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[ccp4bb] degradation of protein durring freez thaw
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CCP4bb <-- 1999 <-- November 1999 <-- 30 November 1999
Previous message:
Subject: precipitation of protein in elution buffer
From: peter hudson peter {- dot -} hudson {- dot -} peter {- at -} GMAIL {- dot -} COM
Date: 2010-04-22		Next message:
Subject: Re: precipitation of protein in elution buffer
From: Kornelius Zeth kornelius {- dot -} zeth {- at -} TUEBINGEN {- dot -} MPG {- dot -} DE
Date: 2010-04-22


Subject: degradation of protein durring freez thaw
From: Jhon Thomas jhon1 {- dot -} thomas {- at -} GMAIL {- dot -} COM
Date: 2010-04-22

Hello BB

I apolozize an off topic query.

I am working with small proetin-protein complex of 24kDa. I purify this
N-terminal His-tagged complex through tylon resin in 20mM of Tris pH-8.0,
0.3M NaCl . After purification this protein complex are dialysed in 20mM
tris pH=8.0.I am able to purify enough amount of protein for
crystallisation, which can be concentrated upto 10mg per ml. Then i check
the dgradation on the polyacrylamide gel after concentration of the protein.
I donot see any degdation protein band on the gel. I store the protein at
-80 in aliquotes of 100ul immedaitely after concentration in same buffer.
protein concentartion are done at 4 degree by centrifugation. Next day
before setting up the trays for crystallisation screening, protein solution
concentration check is being done. it turns out that this complex has
degraded and concentration is only 1-2 mg per ml. i would appreciate the
suggestions to prevent the degradation of complex or How should i make it
more stable? so, that i can proceed for the crystallisation. I would really
appreciate the suggestions.


Thanks in advance

Thomas

CCP4bb navigation

CCP4bb <-- 1999 <-- November 1999 <-- 30 November 1999
Previous message:
Subject: precipitation of protein in elution buffer
From: peter hudson peter {- dot -} hudson {- dot -} peter {- at -} GMAIL {- dot -} COM
Date: 2010-04-22		Next message:
Subject: Re: precipitation of protein in elution buffer
From: Kornelius Zeth kornelius {- dot -} zeth {- at -} TUEBINGEN {- dot -} MPG {- dot -} DE
Date: 2010-04-22
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