Re: [ccp4bb]: NAD-binding Proteins

["Deshmukh N. Gopaul" <gopauld@pasteur.fr>]

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Hi Chris,
Try loading at a low pH followed by a pH gradient to a higher value.
On RedA (Amicon), pH 6.5 (loading) to 8.5 worked well for Nucleoside 
hydrolase from C. fasciculata.

Hope this helps.
Regards,

Deshmukh

Chris Snook wrote:

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>
>I now that this is not a CCP4 question, but I need some help/advice with 
>regard to Affinity Chromatography of an NAD binding protein.
>
>I am using Bio-Rad's AffiBlue gel but am unable to get the protein to bind to 
>the column.  Extensive dialysis, with and without the substrate, fails to 
>elicit binding to the column. The protein is loaded in low ionic strength 
>buffers.
>
>Does anybody have a preference of gel type for  for this type of column? Are 
>there any tricks to aid binding to the column?
>
>Many thanks for any help
>
>Chris
>

-- 
Deshmukh N. Gopaul, Ph.D.
Institut Pasteur
Departement de Biologie Structurale et Chimie
Laboratoire de Biochimie et Biophysique Des Macromolecules
Bat. Metchnikoff, p3011		email: gopauld@pasteur.fr
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