[RASMB] Analytical Centrifuge Question

Arthur Rowe arthur.rowe at nottingham.ac.uk
Wed Jul 28 06:04:00 PDT 2004


Hi Anna

It is certainly possible to study saccharides in the AUC. I have done it.
You will need to observe the following precautions:

(1) a cell blank - always a good thing - is absolutely vital for
distributions which are clearly minimal in their range of fringe increment.

(2) a long column will give the best data range. The standard formula (e.g.
from BIOMOLS - RASMB archive) gives us around 12 hours equilibration time
for a typical saccharide (maltose, D = 42 E-7) if we use a 5 mm column at
20º. You will need to run at the highest speed you can get away with in
terms of avoiding window (and hence fringe) distortion. But even being
restricted to 40000 things should still be OK

(3) get the concentration up. NO danger of big fringe increments or
schlieren effects. Obviously if you are looking for a Ka value then that may
influence your choice of loading concentration. In general around 10 mg/ml
is a good choice. Of course there is the thermodynamic non-ideality to take
into account (which would make an apparent monomer mass around 10% - very
roughly - too low).

(4) to cope with estimating Ka in the presence of significant 2BM you can
try fixing a pluasible B value in NONLIN (floating both it and K1,2 is a
no-no). I had a poster at the recent AUC Meeting in Oxford showing off INVEQ
- an algorithm which specifically copes with both Ka and BM as variables.

Good luck! It should work.

Arthur

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Arthur J Rowe
Professor of Biomolecular Technology
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School of Biosciences
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Leicestershire LE12 5RD   UK

Tel:        +44 (0)115 951 6156
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From: anna_ber at alcor.concordia.ca
Date: Tue, 27 Jul 2004 10:01:53 -0400
To: rasmb at server1.bbri.org
Subject: [RASMB] Analytical Centrifuge Question


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Hello,

I am working in Dr. Muchalls lab and I have to study a monomer dimer
equilibrium. I was wondering whether it is possible to study this
equilibrium 
with the analytical centrifuge? I have read that molecules as small as
sucrose 
have been experimented with in the analytical centrifuge but I have no clue
as 
to the lower limit of the apparatus? Is it possible to study a molecule with
a 
molar mass of 188.65g/mol and a dimer of 377.3g/mol.

Thank You 

Anna Bernardi  
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