[RASMB] sedfit vs SOMO vs structure

[Arthur Rowe]

Many apologies, Ewa, you did indeed write it like that. And of course I  
agree (and wrote) that MALLS is certainly one way to settle the issue.

Kind regards

Arthur

************************************************************************ 
*******
Arthur J Rowe
Professor of Biomolecular Technology / Director NCMH Business Centre
School of Biosciences
University of Nottingham
Sutton Bonington
Leics LE12 5RD

TEL:  0115 9516156

************************************************************************ 
*******



On Jul 8, 2011, at 17:08, Ewa Folta-Stogniew wrote:

>
>
>
> At 10:37 AM 7/8/2011, Arthur Rowe wrote:
>
>> SEC column MW's are not credible, for well-known reasons. And I don;t
>> agree with Ewa that having DLS on-line would totally resolve the  
>> issue,
>> since SEC-DLS remains a non-absolute method. SEC-MALLS of course would
>> be excellent - if you have the kit.
>
>
> I suggested SEC/MALLS with DLS so I am not sure what there is to  
> disagree about?
>
> Ewa
>
>
>
>> Regards to you, and to all
>>
>> Arthur
>>
>> ********************************************************************** 
>> ** *******
>> Arthur J Rowe
>> Professor of Biomolecular Technology / Director NCMH Business Centre
>> School of Biosciences
>> University of Nottingham
>> Sutton Bonington
>> Leics LE12 5RD
>>
>> TEL:  0115 9516156
>>
>> ********************************************************************** 
>> ** *******
>>
>> On Jul 8, 2011, at 13:54, Ewa Folta-Stogniew wrote:
>>
>>> At 06:08 AM 7/8/2011, Mark Agacan wrote:
>>>
>>>> Hello RASMB,
>>>> Sedfit c(s) analysis of a protein I'm working with shows a 30 kDa
>>>> peak with S = 2.60 and f/f0 of 1.2, that corresponds to monomer.
>>>> SOMO in Ultrascan using the pdb file for this protein predicts a 30
>>>> kDa monomer at 2.42, with f/f0 of 1.19.  It's good agreement with  
>>>> the
>>>> sedimentation velocity results.
>>>> However, the crystal structure, SEC, etc. indicate this protein
>>>> should be a dimer.
>>>> Starting with the default value of 1.2 for the frictional ratio and
>>>> floating in sedfit returns a value of 1.2, with MW that corresponds
>>>> to monomer.  RMSD = 0.009756
>>>> Doubling this value to 2.4 and floating gives a value of 2.3, and
>>>> assigns a mass that corresponds to dimer to the main peak in the  
>>>> c(s)
>>>> analysis.  RMSD = 0.009324
>>>> Thus there appear to be two energy minima for the frictional ratio,
>>>> depending which initial value is used.  The RMSD value is lower when
>>>> the higher value of f/f0 is used.
>>>> Would you, on the basis of RMSD values, assign monomer or dimer to
>>>> this peak?
>>>
>>> I would run SEC/MALLS with DLS on line- to get direct measurement of
>>> MW and Rh. which will allow you to determine f/fo value. .
>>>
>>> See attached.
>>>
>>> Ewa
>>>
>>>
>>>> Thanks,
>>>> Mark
>>>>
>>>> ~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~
>>>> Dr Mark Agacan, Scientific Officer for the Division of Biological
>>>> Chemistry and Drug Discovery,
>>>> Wellcome Trust Biocentre, College of Life Sciences, University of
>>>> Dundee, Dundee, DD1 5EH
>>>> Tel: +44 1382 386095    Fax: +44 1382 345764    Mobile: 07525 451  
>>>> 117
>>>> ~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~
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>