I generally add Protease inhibitor cocktail in cell lysate (calbiochem set II) which have almost all required protease inhibitors, and each time I got the crystal, without any effect due to this. You should be also careful if your protein itself is a protease as many times people find these inhibitors bound in the active site, so optimization of concentration PIC is also important. You can try EDTA and PMSF together once, as most of the time it works. But better you add cocktail of many. If your protein is an ATPase then try to avoid EDTA into it as it interfere its activity.
Good luck Prem On Tue, Dec 12, 2017 at 4:21 PM, Liuqing Chen <[email protected]> wrote: > HI! > I purified a protein in last year, and 1.jpg showed the sds-page of this > protein, and it can grow crystals in one condition. But i can't repeated > it , i have purifed many times, all failed to grow crystal. I can't get > the phase by metal soak or MR. the 2.jpg showed the same protein i > purified in last week, then i treated it with different concentration of > chymotrypsin within 30min, the result showed in 3.jpg, it showed my > protein is easy to be degraded. > My question is should i add protease inhibitor when purify my protein > (expressed in E.coli Bl21 de3), PMSF? or others? the inhibitor is > expensive. > thanks in advance. > Liuqing chen >
