Synthesis of a fluorescent probe for measuring the activity of endo-ß-N-acetylglucosaminidases recognizing hybrid-type N-glycans.
Bioorg Med Chem
; 100: 117612, 2024 Feb 15.
Article
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| MEDLINE
| ID: mdl-38290307
ABSTRACT
A fluorescence-quenching-based assay system was constructed to determine the hydrolytic activity of endo-ß-N-acetylglucosaminidases (ENGases) interacting with hybrid-type N-glycans. This was achieved using a dual-labeled fluorescent probe with a nonasaccharide structure. We produced the nonasaccharide skeleton by the stepwise glycosylation of the galactose residue on a galactosyl chitobiose derivative. Next, we introduced azido and acetoxy groups into the nonasaccharide derivative in a stepwise manner, which led to stereochemistry inversion at both the C-4 and C-2 hydroxy groups on its galactose residue. The protecting groups of the resulting nonasaccharide derivative were removed, and the derivative was labeled with an N-methylanthraniloyl group to obtain a reporter dye and a 2,4-dinitrophenyl group as a quenching molecule to obtain target probe 1. The use of this probe along with a microplate reader enabled a facile evaluation of the hydrolytic activities of ENGases Endo-H, Endo-M, Endo-F3, Endo-S, and Endo-CC. Furthermore, this probe could also assist in the search for novel ENGases that are specific to hybrid-type N-glycans.
Palabras clave
Texto completo:
1
Banco de datos:
MEDLINE
Asunto principal:
Acetilglucosaminidasa
/
Colorantes Fluorescentes
Idioma:
En
Año:
2024
Tipo del documento:
Article