[ccp4bb] Antwort: [ccp4bb] removal of sulfate ion from the active site

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CCP4bb <-- 1999 <-- November 1999 <-- 30 November 1999

Previous message:
Subject: Re: removal of sulfate ion from the active site
From: Dirk Kostrewa dirk {- dot -} kostrewa {- at -} PSI {- dot -} CH
Date: 2007-05-25

Next message:
Subject: Re: Antwort: removal of sulfate ion from the active site
From: Frank von Delft frank {- dot -} vondelft {- at -} SGC {- dot -} OX {- dot -} AC {- dot -} UK
Date: 2007-05-25

Subject: Antwort: removal of sulfate ion from the active site
From: Djordje {- dot -} Musil {- at -} MERCK {- dot -} DE Djordje {- dot -} Musil {- at -} MERCK {- dot -} DE
Date: 2007-05-25

Hi Simon,

One solution would be to do as Herman has suggested. But if your crystals
do not survive for any reason (sulfate ion may be needed for stuctural
stability of the crystal) you may try to cross-link your crystals using
glutaraldehyde.

A few more things to think of: what is the affinity of your ligand to the
protein, compared to the affinity of the sulfate ion? Do they compete for
the same site? Then you need to compete the suflate out of the
structure.The maximal concetration of the ligand in solution is determined
by its solubility. Since you have a phosphate group in your ligand I
supposse that the solubility is higher than 10 mM. Is it? How long have you
soaked, i.e. was it enough time to reach the equilibrium?

Best regards,

Djordje

______________________________________________

Yue Li
U> An
Gesendet von: CCP4BB@JISCMAIL.AC.UK
CCP4 bulletin Kopie
board
AC.UK> [ccp4bb] removal of sulfate ion
from the active site

24.05.2007 20:25

Bitte antworten
an
Yue Li
U>

Hi all,

I have crystals of the apo enzyme growing from 1.7M ammonium sulfate
condition. After soaking with 10mM ligand (substrate analog) which has a
phosphate group, the ligand did not enter the active site of the enzyme
because of the competition of sulfate ion and phosphate group. So I am
wondering if anyone knows how to get rid of ammonium sulfate from
crystal before soaking it with the ligand solution? Thanks a lot.

Simon

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CCP4bb navigation

CCP4bb <-- 1999 <-- November 1999 <-- 30 November 1999

Previous message:
Subject: Re: removal of sulfate ion from the active site
From: Dirk Kostrewa dirk {- dot -} kostrewa {- at -} PSI {- dot -} CH
Date: 2007-05-25