Hi Jonathan, This is purely a chemistry guess, but if you have a PEG400 derived compound like:
(HO)-CH2-CH2-O-CH2-CHO, i.e, part oxidised, then the CHO might form a N,O acetal ring with the free amine and side chain. Five membered ring formation is kinetically favourable, though the chemistry might be a bit unlikely at your pH. Cheers, Partha On Tue, Jun 23, 2009 at 3:10 PM, Jonathan Elegheert < jonathan.eleghe...@ugent.be> wrote: > Dear bb, > > I have a high resolution dataset showing that the N-terminal threonine of > my protein has been modified and cyclized, with the main chain N and side > chain OG1 now involved in a non-planar 5 membered ring sugar-like structure. > The molecule seems to be 5-6 atoms long with some 'head group'. > > http://studwww.ugent.be/~jeleghee/ccp4/coot_front.png<http://studwww.ugent.be/%7Ejeleghee/ccp4/coot_front.png> > http://studwww.ugent.be/~jeleghee/ccp4/coot_side.png<http://studwww.ugent.be/%7Ejeleghee/ccp4/coot_side.png> > http://studwww.ugent.be/~jeleghee/ccp4/coot_angle.png<http://studwww.ugent.be/%7Ejeleghee/ccp4/coot_angle.png> > > Modification must have occurred in the drop since N-terminal sequencing > through Edman degradation after it came from the column gave a signal, i.e. > the N-terminal Thr amine was free. This also confirmed that the initiator > Met is definitely cleaved off. > > The crystallization condition is as follows; > - 0.1 M Tris pH 8.2 > - 1.65 M (NH4)2SO4 > - 2 % PEG400 > - 0.25 % (v/v) β-octyl glucoside > > It is worthy to mention that in the periphery of the protein, ethylacetate > is bound (the reaction product of the ethanol and acetate > impurities/breakdown products of the PEG400 stock solution). I don't know > whether any PEG400 breakdown products or others derived from some of my > crystallization components could show that sort of reactivity. > > Many thanks in advance if anyone has any pointers, > > Jonathan > > -- > Jonathan Elegheert > Ph.D. Student > Ghent University <jonathan.eleghe...@ugent.be> > > -- MRC National Institute for Medical Research Division of Molecular Structure The Ridgeway, NW7 1AA, UK Email: pcha...@nimr.mrc.ac.uk Phone: + 44 208 816 2515
