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Multi-Angle Light Scattering (MALS) coupled with an HPLC column attached to an analytical SEC column is my standard "go-to" method for determining molecular weights/oligomeric states of peaks from SEC runs. I've only ever used the detectors from Wyatt ( http://www.wyatt.com/solutions/hardware/hardware.html), not sure if there are any other manufacturers in the business. Cheers, Jim On Mon, Aug 13, 2012 at 10:04 AM, Sangeetha Vedula <[log in to unmask]>wrote: > Hello all, > > I am working with a protein that is probably a hexamer based on homology > with other proteins but when I ran it on an analytical size gel filtration > column, I see multiple peaks. I would like to determine the exact > oligomerization state of the mixture and have considered blue native gel > electrophoresis and gradient centrifugation. The monomer is about 54 kDa. > All of the peaks are active enzymatically. I wish I could test higher > concentrations to see if the equilibrium will shift towards a hexamer (or > whatever the naturally highest state is) but the protein crashes out. > > The protein was expressed in baculovirus and the multitude of states could > be an artifact but I'd like to know what I am working with. Also, down the > line, I'll need to pick one of the states for crystallization trials. > > Has anyone encountered this problem? How did you determine the > oligomerization state(s)? > > I have basic biorad gel apparatus available and no gradient mixer. > > Thanks for any input. > > Best regards, > > Sangeetha. > > > -- Jim Fairman, Ph D. Crystal Core Leader I Emerald BioStructures <http://www.emeraldbiostructures.com/> Tel: 206-780-8914 Cell: 240-479-6575 E-mail: [log in to unmask] [log in to unmask]