A model system for convenient fluorescent labeling of sugar chain in taka-amylase A.
A convenient detection of sugar chains in Taka-amylase A (TAA) was done by using 40 micrograms of enzyme, where a decrease in the UV absorption of NaIO4 during the periodate oxidation reaction was monitored. The periodate-oxidized sugar chain was labeled with a fluorescent reagent, N-1-ethylenediaminonaphthalene (EDAN), by incubation at pH 9.5 and 30 degrees C for 1 h. The excess EDAN was removed by either quenching with o-phthaladehyde or Bio-Gel P-2 gel adsorption. Among the peptide fragments prepared from the EDAN-labeled TAA, a fluorescent peptide corresponding to the sugar chain was distinguished by the ODS column. These results suggest that periodate oxidation and subsequent fluorescent labeling were useful for the sensitive analysis of various glycoprotein samples.
Kobayashi
M
M
National Food Research Institute, Tsukuba, Japan.
Sasaki
Y
Y
Chiba
Y
Y
D002241:Carbohydrates / Q000032:analysis*; D005029:Ethylenediamines; D005456:Fluorescent Dyes; D010084:Oxidation-Reduction; D010504:Periodic Acid / Q000737:chemistry; D012680:Sensitivity and Specificity; D013056:Spectrophotometry, Ultraviolet; D000516:alpha-Amylases / Q000032:analysis*
D002241:Carbohydrates; D005029:Ethylenediamines; D005456:Fluorescent Dyes; C008588:N-(1-naphthyl)ethylenediamine; D010504:Periodic Acid; D000516:alpha-Amylases