Hi Francesca, since your protein is expressed in eucarytic cells, glycosylation might be an issue. Glycosylation usually enhances the solubility and can influence the crystallisation negativly because of high flexibility. There are commercially available deglycosylation kits.
You can try methylation of primary amines of your protein. For this purpose commercially available kits are existing too. My experience was a drastic reduction of solubility. Good Luck Max -------- Original message -------- From: "Mattiroli,Francesca" <[email protected]> Date:17/02/2015 04:34 (GMT+00:00) To: [email protected] Cc: Subject: [ccp4bb] how to reduce protein solubility Hi all, I am struggling with a protein complex that is too soluble. I have reached about 20 mg/ml but I still observe very little precipitation (clear drops in 90-95% of the tested conditions). The proteins are expressed in insect cells and going to higher concentration is not easily achievable. I have tried different buffer conditions (salt concentration and pH) and I am testing temperatures. I am at a loss with what to try next. Do you think PTMs (phosphorylation, acetylation) might be causing this? Any input on how to decrease solubility? Thank you very much in advance, Francesca ------------------------------------------------------------------------------------------------ ------------------------------------------------------------------------------------------------ Forschungszentrum Juelich GmbH 52425 Juelich Sitz der Gesellschaft: Juelich Eingetragen im Handelsregister des Amtsgerichts Dueren Nr. HR B 3498 Vorsitzender des Aufsichtsrats: MinDir Dr. Karl Eugen Huthmacher Geschaeftsfuehrung: Prof. Dr.-Ing. Wolfgang Marquardt (Vorsitzender), Karsten Beneke (stellv. Vorsitzender), Prof. Dr.-Ing. Harald Bolt, Prof. Dr. Sebastian M. Schmidt ------------------------------------------------------------------------------------------------ ------------------------------------------------------------------------------------------------
