native and oxidized protein.. separation on Ion exchange chromatography? - possible? method suggestions? (Aug/01/2008 )
hii i m trying to separate a native and oxidized protein (could be a single methionine oxidation also).. on Ion exchange chromatography? u think its possible? any method suggestions? shud i do it by pH gradient or salt gradient? which works better for resolution for such close species?
-alice!-
you can run them on ief to find out if there is a difference in pI. if there is then you can separate with a pH gradient.
-mdfenko-
QUOTE (alice! @ Aug 1 2008, 12:53 AM)
hii i m trying to separate a native and oxidized protein (could be a single methionine oxidation also).. on Ion exchange chromatography? u think its possible? any method suggestions? shud i do it by pH gradient or salt gradient? which works better for resolution for such close species?
beside the technical problem, isn´t there the bigger problem to conserve the forms of native and oxidized protein? during run, you normally have to prevent oxidaztion, f.i. by adding DTT or DTE;
if you do this not, the native fration will easily become oxidized during column run, and if you do, you may reduce your oxidized protein to the native state...
-The Bearer-