulldown is significantly higher when peptide is added. Another peptide (size and sequence are very similar and same amount) causes no change in pulldown so the result seen by the peptide in question seems specific. I am trying to understand what could be the reason/mechanism for this.
Any insights would be helpful.
Thank you
John Carter Thank you for replying. It could be one possibility. If I can bother you more, I still would like your expertise in understanding few more things.
So if the peptide mimics the active site of protein A, can it still allosterically bind to protein A or B and enhance the interaction? There's also a protein C in the equation and presence of the peptide increases pulldown of all 3.
Under normal conditions, proteins A, B, and C can pulldown each other.
No peptide scenario: All 3 can still get pulled down but presence of A affects the pulldown of C by B.
Peptide added: Increased pulldown of all 3 compared with levels seen when no peptide was added.
What is it that the peptide is doing which is causing an increased pulldown is something I cannot understand.
I'm guessing A and B normally bind cooperatively, and the peptide can substitute for B in producing the cooperative effect.
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