20
P. ROEPSTORFF
. .
~===::::::::"i "
o
~
r-------------c-------------~ o~
u
C/)
o
~
.;-----,.~;:===t
~
Fig. 2.4. Example of protein identification based on a peptide sequence tag. (A) nanoESI-spectrum
obtained on a QTOF mass spectrometer of the peptide mixture obtained after in-gel digestion and
micropurification of a protein spot from a 2D-gel of an immune precipitated human protein complex.
(B) The resolution is sufficient to identify the selected peak as doubly charged based on the spacing
between the isotope peaks. (C) MS/MS spectrum obtained upon selection and collision induced dissociation of the peaks in the isotope envelope at m/z 775-777. Based on singly charged fragment ions
(D) a sequence tag "850.5 Mo x SY1247.6" can be generated. Upon search in the EBI non-redundant
database the peptide is identified as a tryptic peptide from the constant region of a mouse antibody
kappa light chain. Thus, the protein in the gel-spot is the antibody used for immune precipitation
P. ROEPSTORFF
. .
~===::::::::"i "
o
~
r-------------c-------------~ o~
u
C/)
o
~
.;-----,.~;:===t
~
Fig. 2.4. Example of protein identification based on a peptide sequence tag. (A) nanoESI-spectrum
obtained on a QTOF mass spectrometer of the peptide mixture obtained after in-gel digestion and
micropurification of a protein spot from a 2D-gel of an immune precipitated human protein complex.
(B) The resolution is sufficient to identify the selected peak as doubly charged based on the spacing
between the isotope peaks. (C) MS/MS spectrum obtained upon selection and collision induced dissociation of the peaks in the isotope envelope at m/z 775-777. Based on singly charged fragment ions
(D) a sequence tag "850.5 Mo x SY1247.6" can be generated. Upon search in the EBI non-redundant
database the peptide is identified as a tryptic peptide from the constant region of a mouse antibody
kappa light chain. Thus, the protein in the gel-spot is the antibody used for immune precipitation