From: Jo Butler <pjgb@mrc-lmb.cam.ac.uk>
  To  : Joel Mackay , rasmb@alpha.bbri.org
  Date: Wed, 24 Nov 1999 10:00:44 +0000

Re: monomer-dimer-tetramer

I hope people will forgive me for adding a further comment, namely that
this is exactly the sort of situation where using a completely different
technique can be very helpful.

Centrifugation alone, as Joel points out, is not good at resolving such
models as one can only fit the data as closely as possible and then
conclude which model is better fitted.  One example of an alternative is
cross-linking the protein and running a time course in SDS/PAGE.  It is
often very easy to follow the bands from monomer up through dimer to a
maximum - in these cases either timer alone or trimer plus tetramer.
Sometimes one also sees multimers of the main aggregate, but this is almost
always easily distinguished and often controllable by adjusting the initial
protein concentration.

I would argue very strongly in favour of using more than a single
technique, particularly where these rely upon radically different
properties and thus are not prone to similar artifacts/problems in
interpretation.  It gives one more confidence in the overall conclusion.

Jo Butler

--On Wed, Nov 24, 1999 12:56 pm +1100 Joel Mackay
<j.mackay@biochem.usyd.edu.au> wrote:

> Dear all,
> I would just like to thank all of those who replied to me a month or two
> ago regarding my problem trying to distinguish a monomer-trimer from a
> monomer-dimer-tetramer equilibrium (original mail below).
> It seems that this is a common problem, and that the best way around it is
> to record data over as wide a concentration range as possible. This is
> fairly obvious I hear you say, but it was reassuring to hear that
> distinguishing these two models is often tricky. I have placed the replies
> to this message on my web site, if people want to read them or download
> them, go to:
> 
> http://www.biochem.usyd.edu.au/~joel/extras/extras.html
> 
> I would also just like to say that I think this newsgroup is an extremely
> valuable resource for us all (especially for people like myself with only
> a few years experience with AU), and I hope it continues on well into the
> future.
> Thanks again.
> Best regards,
> Joel Mackay
> 
> 
>> Dear all,
>> I have a question about deciding which model describes one's data best. I
>> have recorded sedimentation equilibrium data at three speeds with three
>> different dilutions for a protein which undergoes some self-association.
>> I have been fitting the data in NONLIN. If i fit the data by fixing
>> sigma to the monomer mass, allowing delta y and lnA values to float, and
>> permitting a single association constant to float, i get the best fit
>> with a monomer-trimer model (both monomer-dimer and monomer-tetramer
>> have worse residuals and higher chi-squared etc according to NONLIN). If
>> I instead allow an extra equilibrium constant to float, and call the two
>> associations monomer-dimer and monomer-tetramer, i get a slightly lower
>> chi-squared (0.0138 vs 0.014 for the monomer-trimer model). My question
>> is, how do i decide if the extra complexity of the model is justified. I
>> know there is a thing called an F-test, and thought that might be
>> appropriate. If so, how does one apply it in this case? What do you all
>> do in these situations? It seems that the extra variable is pretty
>> risky, but presumably a
>> sufficiantly large reduction in the chi-squared would justify its
>> inclusion. cheers and thanks in advance for any help,
>> Joel Mackay
> 
> ************************************************************************
> Dr Joel Mackay                                 ph +61-2-9351-3906
> ARC Research Fellow                        fax +61-2-9351-4726
> Department of Biochemistry
> University of Sydney
> NSW 2006 Australia
> http://www.biochem.usyd.edu.au/~joel/
> ************************************************************************ 



P.J.G. Butler,
MRC Laboratory of Molecular Biology,
Hills Road,
Cambridge, CB2 2QH,
UK.