Open AccessDevelopment of Fluorogenic Substrates of α-l -Fucosidase Useful for Inhibitor Screening and Gene-expression Profiling
Author(s) -
Kazuki Miura,
Takumi Tsukagoshi,
Tetsufumi Hirano,
Takeshi Nishio,
Wataru Hakamata
Publication year - 2019
Publication title -
acs medicinal chemistry letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.065
H-Index - 66
ISSN - 1948-5875
DOI - 10.1021/acsmedchemlett.9b00259
Subject(s) - fucosidase , hela , enzyme , biochemistry , chemistry , cell culture , glycolipid , glycoprotein , cell , microbiology and biotechnology , biology , fucose , genetics
Inhibitors of human α-l-fucosidases, tissue α-l-fucosidase (tFuc), and plasma α-l-fucosidase reportedly play roles in multiple diseases, suggesting their therapeutic potential for gastric disease associated with Helicobacter pylori and fucosidosis. Terminal fucose linkages on glycoproteins and glycolipids are a natural substrate for both enzymes; however, there are currently no fluorogenic substrates allowing their cellular evaluation. Here, we described the development of novel three-color fluorogenic substrates for lysosome-localized tFuc that exhibited excellent specificity and sensitivity in three human cell lines. Additionally, we developed a cell-based high-throughput inhibitor screening system in a 96-well format and a cell-based inhibitory activity evaluation system in a 6-well format for tFuc inhibitors using this substrate, which allowed accurate quantification of the inhibition rate. Moreover, analysis of significant changes in gene expression resulting from 30% inhibition of tFuc in HeLa cells revealed potential roles in gastric disease.