On the PE-ABI PROCISE platforms PTH-Met sulphoxide co-elutes with
PTH-threonine and, coincidentally, PTH-homoserine. We frequently observe
high PTH-Thr from "real" protein samples with N-terminal methionine and
less obviously from internal methionine. We rationalise this by noting
that methionine sulphoxide is not stable in acidic conditions in the
presence of sulphur in a low oxidation state.... thiol, thioether etc. The
sequencing chemistry should thus reduce all PTH-Met sulphoxide to PTH-Met.
Clearly, on the PROCISE platform this reduction is not complete in the
early cycles if RELATIVELY high molar amounts of the sulphoxide are
present. One would expect that the DTT-containing aqueous TFA (R4A) should
be an especially effective reductant so I would expect that the
detectability of PTH-Met sulphoxide would vary significantly between
platforms.
Noting that PTH-homoserine also co-elutes with both PTH-methionine
sulphoxide and PTH-Thr confounds the diagnosis of failed CNBr cleavages.
PTH-methionine sulphone elutes after DMPTU, slightly later than
PTH-S-carboxamidomethylcysteine. I suspect that PTH-methionine sulphone
would coelute with PTH-S-carboxamidomethylmethionine but I have not
verified this. The identification of PTH-methionine sulphone would thus be
confounded by exposure to iodoacetamide.
Personally, I would investigate this problem by using enzymatic digestion
to generate a set of native peptides and use mass analysis to pick peptides
which are more massive than expected by multiples of oxygen atoms.
Assuming that you are dealing with incompletely oxidised protein the
patterns should be fairly easy to pick.
cheers
Milburn
Dr.Peter Milburn
Biomolecular Resource Facility
The Australian National University
GPO Box 334 Canberra ACT 0200 AUSTRALIA
Phone No. <61> <2> 6249 4326
Fax No. <61> <2> 6249 4326
E-mail Address Peter.Milburn@anu.edu.au