Re: 2D electrophoresis
Gary Hathaway (hathaway@cco.caltech.edu)
Wed, 6 May 1998 02:36:37 -0700
>Date: Wed, 6 May 1998 02:16:12 -0700
>To: "Mark Lively" <mlively@medcenter.wpmail.wfu.edu>
>From: Gary Hathaway <hathaway@cco.caltech.edu>
>Subject: Re: Misc Strength of protein protein interactions -Reply
>Cc:
>Bcc:
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>At 1:14 PM -0700 5/5/98, Mark Lively wrote:
>>Gary,
>> Thanks for the input. We actually do have a) and b). Our Beckman
>>Analytical Ultra is four doors down the hall from my colleague's office.
>>What I did not communicate well is the main question we have is one of
>>magnitude of interactions of this type. We know, for example, that
>>biotin-avidin complexes have association constants in the range of 10E13.
>>But what he and I have no feel for is the lower end of that scale. If one
>>sees only dimer during gel filtration chromatography then one should be
>>able to sat that the association constant is "at least ..." Do you have a
>>feel for that lower limit?
>>Thanks for your interest. I will tell my friend to learn to use the AU.
>>Mark
>>
>Mark,
> I spent a good deal of my graduate research trying to decide whether
>lactic acid dehydrogenase dissociated in dilute salt solutions. The final
>conclusion I made was that it did not. I spent the next 4 years trying to
>decide whether tryptophan synthase B subunit dissociated in dilute
>solution. Then answer was, yes, but only as the apoprotein not as the form
>with pyridoxal phosphate bound. So,if we're talking about measuring
>dissociation constants in dilute "physiological" solutions then my feeling
>is that the equilibrium constants for most multisubunit proteins and
>certainly avidin-biotin complexes will be difficult to measure by this
>technique. But then, they probably wont be by Gary Ackers' gel filtration
>method either. I can't give you a hard number for the range of
>dissociation constants that could be measured, but I'm certain such info.
>exists in the literature. Perhaps some of Beckman Instruments' literature
>on their centrifuge covers it, or should anyway. The beauty is that weaker
>interactions are probably easier to measure than strong ones with the
>centrifugal method and the measurements are precise. I wasn't just being
>facetious about needing a physical biochemist. There are many ways to do
>this type of experiment all with their unique artifacts that must be
>understood and avoided or overcome. You can get in a lot of trouble even
>if you know the subject in depth, but when it works, the data are elegant
>and (I think) the best.
best regards
>
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Gary M. Hathaway, Director
PPMAL - Protein/Peptide Micro Analytical Laboratory
California Institute of Technology
139-74, Division of Biology
Pasadena, CA 91125
http://www.cco.caltech.edu/~ppmal
email Gary: hathaway@cco.caltech.edu
email facility: ppmal@cco.caltech.edu
phone: lab (626) 395-6388 or office (626) 395-2769
FAX (626) 449-3414
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