Rongjin, Yes, there most definitely are numerous examples of phosphorylated, glycosylated, lipidilated, etc. etc. proteins crystallized. Since you are asking specifically about phosphorylation here are my two cents:
a. over one half of all heterogenously expressed kinases are phosphorylated b. about half of those phosphorylate themselves, sometimes to a great degree. The most common example of the latter is PKA which under the right expression conditions can be phosphorlyated on over 20 sites c. there are several major techniques people have employed to crystallize phosphorylated proteins. Nearly all of them focus on obtaining homogenous species. d. some proteins can crystallize even when heterogenously phosphorylated, although the crystals are not always any good. Yet others absolutely require homogeneity. Several important notes: hyperphosphorylating kinases can be very toxic to their expression hosts. Depending on the host, some kinases may be super-active, silent, or somewhere in between. Some kinases become insoluble when they're not phosphorylated, the opposite also can happen. Methods to control phosphorylation include: expression host choice coexpression with other kinase(s) coexpression with phosphatase(s) in vitro enzymatic treatment (phosphorylation or dephosphorylation) mutagenesis of key residues - either of the phosphorlyation sites or of the active site coexpression with regulatory proteins expression in the presence of inhibitors modulation of growth conditions (which is how PKA was done) etc. Artem > Dear all, > > Below is a question my friend asked me, but I have never worked on > phosphorylated proteins. > Has anyone worked on crystallizing phosphorylated proteins and can you > comment on it? > > Thanks > > Rongjin Guan > ---------------------------------------------------------------------- > I would like to ask how feasible it is to crystalize protein containing > post-translational modification, such as phosphorylation. To my limited > knowledge, I think the heterogenicity of modified and unmodified proteins > causes the major obstacle in the crystal structure. Is there any specific > method to crystalized modified proteins (like phosphorylated proteins ) > these days? >
