Did anyone suggest adding any known ligand? If your protein happens to
bind some compound/peptide/DNA/RNA/whatever, including that other partner
could dramatically change it's solution properties and be an easy fix for
your handling/formulation issues.
Good luck!
On Fri, Jul 14, 2017 at 6:33 AM,
Hi Chris,
In your Ni-NTA buffer, the total [Na+] could be greater than 530mM after
titration. Likely, your protein likes higher concentration salt for surface
charge stabilization. Adding Glycerol may further modify the charge
distribution and make the protein happy in the solution. For further
HEPES pH 7.5, 10% glycerol)
works. It should work well with crystallization.
Cheers,
Chun
From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of Chris Fage
Sent: Friday, July 14, 2017 3:33 AM
To: CCP4BB@JISCMAIL.AC.UK
Subject: Re: [ccp4bb] Protein rapidly precipitates when off ice
To: CCP4BB@JISCMAIL.AC.UK
Subject: Re: [ccp4bb] Protein rapidly precipitates when off ice
Dear All,
Thank you for the many suggestions. After sending my first message to the BB, I
tried exchanging the sample into buffer containing 5 mM EDTA and also into
buffer at pH 9.0 (using BICINE). Neither
Dear All,
Thank you for the many suggestions. After sending my first message to the
BB, I tried exchanging the sample into buffer containing 5 mM EDTA and also
into buffer at pH 9.0 (using BICINE). Neither of these appear to have
helped the instability--precipitation still occurred within ~1 min
ght try try solubility screening as
> outlined...
>
> http://scripts.iucr.org/cgi-bin/paper?dz5020
>
> HTH,
>
> Dave
>
> --
> Dr David C Briggs
> Hohenester Lab
> Department of Life Sciences
> Imperial College London
> UK
> http://about.me/david_brig
ndon
>> UK
>> http://about.me/david_briggs
>>
>>
>> From: CCP4 bulletin board <CCP4BB@JISCMAIL.AC.UK> on behalf of Chris Fage
>> <fage...@gmail.com>
>> Sent: Thursday, July 13, 2017 11:40:34 PM
>> To: CCP4B
>
>
> From: CCP4 bulletin board <CCP4BB@JISCMAIL.AC.UK> on behalf of Chris Fage
> <fage...@gmail.com>
> Sent: Thursday, July 13, 2017 11:40:34 PM
> To: CCP4BB@JISCMAIL.AC.UK
> Subject: [ccp4bb] Protein rapidly precipitates when off ice
>
&g
ege London
UK
http://about.me/david_briggs
From: CCP4 bulletin board <CCP4BB@JISCMAIL.AC.UK> on behalf of Chris Fage
<fage...@gmail.com>
Sent: Thursday, July 13, 2017 11:40:34 PM
To: CCP4BB@JISCMAIL.AC.UK
Subject: [ccp4bb] Protein rapidly precipit
Dear CCP4BB Community,
This week, I purified a nicely overexpressing protein by Ni-NTA followed by
gel filtration. In a 4 C centrifuge, I concentrated my gel filtration
fractions to ~1 mL, transferred the spin filter to ice, and then collected
2 uL for measurement on the Nanodrop. Sadly, the
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