Rapid and sensitive analysis of mucin-type glycans using an in-line flow glycan-releasing apparatus.
Rapid and sensitive analysis of glycans is essential for glycomics. We previously reported an apparatus, the AutoGlycoCutter (AGC), for rapid release of O-linked glycans under alkaline conditions and its application to rapid analysis of glycans in proteoglycans. We now report an application of the AGC to obtain mucin-type glycans with reducing end (i.e., hemiacetal group) within only 3 min. The released oligosaccharides could be labeled with fluorescent 2-aminobenzoic acid for analysis by normal-phase high-performance liquid chromatography (NP-HPLC) and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS). We could detect O-glycans from as low as 5 pmol of bovine caseino glycomacropeptide (CGMP) by the proposed procedures. The validity of the current method was shown by the analyses of the released O-glycans from some standard glycoproteins: bovine submaxillary mucin, bovine fetuin, porcine stomach mucin, and human colostrum immunoglobulin A. The advantage of the current method was also demonstrated in comparative analysis of mucin-type glycans in CGMP derived from three different animal species.
Yamada
Keita
K
Faculty of Pharmaceutical Sciences, Kinki University, Higashi-Osaka, Osaka 577-8502, Japan.
Hyodo
Satomi
S
Matsuno
Yu-ki
YK
Kinoshita
Mitsuhiro
M
Maruyama
Shu-zou
SZ
Osaka
Yu-suke
YS
Casal
Enriqueta
E
Lee
Yuan C
YC
Kakehi
Kazuaki
K
D000818:Animals; D002240:Carbohydrate Sequence; D002364:Caseins / Q000032:analysis; D002417:Cattle; D002851:Chromatography, High Pressure Liquid; D003126:Colostrum / Q000737:chemistry; D019075:Electrophoresis, Capillary; D005260:Female; D005753:Gastric Mucosa / Q000737:chemistry; D006041:Goats; D006801:Humans; D007070:Immunoglobulin A / Q000032:analysis; D009077:Mucins / Q000032:analysis* / Q000737:chemistry; D010446:Peptide Fragments / Q000032:analysis; D011134:Polysaccharides / Q000032:analysis* / Q000737:chemistry; D011247:Pregnancy; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D012756:Sheep; D013045:Species Specificity; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013363:Submandibular Gland / Q000737:chemistry; D013552:Swine; D013997:Time Factors; D000509:alpha-Fetoproteins / Q000032:analysis; D062367:ortho-Aminobenzoates / Q000737:chemistry
D002364:Caseins; D007070:Immunoglobulin A; D009077:Mucins; D010446:Peptide Fragments; D011134:Polysaccharides; D000509:alpha-Fetoproteins; C065989:caseinomacropeptide; D062367:ortho-Aminobenzoates; C031385:anthranilic acid