Re: [ccp4bb] Thermofluor experiment

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Subject: beamtime proposal deadline for BM14/ESRF -April/May run 2008 Deadline 20-2-2008
From: "Martin A {- dot -} Walsh" walsh {- at -} ESRF {- dot -} FR
Date: 2008-02-13

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Subject: Re: xprep vs shelxc
From: "George M {- dot -} Sheldrick" gsheldr {- at -} SHELX {- dot -} UNI-AC {- dot -} GWDG {- dot -} DE
Date: 2008-02-13

Subject: Re: Thermofluor experiment
From: Andreas_Förster docandreas {- at -} GMAIL {- dot -} COM
Date: 2008-02-13

Dear Kornelius,

I found the idea of doing Thermofluor on membrane proteins really
intriguing - for identifying the best buffer and detergent, secondary
detergents, even for checking crystallization drops that stayed clear.
(This latter experiment should theoretically be possible with large
drops, even though you'd be working very near the limits claimed in the
publications.)

In the end, I didn't find the method too useful because of noise issues
due to detergent and exposed hydrophobic portions of the protein. I
always felt that, in order to get reasonable signal, I'd have to use
protein at unreasonable concentrations.

The method works much better for soluble protein, though I can't tell
you a success story where it led to crystallization that seemed
impossible before.

Andreas

Kornelius Zeth wrote:
> Dear all,
>
> a question very related to the discussion before. I have been reading the papers about the thermofluor experiment with great interest.
>
> I wonder what people think about the underlying principles/ideas and the success that the method yielded in their own labs for crystallization or related purposes?
>
> Has anybody used this method with membrane proteins in order to find out the stability of the protein in the presence of a second detergent?
>
> Is the method limited to this certain dye (sypro orange)?
>
> Have a nice day
>
> Kornelius
>
> P.S.: I will make a summary of all opinions.
>
> ----------------------------------------------
> Kornelius Zeth
> Max Planck Institute for Developmental Biology
> Dept. Protein Evolution
> Spemannstr. 35
> 72076 Tuebingen, Germany
> kornelius.zeth@tuebingen.mpg.de
> Tel -49 7071 601 323
> Fax -49 7071 601 349
>

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CCP4bb <-- 2008 <-- February 2008 <-- 13 February 2008

Previous message:
Subject: beamtime proposal deadline for BM14/ESRF -April/May run 2008 Deadline 20-2-2008
From: "Martin A {- dot -} Walsh" walsh {- at -} ESRF {- dot -} FR
Date: 2008-02-13

Next message:
Subject: Re: xprep vs shelxc
From: "George M {- dot -} Sheldrick" gsheldr {- at -} SHELX {- dot -} UNI-AC {- dot -} GWDG {- dot -} DE
Date: 2008-02-13