Dear Raji, The best way to find out is to run a SEC-MALLS (Size Exclusion Chomatography - Multi-Angle Laser Light Scattering) experiment.
Best, Isabel ----------------------------------------------------------------------------------------- Dr. Isabel De Moraes, MRSC Membrane Protein Laboratory Facility Co-ordinator Membrane Protein Laboratory Diamond Light Source Ltd, Chilton, Didcot, Oxfordshire, OX11 ODE, UK Tel (direct): 01235 778664 ------------------------------------------------------------------------------------------ ________________________________ From: CCP4 bulletin board [CCP4BB@JISCMAIL.AC.UK] on behalf of Jacob Keller [j-kell...@fsm.northwestern.edu] Sent: 21 June 2012 17:11 To: ccp4bb Subject: Re: [ccp4bb] Detergent and protein oligomerization First of all, isn't the choice either dimer or trimer, and second, as a protein-detergent complex (PDC), it would be very unlikely that a trimer of 99 kD would run at 100 kD, although all is fair in love, war, and membrane proteins. JPK On Thu, Jun 21, 2012 at 10:50 AM, Raji Edayathumangalam <r...@brandeis.edu<mailto:r...@brandeis.edu>> wrote: Hi Everyone, Sorry for the non-CCP4 post. I have a very basic question about detergents, critical micelle concentration and behavior on gel filtration. A 33kDa membrane protein was purified by gel filtration in a buffer containing 0.4%(w/v) beta-NG (CMC: 6.5mM) and 0.046%(w/v)LDAO (CMC: 0.14mM). So the concentrations of beta-NG and LDAO in the gel-filtration buffer are ~2X and ~14X that of the CMCs of the respective detergents. The elution volume of the protein peak (plus detergent) on Superdex200 corresponds to a molecular mass of 100kDa. I think that the 100kDa mass above includes contributions from both the protein as well as the detergent micelles. If this is correct, is it then accurate to try to glean the oligomerization state of the protein (and conclude that it is a trimer or tetramer) without taking into account detergent micellar mass and its influence on elution volume? How should one interpret the 100kDa mass estimate from the gel filtration? Thanks. Raji -- Raji Edayathumangalam Instructor in Neurology, Harvard Medical School Research Associate, Brigham and Women's Hospital Visiting Research Scholar, Brandeis University -- ******************************************* Jacob Pearson Keller Northwestern University Medical Scientist Training Program email: j-kell...@northwestern.edu<mailto:j-kell...@northwestern.edu> *******************************************
