Phosphopeptide analysis by directly coupling two-dimensional planar electrochromatography/thin-layer chromatography with matrix-assisted laser desorption/ionization time-of-flight mass spectrometry.
A novel strategy is presented for the fractionation of complex peptide mixtures using two-dimensional planar electrochromatography/thin-layer chromatography (2D PEC/TLC). Phosphopeptides migrate more slowly in the first dimension, based upon their anionic phosphate residues, and certain predominantly acidic phosphopeptides even migrate in the opposite direction, relative to the bulk of the peptides. Phosphopeptides are further distinguished based upon hydrophilicity in the second dimension. This permits a restricted region of the plate to be directly interrogated for the presence of phosphopeptides by mass spectrometry (MS). Phosphopeptide analysis from the plates by matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF)-MS and tandem MS enabled peptide sequencing and identification.
Panchagnula
Venkateswarlu
V
Biochemistry Department, PerkinElmer Life and Analytical Sciences, Waltham, MA 02451, USA.
Mikulskis
Alvydas
A
Song
Linan
L
Wang
Yang
Y
Wang
Mei
M
Knubovets
Tanya
T
Scrivener
Elaine
E
Golenko
Eva
E
Krull
Ira S
IS
Schulz
Michael
M
Heinz-Emil-Hauck
Patton
Wayne F
WF
D053801:Capillary Electrochromatography / Q000379:methods*; D002855:Chromatography, Thin Layer / Q000379:methods*; D010455:Peptides / Q000032:analysis* / Q000302:isolation & purification / Q000378:metabolism; D010750:Phosphoproteins / Q000032:analysis / Q000302:isolation & purification / Q000378:metabolism; D040901:Proteomics / Q000379:methods; D015203:Reproducibility of Results; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods*; D014357:Trypsin / Q000378:metabolism
D010455:Peptides; D010750:Phosphoproteins; D014357:Trypsin