Investigations with O-linked protein glycosylations by matrix-assisted laser desorption/ionization Fourier transform ion cyclotron resonance mass spectrometry.
Posttranslational modifications such as glycosylation can play a fundamental role in signaling pathways that transform an ordinary cell into a malignant one. The development of a protocol to detect these changes in the preliminary stages of disease can lead to a sensitive and specific diagnostic for the early detection of malignancies such as ovarian cancer in which differential glycan patterns are linked to etiology and progression. Small variations in instrument parameters and sample preparation techniques are known to have significant influence on the outcome of an experiment. For an experiment to be effective and reproducible, these parameters must be optimized for the analyte(s) under study. We present a detailed examination of sample preparation and matrix-assisted laser desorption/ionization Fourier transform ion cyclotron resonance mass spectrometry (MALDI-FT-ICR-MS) analysis of O-linked glycans globally cleaved from mucin glycoproteins. Experiments with stable isotope-labeled biomolecules allowed for the establishment of appropriate acquisition times and excitation voltages for MALDI-FT-ICR-MS of oligosaccharides. Quadrupole ion guide optimization studies with mucin glycans identified conditions for the comprehensive analysis of the entire mass range of O-linked carbohydrates in this glycoprotein. Separately optimized experimental parameters were integrated in a method that allowed for the effective study of O-linked glycans.
Williams
Taufika Islam
TI
W.M. Keck FT-ICR Mass Spectrometry Laboratory, Department of Chemistry, North Carolina State University, Raleigh, NC 27695, USA.
Saggese
Diana A
DA
Toups
Kristina L
KL
Frahm
Jennifer L
JL
An
Hyun Joo
HJ
Li
Bensheng
B
Lebrilla
Carlito B
CB
Muddiman
David C
DC
D000809:Angiotensins / Q000737:chemistry; D000818:Animals; D006031:Glycosylation; D009077:Mucins / Q000737:chemistry*; D009844:Oligosaccharides / Q000737:chemistry; D015203:Reproducibility of Results; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods*; D017550:Spectroscopy, Fourier Transform Infrared / Q000379:methods*; D013552:Swine; D053719:Tandem Mass Spectrometry / Q000379:methods*
D000809:Angiotensins; D009077:Mucins; D009844:Oligosaccharides