[ccp4bb] Cannot running NTA to purify the protein having His-tag?

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CCP4bb <-- 1999 <-- November 1999 <-- 30 November 1999

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Subject: Cannot running NTA to purify the protein having His-tag?
From: ngoductri {- at -} GMAIL {- dot -} COM
Date: 2007-02-28

Dear CCP4 users,

I'm purifying a kind of protease having His-tag. The protein is expressed in
insect cells and broken by sonication.
I used NTA resin to purify this protein.
Buffer A is 50mM phosphate buffer pH 7.5 and 300mM NaCl. Buffer B is 50mM
phosphate buffer pH 7.5, 300 mM NaCl and 300 mM Imidazole.
However, all proteins cannot bind to NTA resin. My protein is eluted in
Flow-through. I also check the NTA resin with the control His-tag. The
western blot also shows that my protein has His-tag.

Do you have any ideas about my problem? I'm really appreciate all of your
advices how to solve this. Thank you very much!

My best regards,
TriNgo
Sungkyunkwan University

CCP4bb navigation

CCP4bb <-- 1999 <-- November 1999 <-- 30 November 1999

Previous message:
Subject: questions on SF likelihood
From: pam52 {- at -} CORNELL {- dot -} EDU
Date: 2007-02-28

Next message:
Subject: Re: Oxford Xcalibur Vs Rigaku micromax
From: rangel {- at -} VT {- dot -} EDU
Date: 2007-02-28