Hi everyone,
I think if you have a highly concentrated ligand stock solution you
typically disregard
the change of volume/receptor concentration.
As long as you can justify the above assumption the thing you are observe
has more to do with ligand excess / the ligand/receptor ratio. And
this is independent
of the total volume.
This all may of course not apply if you are unlucky enough to have a ligand that
is only available in dilute solution.
Regards,
jcw
On Wed, Feb 8, 2012 at 6:24 PM, Wayne Boucher <[log in to unmask]> wrote:
> Ah, Tim/Magnus are suggesting that if V is the original volume then you are
> adding dV extra volume and so the concentration of A (including that in AB)
> has changed from a to aV/(V+dV). If dV is large enough then that could be a
> problem. Is dV large enough? And if it is a problem then we would have to
> have some way for dV/V to be input (not for each point, because presumably
> this is proportional to how much B is added, so if you add twice as much
> extra volume that is because you are adding in twice as much B).
>
> Wayne
>
> On Wed, 8 Feb 2012, Wayne Boucher wrote:
>
>> Hello,
>>
>> Just to make sure what we are talking about (!), if we have the reaction A
>> + B <--> AB where A is the protein (say) and B the ligand, then the total
>> concentration of A is a = [A]+[AB] and the total concentration of B is b =
>> [B]+[AB]. So are you just saying that [A] decreases when you add B, because
>> that I agree with. The Analysis code assumes not that [A] is fixed but that
>> a is fixed (and perhaps I'm using the wrong terminology to describe that
>> situation). So, alternatively, are you saying that a itself might change??
>>
>> Wayne
>>
>> On Wed, 8 Feb 2012, Fowler, Andrew wrote:
>>
>>> I may be wrong, but I think what Aldino means by changing both the ligand
>>> and protein concentration in a titration is that one starts with free
>>> protein at a given concentration and volume and adds increasing amounts
>>> of
>>> ligand to that. So the protein concentrations decreases while the ligand
>>> concentration increases, but both are known at each point so you really
>>> don't have to fit two independent variables. It's not be the ideal case,
>>> but for various reasons many (most?) titrations get done this way.
>>>
>>> Analyzing this sort of titration data would be a very nice addition to
>>> Analysis - I know I would find it useful. The equation to fit the data is
>>> a bit messier but input would be both ligand and protein concentration
>>> for
>>> each point (or if you wanted to be really nice, starting concentration
>>> and
>>> volume of protein, ligand stock concentration, and the volume of ligand
>>> added at each step with the program calculating from that).
>>>
>>> Cheers,
>>> Andrew
>>>
>>> --
>>> C. Andrew Fowler, Ph.D. | University of Iowa
>>> Associate Director | B291 Carver Biomedical Research
>>> Building
>>> Medical NMR Facility | Iowa City, IA 52242
>>> 319-384-2937 (office) | 319-335-7273 (fax)
>>> [log in to unmask]
>>>
>>>
>>>
>>>
>>> On 2/8/12 10:22 AM, "Wayne Boucher" <[log in to unmask]> wrote:
>>>
>>>> Hello,
>>>>
>>>> The two titration type functions that are in Analysis are for
>>>> protein-ligand in fast exchange where you vary the ligand concentration
>>>> but keep the protein concentration fixed, and a monomer-dimer in fast
>>>> exchange, where you vary the total concentration. The equations are
>>>> discussed in a PDF file obtainable at:
>>>>
>>>>
>>>> https://sites.google.com/site/ccpnwiki/Home/documentation/ccpnmr-analysis/
>>>> FAQs/data-analysis
>>>>
>>>> If you want to vary two concentrations then that complicates any attempt
>>>> at fitting (it looks like to me but someone out there probably knows
>>>> better).
>>>>
>>>> The current Analysis does the fitting in C functions. Unfortunately
>>>> that
>>>> turns out to be some of the worst C code in Analysis because stuff kept
>>>> getting bolted onto it, so I wouldn't recommend anyone try to adjust
>>>> that
>>>> (although if you want to I can tell you what needs doing).
>>>>
>>>> I've written a Pythonic version of the fitting but it has not been
>>>> hooked
>>>> into the user interface. I will send you information about that in a
>>>> separate post, because it's likely to be the easiest way for you to add
>>>> your own functions, it's just that the visualisation is not there, so
>>>> would have to be done in (say) Excel. (You can also let us know what
>>>> you
>>>> want added and I can add it into the C functions.)
>>>>
>>>> Wayne
>>>>
>>>> On Wed, 8 Feb 2012, Aldino wrote:
>>>>
>>>>> Hi all,
>>>>> I have some questions regarding curve fitting on analysis:
>>>>>
>>>>> 1) is there any function on analysis to fit a concentration titration
>>>>> set in
>>>>> which you change both the ligand and the receptor concentrations?
>>>>>
>>>>> 2) is the Michaelis-Menten funtion defined?
>>>>>
>>>>> 3) where can I find information on the several funtions in avalysis?
>>>>>
>>>>> 4) can I write my own funtions?
>>>>>
>>>>>
>>>>> Cheers,
>>>>> Aldino Viegas
>>>>>
>>>>>
>>>>> ______________________________________________
>>>>> Aldino Viegas, PhD Student
>>>>> Dep. Química, REQUIMTE
>>>>> Faculdade de Ciências e Tecnologia
>>>>> Universidade Nova de Lisboa
>>>>> 2829-516 Caparica, Portugal
>>>>>
>>>>> Tlf. +351 212948300
>>>>> Ext. 10900 Lab. 106-A
>>>>> [log in to unmask]
>>>>>
>>>
>>>
>>>
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