I am trying to crystallize a protein that I am phosphorylating in-vitro. There
is no way that I can purify the phosphorylated protein from unphosphorylated
one. I tried Ion exchange and gel filtration for separating them and they are
not working.
Mass spec result shows the phosphorylation at
Thank you Prof Lewis.
Its serine residue which is getting phosphorylated. the pI of my protein is
around 8.2. So native gel ( around pH 8.3) is also not working for me. I tried
ion exchange at pH 7.0, 7.5 and 8.5 but my protein is either coming in flow
through or very poorly binding to Q and SP
of Srivastava,
Dhiraj [dhiraj-srivast...@uiowa.edu]
Sent: Monday, January 19, 2015 5:39 PM
To: CCP4BB@JISCMAIL.AC.UK
Subject: Re: [ccp4bb] phosphoprotein crystallization
Thank you Prof Lewis.
Its serine residue which is getting phosphorylated. the pI of my protein is
around 8.2. So native gel
-srivast...@uiowa.edu]
Sent: Monday, January 19, 2015 5:39 PM
To: CCP4BB@JISCMAIL.AC.UK
Subject: Re: [ccp4bb] phosphoprotein crystallization
Thank you Prof Lewis.
Its serine residue which is getting phosphorylated. the pI of my protein is
around 8.2. So native gel ( around pH 8.3) is also
an effective modification to your protein preparations would be to add
phosphatase inhibitor tablets during all
stages of purification. Pierce sell a combination protease and PI tablet
that is effective and economical to be added during protein preps. Also you
invitro buffers should have
