How long did you wait before freezing the crystal? Sometimes I have to wait
days before the ligand finds it way into the binding site.
Casper Wilkens
Asst. Prof.
Structural Enzymology & Biorefining
DTU Bioengineering
Technical University of Denmark
[http://www.dtu.dk/%7E/media/DTU_Generelt/Andet/DTU_email_logo_01.gif]
Department of Biotechnology and Biomedicine
Søltofts Plads
Building 224
Room 028
2800 Kgs. Lyngby
[email protected]<mailto:[email protected]>
www.bioengineering.dtu.dk/<http://www.bioengineering.dtu.dk/>
https://www.cazypedia.org/index.php/User:Casper_Wilkens
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________________________________
Fra: CCP4 bulletin board <[email protected]> på vegne af Gourab Basu
Choudhury <[email protected]>
Sendt: 24. april 2021 07:40:20
Til: [email protected]
Emne: Re: [ccp4bb] Co crystalization with less soluble ligand.
I tried sokaing ,high concentration of DMSO is effecting crystal, so I tried
other contidtion where 10%DMSO is there. Got 1.9 A data with protein co
crystalization with 10mM ligand. But ligand occupancy is not visble.
I got the KD value from ITC, with reverse titration. With 10uM ligand in cell
and 150uM protein in syringe. .
Can anybody suggest some views.please feel free to share your experiences.
On Sat, Apr 24, 2021, 10:49 AM Peat, Tom (Manufacturing, Parkville)
<[email protected]> wrote:
Hello Gourab,
DMSO is not the only possible solvent- there are others to try.
I don't want to sound negative, but 40 micromolar is not a very tight binding
compound. It would also depend on how that binding constant was measured- how
did someone get enough in solution to measure that?
Best of luck, tom
Tom Peat, PhD
Proteins Group
Biomedical Program, CSIRO
343 Royal Parade
Parkville, VIC, 3052
+613 9662 7304
+614 57 539 419
[email protected]
________________________________
From: CCP4 bulletin board <[email protected]<mailto:[email protected]>>
on behalf of Gourab Basu Choudhury
<[email protected]<mailto:[email protected]>>
Sent: Saturday, April 24, 2021 12:46 PM
To: [email protected]<mailto:[email protected]>
<[email protected]<mailto:[email protected]>>
Subject: [ccp4bb] Co crystalization with less soluble ligand.
Hello everyone,
I am finding it difficult for getting a ligand density inside the protein as
the ligand is very much insoluble. It's only soluble in 100% DMSO. I tried for
co crystalization. Kd value of the ligand is near 40um. Any suggestion what to
do?
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