[RASMB] NONLIN

[Mary Judith Kornblatt]

Daryl, I am not an expert - in fact, I haven't been happy with any of my
fits but here are a few things that I have learned. Set sigma for what you
know it is and do not fit; you know the monomer molecular weight! I usually
make a educated guess of Ka and enter that value (ie, I don't start with it
at zero). I have found, especially when I try varying other things, that
NONLIN can get stuck in a bad fit and not get out again. So you want to
start by fitting as few things as possible and start with reasonable
guesses. Judith Kornblatt



At 09:07 AM 5/8/02 -0400, Daryl Bosco wrote:
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>Hello
>
>I have recently started using NONLIN in an effort to calculate Kd's for
>a self associating system (comparing WT and mutants).
>
>I first used the Beckman/ Origins software to calculate a Ka in
>absorbance units which I converted to a Kd.  I then went to NONLIN and
>found that the Ka in absorbance units (I assume that is what I am
>calculating) is many orders of magnitude off.  I assume this is because
>I am not experienced with the program and how it is working.
>
>First question:
>
>Can anyone tell me explicitly where I may find a comprehensive manual
>for NONLIN?  I have been on the web and found useful material (including
>past emails to RASMB), but not a manual.  For example, I was not able to
>find how to plot the residuals under the plot menu.  (?)
>
>Second more detailed question (for those who have the time):
>
>I set up the NONLIN fitting procedure according to the instructions for
>self association in the README file that accompanied NONLIN.  First I
>read in 8 files (one speed, one detected wavelength, 8 concentrations).
>Set the value for LnK2 to 0 (allow to converge).  Delta y is 0 (b/c this
>is absorbance), LnA refers to the lowest concentration and so I set this
>to -12 (for 7 uM)--allow to converge.  Set sigma to 0.8 (calculated with
>SEDNTERP) and allow to converge.  Do not put anything in for
>concentration and speed factors b/c only one speed used here and one
>wavelength detected.  LnK2 is calculated by NONLIN as -13.  I assume
>this means the Ka in absorbance is 3.1 e-6 (in Beckman I get Ka in
>absorbance of 0.151).  I know the accuracy will improve by using another
>speed, but I am first trying to get a handle on the program and hope to
>get something that is reasonable (Ka of 3.1 e-6 is not reasonable).  If
>the more experienced NONLIN user can already see a fatal error in my
>fitting approach, would you please let me know?
>
>Your responses are greatly appreciated, thank you
>
>Daryl Bosco
>
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Mary Judith Kornblatt
Department of Chemistry and Biochemistry
enzyme research group
Concordia University
1455 de maisonneuve ouest
montreal, qc, canada h3g 1m8
tel 1 514 848 3384, fax 1 514 848 2868