In addition to the previous suggestions, if you have a metal-binding protein, beware of acidic compounds chelating the metal and stripping it out of the protein, as this often lead to the effect you observed as well.
Isaac On 5 Feb 2015 13:45, "Monica Mittal" <[email protected]> wrote: > Hi all, > I am working to crystallize a protein-ligand complex. I did a preliminary > melting curve analysis for the protein in the absence and presence of 2 > ligands (dissolved in protein buffer). I kept the other controls as buffer > an a known standard to confirm instrument performance. All expts done in > triplicates. > Now the results are like : Tm of protein alone is 56 deg, Tm in the > presence of Lig1 conc. of 0.05mM, 0.1mM, 1.0mM and 10.0mM is 56.2, 56.1, > 56.0 and 53.5 respectively !! Tm in the presence of Lig 2 conc. of 0.05mM, > 0.1mM, 1.0mM, 4.0mM and 10.0mM is 56.2, 56.1, 54.9, 52.3 and 50.6 > respectively !! > Although the effective delta Tm for both is different at higher > concentration, but both are kind of making protein less stable. So i was > wondering, will it be difficult to co-crystallize them !! Any suggestions > in this regard are highly appreciated !! > > Thanks > Monica >
