The salt in PBS won't prevent binding if it's a strong interaction. If you have a very weak or transient binding it might be something to consider (maybe use 10% PBS), but if the protein binds the peptide in vivo, then the similar salt concentration in PBS shouldn't interrupt that binding. But, again, it depends on your specific protein/peptide.
On Tue, Jun 19, 2018 at 9:35 AM Andreas Stuermer <andreas.t.stuermer@gmail.com> wrote:
So you're saying PBS is probably good? I thought the salts may prevent bindingOn Tue, Jun 19, 2018, 16:30 Bryan Jones <bryanjjones@gmail.com> wrote:While it's always dependant on the specific protein and peptide, a good place to start would be with something physiological-ish, like the extraction buffer or PBS. Depending upon how tightly your protein and peptide bind and how specific you need the interaction to be, you might want to add a bit of detergent to prevent nonspecific interactions. With antibody binding the standard is PBS with 0.1% tween20.--On Tue, Jun 19, 2018, 6:31 AM Andreas "Mega" Stuermer <andreas.t.stuermer@gmail.com> wrote:Will deionized water work, maybe? If I do 1% extraction buffer and 99% dH2O, the pH might still be stable (measure it) and there's not much NaCl to prevent binding of the partners--
On Tuesday, June 19, 2018 at 1:26:52 PM UTC+2, Andreas "Mega" Stuermer wrote:Hi guys,I want a peptide to bind to a protein.After I extracted the protein from the organism (extraction buffer 0.1M Tris-HCl, 0.1M NaCl, pH 8) I will put them into a MWCO centrifuge thing to remove small molecules with 3 kDa cut-off (everything smaller than 30 amino acids flows through the filter and is discarded).What buffer should I put the protein and the peptide in? I know they react under physiological conditions in the cytosol. Maybe I should not use extraction buffer at all and just use the cell pulp plus protease inhibitors?Looking forward to some input
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