Hi Amala, In addition to Limbini´s suggestions.....
1. You can always do MALS, SLS, Native Page or cross linking experiments to check whether your protein is monodisperse in the solution or not. This is very important step. 2. When you set up crystallization and if you have 40-50% drops precipitated that would be ideal concentration for protein crystallization. 3. As Lumbini said fresh gel filtration passed protein samples behave much better than frozen one. 4. You can also try sitting drop OR under oil method of crystallization too because they follow different crystallization phase diagram. 5. Adding ligands and cofactor would be good idea as well. 6. Add some DTT if you think you have several Cys residues, which may be surface exposed. You can always contact me for more help. Best wishes, ---xxxxx---- With regards Rajesh K. Harijan, Ph.D. Albert Einstein College of Medicine 1300 Morris Park Ave., Bronx, NY 10461 Tel: 718.430.2777 | Fax: 718.430.8565 On Fri, Dec 27, 2019 at 8:30 AM amala mathimaran <[email protected]> wrote: > Dear all, > > Can you suggest me how to get protein crystal??? > > > > I purified my target protein and concentrated to 12mg/ml (pI 6.04). Final > purified protein contains 50mM HEPES pH 8.0 and 50mM NaCl. Then the initial > screening was done using hanging drop method but no crystal. So 2mM NADP > cofactor was added again screen with Hampton (PEG ion, PEG RX, crystal > screen, Index) and Molecular dimensions conditions etc. I got precipitate > like formation the image was attached below. From this formation what I can > do… mean while I increase the protein concentration and did screening for > that selected conditions again I got same kind of formations. I am new to > protein crystallography kindly suggesting me. And how much concentration is > suitable for protein crystallization?? How to find which concentration is > enough for our target protein crystallization?? Thanks in advance > > ------------------------------ > > To unsubscribe from the CCP4BB list, click the following link: > https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB&A=1 > ######################################################################## To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB&A=1
