Cell-permeable probe for identification and imaging of sialidases.
Alkyne-hinged 3-fluorosialyl fluoride (DFSA) containing an alkyne group was shown to be a mechanism-based target-specific irreversible inhibitor of sialidases. The ester-protected analog DFSA (PDFSA) is a membrane-permeable precursor of DFSA designed to be used in living cells, and it was shown to form covalent adducts with virus, bacteria, and human sialidases. The fluorosialyl-enzyme adduct can be ligated with an azide-annexed biotin via click reaction and detected by the streptavidin-specific reporting signals. Liquid chromatography-mass spectrometry/mass spectrometry analysis on the tryptic peptide fragments indicates that the 3-fluorosialyl moiety modifies tyrosine residues of the sialidases. DFSA was used to demonstrate influenza infection and the diagnosis of the viral susceptibility to the anti-influenza drug oseltamivir acid, whereas PDFSA was used for in situ imaging of the changes of sialidase activity in live cells.
Genomics Research Center,  Academia Sinica, Taipei 115,  Taiwan.
Charng-Sheng
CS
Tsai
Hsin-Yung
HY
Yen
Meng-I
MI
Lin
Tsung-I
TI
Tsai
Shi-Yun
SY
Wang
Wen-I
WI
Huang
Tsui-Ling
TL
Hsu
Yih-Shyun E
YS
Cheng
Jim-Min
JM
Fang
Chi-Huey
CH
Wong
D000480:Alkynes / Q000737:chemistry; D002853:Chromatography, Liquid; D057930:Click Chemistry / Q000379:methods*; D018736:DNA Adducts / Q000378:metabolism; D006801:Humans; D007251:Influenza, Human / Q000175:diagnosis; D015336:Molecular Probe Techniques; D015335:Molecular Probes / Q000737:chemistry*; D015394:Molecular Structure; D009439:Neuraminidase / Q000737:chemistry* / Q000378:metabolism / Q000648:ultrastructure*; D040901:Proteomics / Q000379:methods; D019809:Streptavidin / Q000737:chemistry; D053719:Tandem Mass Spectrometry
D000480:Alkynes; D018736:DNA Adducts; D015335:Molecular Probes; D019809:Streptavidin; D009439:Neuraminidase