I want to determine which proteins in a cell lysate bind to a short alpha helical synthetic peptide. Is there a good functional group (like an azide) that I could add to the peptide that would cross link the peptide to any interacting protein? Some kind of click-it chemistry would be best, or a photo-activated cross link.
There are loads of chemical crosslinkers that will link your peptide to any interacting protein. You find them in all forms: There are reducible crosslinkers, meaning they can be unlinked again; there are different lengths in the linker; you find soluble and non-soluble linkers, in case you care about membrane proteins. I worked with DTSSP, DSP, and BS3 so far. It is basically click-it chemistry ;) Just avoid Tris buffers, as they are used for quenching. I found it helpful to try different concentrations at first, but I ended up with 0.5mM and 2mM, 1h on ice, quench with 30-150mM Tris.
One question though: How do you want to identify your linked protein in the end? Do you have an antibody against your synthetic peptide? In that case you could try a pull-down. That would also be possible after the crosslinking your protein. If you don't have an antibody, you might actually look for a way to link your peptide to a matrix. That is something I never tried, but I am sure there is someone here ;)
Diazirines are good photo-activated covalent cosslinkers. They are small (less likely to disrupt the native binding you're looking for), and non-specific (they will react with a large variety of functional groups). Some are commercially available as the NHS ester for coupling onto your peptide. http://www.piercenet.com/product/nhs-diazirine
There are lots. Diazirines are great. Some others to consider are benzophenone, and aryl azides. You could also consider biotinylating your peptide then performing a simple streptavidin pull-down. If the interaction is low affinity you could try an non-specific gluteraldehyde x-linking followed by streptavidin pull-down. Lots of options.
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