Delta M Mode is a feature for identifying proteoforms containing unknown mass shifts, for example, PTMs. The delta is the difference between the observed precursor mass and the theoretical precursor mass. When you perform a search in Delta M Mode, the ProSightPD application concurrently performs three queries per sequence to compare the following:

  • The theoretical fragment ion masses of the protein sequence to the observed fragment ion list as usual
  • The theoretical fragment ion masses derived from the sequence and the Delta M applied N terminal to the observed fragment ion mass list
  • The theoretical fragment ion masses derived from the sequence and the Delta M applied C terminal to the observed fragment mass list

A Delta M Mode search takes approximately two times longer than the same search without Delta M Mode.

By carefully observing the pattern of fragments with and without Delta M Mode, you can locate the source of the mass shift. For example, a particular result returns with the two smallest N-terminal fragments matching without the mass shift. If all other matching N-terminal fragments contain the mass shift, the unknown mass can be localized on an amino acid between the second and third N-terminal fragments:

Figure Schematic of Δm mode
Δm=+2 Da Δm=+1 Da Δm=0 Da Δm=-1 Da Δm=-2 Da b, b+Δmy, y+Δm b, b+Δmy, y+Δm 2500 Da7500 Da 10000 Da Observedfragmention masses Theoretical mass for each protein candidate Theoretical fragment ion values for eachprotein candidate 9998 Da b, b+Δmy, y+Δm 9999 Da 10000 Da 10001 Da b, b+Δmy, y+Δm 10002 Da b, b+Δmy, y+Δm

When Delta M Mode is enabled, fragments found using the Delta M Mode display on the PrSMs page, along with two additional columns: Delta M Residue and Delta M Fragment(s), where the delta mass and the possible residue where the delta mass was localized. Delta masses also display in the ProSightPD fragment map with an orange square around the potential residue where the delta mass was localized.