We have some samples that came from protein crystals on which we collected data The protein was chemically modified by the addition of one -CH2PO3 group putatively to a single cysteine reside; however, showing this by LC/MS of the trypsin-derived peptides has not been successful, despite repeated attempts. Whole-protein mass spectrometry does show an increase in mass. After diffraction data were collected, the crystals were dissolved and frozen.
We would like to show that the crystals still contain the modification. It occurs to me that the retention time in RP-HPLC or the mass spectrum of the protein in these samples would be useful in showing that the protein that was crystallized was or was not identical to the protein prior to crystallization. However, the protein is generally crystallized in the presence of a high concentration of high-MW polyethylene glycols. Therefore, we need to remove the PEG from the protein and possibly concentrate. I was thinking of ZipTips. Does anyone have suggestions?