From: John Philo <jphilo@mailway.com>
  To  : Borries Demeler <demeler@bioc09.v19.uthscsa.edu>
  Date: Thu, 27 Apr 2000 09:59:52 -0700

RE: variation of local lnks

I think if on your plot versus rotor speed you connected the points for each
individual sample or color-coded them you would in fact see a slight trend
to higher association constants at higher speeds (if they connect the way I
think they do, anyway).

I thought from your earlier message you were always fitting to models
including species higher than dimer. If its just a monomer-dimer fit I can
run these data through my version with incompetent monomer if you like so
you can see how the Ka compares with the data at wavelengths where you only
see the heme.

John

-----Original Message-----
From: Borries Demeler [demeler@bioc09.v19.uthscsa.edu]">mailto:demeler@bioc09.v19.uthscsa.edu]
Sent: Thursday, April 27, 2000 8:11 AM
To: UMJ
Cc: rasmb@alpha.bbri.org
Subject: Re: variation of local lnks


>
> If you are sure the monomer-dimer is the correct model for your system I
> have a way to estimate the correct K2 from samples contaminated by either
> incompetent monomer or incompetent dimer, not by direct fitting though.
> Rather, the method requires NONLIN fitting of a series of data sets with
> different loading concentration and then, the K2 can be calculated using
the
> estimated NONLIN parameters - I call it the secondary calculation. If you
> are interested, I can provide you more info on that.
> John is right, simply including more fitting parameters can always improve
> the RMS of a fit, but it does not guarantee the fitting is correct. To be
> sure that incompetent species is what causes the problem, one should
follow
> Emory's suggestion - to observe the trend of variations of estimated lnK
> with loading concentration and/or speed. If it was incompetent large
> species, you can probably get rid of it at a higher speed. To deal with
> incompetent monomer is more challenging.
>
> Regards
> Yujia
>

Yes, I would be interested, I have sent a copy of a plot to Tom Laue
yesterday that shows the K2's plotted against loading concentration and
against speed, and these plots seem to indicate that I do infact have the
presence of incompetent monomers, which is what I suspected all along,
and it matches well with the biochemical hypothesis (how often can you say
that!). For those that are interested, I put the plot on the web at:

http://demeler.uthscsa.edu/data/k2.gif

It shows a clear decrease of k2 with increasing loading concentration,
but an uncorrelated effect with rotor speed. Tom indicated that this
means "incompetent monomer".

If Yujia or someone else has some software to do some secondary
analysis on this data, please let me know. If there is a way to
extract the k2 for the competent part I would be happy.

Regards, -Borries