A sensitive assay of sialidase activity with fluorogenic oligosaccharide derivatives.
Fluorogenic oligosaccharide derivatives, lactityl-aminopyridine and sialyllactityl-aminopyridines, were synthesized by reductive amination of lactose and sialyllactose with 2-aminopyridine in the presence of sodium cyanoborohydride. Lactityl-aminopyridine showed bluish-white fluorescence at 390 nm by excitation with ultraviolet light at 320 nm, and the intensity of the fluorescence was proportional to the concentration of lactityl-aminopyridine within a range from 20 to 9800 pmole/ml. Two isomers of sialyllactityl-aminopyridine were enzymatically hydrolyzed to sialic acid and lactityl-aminopyridine by the action of both microbial and mammalian sialidases, and their suitability as substrates for sialidase was investigated. The result showed that sialidase activity was determined with about 55-fold higher sensitivity than that of ordinarily used colorimetric methods.
D000631:Aminopyridines / Q000138:chemical synthesis / Q000378:metabolism*; D005456:Fluorescent Dyes / Q000378:metabolism; D008247:Lysosomes / Q000201:enzymology; D009439:Neuraminidase / Q000032:analysis* / Q000378:metabolism; D009844:Oligosaccharides / Q000378:metabolism*; D013050:Spectrometry, Fluorescence
D000631:Aminopyridines; D005456:Fluorescent Dyes; D009844:Oligosaccharides; C037629:2-(N-3'-sialyllactityl)aminopyridine; C037628:2-(N-lactityl)aminopyridine; C037630:2-(N-6'-sialyllactityl)aminopyridine; D009439:Neuraminidase