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Re: [ccp4bb]: substrate soaking into crystals
Yes, the sulfate is probably competing with ATP for the binding to the
phoshate groups binding sites. Actually, i would guess that the map shows clearly a sulfate
ion bound in the nucleotide-binding site. You may try to do soaking experiments with
a really high concentration of ATP (about 10, 20, 50 mM ...). May be you can also
decrease the sulfate concentration in your soaking solution (to 0.1 M or 0.05 M).
The other alternative is to replace the ammonium sulfate by something else in the a
soaking solution (NH4Cl or NaCl).
If you don't see the sulfate ion bound in the nucleotide-binding site, then the problem
is different: the nucleotide-binding-site is not accessible or the affinity for ATP is very low !
May be you have need to add Mg2+ (if your protein use Mg2+ to hydrolyse ATP, in this case of course
you will get ATP-hydrolysis, ...). Then you could try with AMPPNP-Mg2+ or AMPPCP-Mg2+ !!!
Have you tried to soak with ADP-Mg2+ ?
Good luck !
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