omley, Matthew" <[EMAIL PROTECTED]>
To: CCP4BB@JISCMAIL.AC.UK
Sent: Thursday, April 24, 2008 7:35:46 AM
Subject: [ccp4bb] Setting drops with proteins that are hard to concentrate
Setting drops with proteins that are hard to concentrate Dear All,
I have a 50kDa protein that is soluble and mo
Dear Matt,
make sure that you don't have slow or even fast formation of unspecific
intermolecular disulfides by simple checks at several time points on
SDS-PAGE using sample buffer without DTT. Check the content of free
thiols and disulfides of your protein over a period of time. If you need
can't you try different volumes: i.e. 360 nl protein solution in very little
buffer and salt + 40 nl precipitant. Should give roughly 10 mg/ml after
equilibration.
Best wishes
Kornelius
On Thu, 24 Apr 2008 12:42:42 +0100
Roberto Steiner <[EMAIL PROTECTED]> wrote:
> I have a similar problem Ma
I have a similar problem Matthew.
Just got some NVoy from Novexin in which some claim can help if
hydrophobic patches are the main problem. Will see.
Regards,
Roberto
On 24 Apr 2008, at 12:35, Bottomley, Matthew wrote:
Dear All,
I have a 50kDa protein that is soluble and monodisperse at up
Dear All,
I have a 50kDa protein that is soluble and monodisperse at up to approx
1mg/ml (after Ni-affinity and size-exclusion chromatography).
However, it aggregates (probably both via disulphides and via
'sticky/hydrophobic patches') when I concentrate it towards 2-3mg/ml,
even in the presence