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Updated: Oct 11, 2025
Oligopeptide Competition Assay for Phosphorylation Site Determination
Published on: May 18, 2017
Discerning Tyrosine Phosphorylation from Multiple Phosphorylations Using a Nanofluidic Logic Platform
Yuting Xiong1,2, Minmin Li1,2, Wenqi Lu2
1Jiangxi Province Key Laboratory of Polymer Micro/Nano Manufacturing and Devices, School of Chemistry, Biology and Materials Science, East China University of Technology, 418 Guanglan Avenue, Nanchang 330013, P. R. China.
Abstract:
Discerning tyrosine phosphorylation (pTyr) catalyzed by Tyr kinase is central to the revelation of oncogenic mechanisms and the development of targeted anticancer drugs. Despite some techniques, this goal remains challenging, especially when faced with the interference of multiple phosphorylation events, including serine (pSer) and threonine phosphorylation (pThr). We describe here a functional polymer-modified artificial ion nanochannel, which enables the sensitive and selective recognition of phosphotyrosine (pY) peptide by the distinct ionic current change. Such a recognition effect allows for the nanochannel to work in a complex protein digest condition. Further, the implementation of nanofluidic logic functions with the addition of Ca2+ dramatically improves the selectivity of the nanochannel to pY peptide and thus can discern pTyr by the Tyr kinase from pSer by the Ser/Thr kinase through simultaneously monitoring multisite phosphorylation at the same or different peptide substrates in one-pot. This logic sensing platform displays the potential in differentiating Tyr kinase and Ser/Thr kinase and assessing multi-kinase activities in multi-targeted drug design.
Insights
This study introduces a novel nanochannel sensor for detecting tyrosine phosphorylation (pTyr). The technology distinguishes pTyr from other phosphorylation types, aiding cancer drug development.
Area of Science:
- Biochemistry
- Nanotechnology
- Analytical Chemistry
Background:
- Tyrosine phosphorylation (pTyr) is crucial for cancer development and targeted therapies.
- Distinguishing pTyr from serine (pSer) and threonine phosphorylation (pThr) is challenging due to complex phosphorylation events.
Purpose of the Study:
- To develop a sensitive and selective method for recognizing phosphotyrosine (pY) peptides.
- To create a nanofluidic platform capable of differentiating Tyr kinase from Ser/Thr kinase activity.
Main Methods:
- Utilized a functional polymer-modified artificial ion nanochannel for pY peptide recognition.
- Implemented nanofluidic logic functions with Ca2+ to enhance selectivity.
- Monitored multisite phosphorylation in a one-pot system.
Main Results:
- The nanochannel demonstrated sensitive and selective recognition of pY peptides via ionic current changes.
- The platform successfully differentiated Tyr kinase from Ser/Thr kinase activity.
- Simultaneous monitoring of multisite phosphorylation was achieved.
Conclusions:
- The developed logic sensing platform offers a promising approach for discerning Tyr kinase and Ser/Thr kinase.
- This technology has potential applications in assessing multi-kinase activities for multi-targeted drug design.
