[RASMB] (no subject)

[pschuck at helix.nih.gov]

Hi All,
while we're discussing ways to estimate time to equilibrium - I like doing
that with SEDFIT, which also has the function to simulate sedimentation
for given molar mass, solution column, and s-value.  (I find this useful
also for double-checking that the rotor speed generates nicely curved
profiles.)

However, I agree with Arthur in that such estimates are only a starting
point.  In my experience they are not very useful other than giving you a
lower limit for the time it might take.  As you all know, there are two
inherent assumptions: 1) We should know the s-value, i.e. the shape of the
molecules.  In particular with proteins that are naturally unfolded, have
large unstructured domains, or are heavily glycosylated, the hydrodynamic
friction can be much higher, and the time to equilibrium I believe should
directly scale with that. (That could make a potential error in estimated
equilibrium time of perhaps 30%?)  We've had one of those largely
unstructured small proteins recently, and it took extremely long for it to
equilibrate. Lateron I realized that the frictional ratio from velocity
was about f/f0 = 2.  2) In the presence of interactions that are slow,
these may actually be the rate-limiting step. That's maybe not that
common, but apparently not extremely unusual, either.  Also, of course,
the buffer viscosity may be a big factor, for cases where glycerol can't
be avoided.  Therefore, I tend to accommodate for a great deal longer
equilibrium time, while scanning in 6 h intervals, and using MATCH to look
how the pairwise differences between scans (relative to the last scan)
slowly level off and reach a constant value.

Another concern relevant to this topic is that we found the profiles CLOSE
TO equilibrium frequently already look nicely exponential, and can also be
modeled reasonably well with a combination of equilibrium exponentials
(unfortunately not entirely correct ones).

Regards,
Peter