Thioester‐Based Coupled Fluorogenic Assays in Microdevice for the Detection of Single‐Molecule Enzyme Activities of Esterases with Specified Substrate Recognition

Single‐molecule enzyme activity assay is a platform that enables the analysis of enzyme activities at single proteoform level. The limitation of the targetable enzymes is the major drawback of the assay, but the general assay platform is reported to study single‐molecule enzyme activities of esteras...

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Bibliographic Details
Published in:Advanced science 2024-03, Vol.11 (10), p.e2306559-n/a
Main Authors: Ukegawa, Tatsuya, Komatsu, Toru, Minoda, Mayano, Matsumoto, Takuya, Iwasaka, Takumi, Mizuno, Tadahaya, Tachibana, Ryo, Sakamoto, Shingo, Hanaoka, Kenjiro, Kusuhara, Hiroyuki, Honda, Kazufumi, Watanabe, Rikiya, Urano, Yasuteru
Format: Article
Language:English
Subjects:
Acids
chemical biology
Design
Enzymes
enzymomics
Esterases - analysis
Esterases - chemistry
Proteins
single‐molecule analysis
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Summary:Single‐molecule enzyme activity assay is a platform that enables the analysis of enzyme activities at single proteoform level. The limitation of the targetable enzymes is the major drawback of the assay, but the general assay platform is reported to study single‐molecule enzyme activities of esterases based on the coupled assay using thioesters as substrate analogues. The coupled assay is realized by developing highly water‐soluble thiol‐reacting probes based on phosphonate‐substituted boron dipyrromethene (BODIPY). The system enables the detection of cholinesterase activities in blood samples at single‐molecule level, and it is shown that the dissecting alterations of single‐molecule esterase activities can serve as an informative platform for activity‐based diagnosis. The single‐molecule enzyme activity assay platform is reported for esterases with strict substrate recognition utilizing the coupled assay using thioesters as substrate analogues. The highly water‐soluble thiol‐reacting probes are developed based on phosphonate‐substituted boron dipyrromethene (BODIPY), which enables the detection of cholinesterase activities in blood samples at single‐molecule level for activity‐based diagnosis.
ISSN:2198-3844
2198-3844
DOI:10.1002/advs.202306559