I would use a low affinity metal ion binding buffer like MOPS, HEPES, or
MES. The "Good" buffers all have fairly low metal-ion affinity.
Phosphates will be problematic because of magnesium phosphate formation.
Cheers,
_______________________________________
Roger S. Rowlett
Gordon & Dorothy Kline Professor
Department of Chemistry
Colgate University
13 Oak Drive
Hamilton, NY 13346
tel: (315)-228-7245
ofc: (315)-228-7395
fax: (315)-228-7935
email: rrowl...@colgate.edu
On 3/20/2013 2:59 AM, Harsh Bansia wrote:
Sorry for a simple and non-CCP4 question.
I have determined the structures of three different mutants of a
thermostable protein by X-ray crystallography method. I feel that Mg2+
has a role in protein stability.
So I want to perform a thermal denaturation study by CD spectroscopy
both in presence and absence of Mg2+ ion.
In this regards, what should be the suitable buffer for CD studies?
May I use PBS buffer ? Since phosphate sequester divalent cations like
Mg2+. Is it advisable to use PBS buffer. If so, what is maximum
concentration of Mg2+ ion that can be used say e.g. 5 mM? My protein
was in Tris buffer and lyophilized and have theoretical pI =4.56 and
maximum activity at pH 8.4.
Thanking you in advances,
harsh
