Re: Misc Strength of protein protein interactions

Jim.Bloom.B@bayer.com
Tue, 5 May 1998 16:49:48 -0400

I recognize this as a potential application for an analytical
ultracentrifuge...although I have one I haven't done what you are asking but I
will suggest that you contact Dr. John Philo at 805-492-6171 (I converse with
him on email but our "firewall" is such that I don't know his address). John
is well known for his AUC and gel filtration/on-line laser light scattering
work.
As people are sensitive about such things let me give ulterior motives for my
giving you John's name: 1.) he knows his stuff and has been extremely helpful
to me; 2.) he and his group were recently let go by Amgen and replaced by
"small molecule" people...although this was good for the "small molecule"
people it was not so good for the protein people...so John and his group have
started Alliance Protein Laboratories...I apologize to the ABRF for plugging a
company but I felt a cold wind on the back of my neck when I heard John's
story...are proteins soon to be made obsolete?

Weather consequential: I went for my lunch time walk and heard TWO claps of
thunder...in the Bay area that is VERY rare...

Jim
Bayer Corp.
Berkeley
510-705-7760

EXNETWRK.abrfreq@BAYERMAIL on 05/05/98 11:33:10 AM
To: "DDA.RFC-822=abrf@aecom.yu.edu/P=Internet/A= /C=us"@X400
cc:
Subject: Misc Strength of protein protein interactions

Colleagues,

A friend is studying two subunits of a multi subunit
bacterial enzyme complex. The two subunits that he is
studying form a stable heterodimer in solution and a goal of
his research is to evaluate the nature of the interaction
between these two subunits. Both proteins can be expressed
and purified in large amounts for physical analysis.

He has made mutations in secific sites that directly
decrease the affinity of the two subunits for formation of
the heterodimer. Using gel filtration chromatography, he is
readily able to demonstrate the presence of the dimer but
needs to quantify the magnitude of the association constants
for the various mutants.

He has two questions:

1. How does one measure the association constant for two
proteins? (I have suggested using Biacore but we do not have
access to this methodology.)

2. What is the lower limit for an association constant
below which one would not expect to be able to observe
significant dimerization during gel filtration
chromatography?

Thanks in advance for any input and suggestions that might
point us to the right literature.
Mark Lively