matth at NOSPAM.biols.sussex.ac.uk
Thu Oct 21 05:16:20 EST 1999
Size exclusion chromatography is not better than analytical
ultracentrifugation or light scattering. As for detection of, for example,
coiled-coil protein oligomerisation by centrifugation - it works very well
and you can get accurate association constants and molecular weights. Size
exclusion chromatography by comparison is a low resolution technique.
Is the aggregation that happens with the protein in question lots of
different molecular weights or a few discrete ones (centrifugation will give
you the distribution of molecular weights). If the latter is the case it
may be that the aggregated form is the biologically active one...
tob at ssi.dk wrote in message <99Oct20.104506gmt+0100.19744 at fsk.ssi.dk>...
>I agree with the other views but if you have a protein that bind to itself
in a ordered manner I do not believe that you detect it with these methods.
Take for example a coil-coil protein that forms an aggregate or rather dimer
or trimers. I believe you have used the best method, size exclusion
chromatography already. If you continue with size exclusion chromatography
try GuHCl or Urea addition to the buffers to get a monomer and use this as a
reference. If you do not get a monomer, you probably have a covalent
complex. If you do get monmeric protein go on with other additives,
detergents, amino acids, glycerol and so on.
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