Paul,
couple things come to mind:
-make sure your substrate is actually a binder and not an (unspecific)
inhibitor (ITC, Thermofluor, Biacore)
-soak longer and at higher concentration. (3mM concentration from 100mM
stock in DMSO for a week or more)
-Is there evidence that your protein needs to undergo substantial
rearrangement prohibited by the crystal
packing? Then you may be out of luck with soaking, try
co-crystallization.
The folks from GSK had a nice paper about some useful techniques for
ligand incorporation probably a worthwhile read: Acta Cryst. (2007).
D63, 72-79 (doi:10.1107/S0907444906047020)
HTH
Carsten
> -----Original Message-----
> From: CCP4 bulletin board [mailto:ccp...@jiscmail.ac.uk] On Behalf Of
> Paul Lindblom
> Sent: Friday, April 09, 2010 5:15 AM
> To: CCP4BB@JISCMAIL.AC.UK
> Subject: [ccp4bb] Does the substrate has access to the active site?
>
> Dear Bulletin Board,
>
> I am trying to soak substrate into crystals of an enzyme, but so far I
> can't see the substrate in the structure. Does anyone knows a program
> to ensure that the entrance to the central cavity is accessibly? I
> mean based on the whole crystal. I already checked the crystal packing
> manually and it seems that the way is free more or less, but I find it
> hard to interpret.
>
> Thanks in advance,
>
> P.
