Capillary electrophoresis of sialic acid-containing glycoprotein. Effect of the heterogeneity of carbohydrate chains on glycoform separation using an alpha1-acid glycoprotein as a model.
alpha1-Acid glycoprotein (AGP) showed multiple peaks on separation using capillary electrophoresis in a chemically modified capillary with dimethylpolysiloxane at slightly acidic conditions. We analyzed glycoforms of AGP species after separation by ion-exchange chromatography, Con A affinity chromatography, and Cu(II)-chelating affinity chromatography. The AGP species thus obtained were digested with N-glycosidase F, and the released carbohydrate chains were analyzed by high-performance liquid chromatography after labeling with 3-aminobenzoic acid. The results afforded basic information on the contribution of carbohydrate chains to the separation mechanism of glycoforms of AGP by capillary electrophoresis. In addition, we describe an easy method for AGP analysis in serum samples using the electrokinetic injection.
Kakehi
K
K
Faculty of Pharmaceutical Sciences, Kinki University, Higashi-Osaka, Japan. k_kakehi@phar.kindai.ac.jp
Kinoshita
M
M
Kawakami
D
D
Tanaka
J
J
Sei
K
K
Endo
K
K
Oda
Y
Y
Iwaki
M
M
Masuko
T
T
D002240:Carbohydrate Sequence; D002241:Carbohydrates / Q000737:chemistry*; D002853:Chromatography, Liquid / Q000379:methods; D019075:Electrophoresis, Capillary / Q000379:methods*; D006023:Glycoproteins / Q000097:blood* / Q000737:chemistry; D006801:Humans; D008956:Models, Chemical; D008969:Molecular Sequence Data; D019158:N-Acetylneuraminic Acid / Q000737:chemistry*; D009961:Orosomucoid / Q000032:analysis*; D020033:Protein Isoforms / Q000097:blood* / Q000737:chemistry; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
D002241:Carbohydrates; D006023:Glycoproteins; D009961:Orosomucoid; D020033:Protein Isoforms; D019158:N-Acetylneuraminic Acid