From: mmorris <michael.morris@adelaide.edu.au>
  To  : RASMB <rasmb@alpha.bbri.org>
  Date: Mon, 16 Jul 2001 10:40:40 +0930

Re: Differentiating between folded and unfolded states in sed vel

Hi Les,

If the rate of reaction is fast you will not be able to distinguish the
two species. Instead, you will observe only one species whose s and D at
zero concentration should return the correct molecular weight. If the
returned value of molecular weight is lower than expected then the rate
of interconversion is either very slow (effectively zero) or not quite
fast enough to rate as "rapid" - the latter is unpleasant to deal with.

The only way I can see that you can detect the presence and quantify the
ratio of the two species is first to use conditions that provide
effectively 100% of the compact form (lower limit of s20,w) and 100% of
the unfolded form (upper limit of s20,w). Conditions which have a
mixture of the two forms (in rapid equilibrium) should then have a
weighted value of s20,w between these limits from which the percentage
of each form could be calculated.

Yours, Michael

Les Hicks wrote:
> 
> Hi folks,
> 
>     Here's a hypothetical question.  Say you have an approximately 7 kDa
> protein that is known from other techniques to undergo reversible folding /
> unfolding in phosphate buffer.  If the rate of folding / unfolding is fast
> (within the time course of a sed vel run, would it be possible to
> differentiate between the two states from the sed vel data?  I would think
> all you would see is a slightly broader boundary and that you would not be
> able to identify two individual boundaries for the two states.  Any takers
> on this question?  In any event, have a good week-end.
> 
> Regards,
> 
> Les Hicks
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