Diagnostic protein discovery using liquid chromatography/mass spectrometry for proteolytic peptide targeting.
A peptide targeting method has been developed for diagnostic protein discovery, which combines proteolytic digestion of fractionated plasma proteins and liquid chromatography coupled to electrospray time-of-flight mass spectrometry (LC/ESI-TOFMS) profiling. Proteolysis prior to profiling overcomes molecular weight limitations and compensates for the poor sensitivity of matrix-assisted laser desorption/ionization (MALDI) protein profiling. LC/MS increases the peak capacity compared to crude fractionation techniques or single sample MALDI analysis. Differentially expressed peptides are targeted in the mass chromatograms using bioinformatic techniques and subsequently sequenced with MALDI tandem MS. In a model study comparing pancreatic cancer patients to controls, 74% of the peptide targets were successfully sequenced. This profiling method was superior to previous experiments using single sample MALDI analysis for protein profiling or proteolytic peptide profiling, because more potential protein markers were identified.
Koomen
John M
JM
Molecular Pathology, University of Texas M.D. Anderson Cancer Center, 1515 Holcombe Blvd., Houston, TX 77030, USA.
Zhao
Haitao
H
Li
Donghui
D
Nasser
Waleed
W
Hawke
David H
DH
Abbruzzese
James L
JL
Baggerly
Keith A
KA
Kobayashi
Ryuji
R
D000328:Adult; D000368:Aged; D000595:Amino Acid Sequence; D001798:Blood Proteins / Q000032:analysis*; D005591:Chemical Fractionation; D005260:Female; D006801:Humans; D008297:Male; D008875:Middle Aged; D008969:Molecular Sequence Data; D010190:Pancreatic Neoplasms / Q000097:blood / Q000175:diagnosis; D010449:Peptide Mapping / Q000379:methods*; D012680:Sensitivity and Specificity; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods*
D001798:Blood Proteins