Thiol-ene-enabled detection of thiophosphorylation as a labeling strategy for phosphoproteins

Kaelyn E. Wilke; Erin E. Carlson

doi:10.1007/978-1-4939-3049-4_1,

Thiol-ene-enabled detection of thiophosphorylation as a labeling strategy for phosphoproteins

Kaelyn E. Wilke, Erin E. Carlson

Chemistry (Twin Cities)

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Abstract

The adenosine triphosphate (ATP) analogue adenosine 5′-O-(3-thiotriphosphate) (ATPγS) has been applied as a tool to study kinase-substrate phosphorylation. Not only does the transfer of a thiophosphate group represent a unique modification amid the phosphoproteome, but it can also be stable to phosphatase activity. However, detection of this species is complicated due to the similar chemical reactivity of thiophosphate and proteinaceous thiols. Here, we describe a novel method for detection of protein thiophosphorylation utilizing the thiol-ene reaction. By first chemoselectively capping protein thiols through radical chemistry, kinase-catalyzed thiophosphorylation can be visualized specifically.

Original language	English (US)
Title of host publication	Methods in Molecular Biology
Publisher	Humana Press Inc.
Pages	3-15
Number of pages	13
DOIs	https://doi.org/10.1007/978-1-4939-3049-4_1,
State	Published - Jan 1 2016

Publication series

Name	Methods in Molecular Biology
Volume	1355
ISSN (Print)	1064-3745

Bibliographical note

Publisher Copyright:
© Springer Science+Business Media New York 2016.

Keywords

ATPγS
Kinase
Phosphoprotein
Substrate
Thiol capping
Thiol-ene
Thiophosphorylation

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AU - Carlson, Erin E.

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N2 - The adenosine triphosphate (ATP) analogue adenosine 5′-O-(3-thiotriphosphate) (ATPγS) has been applied as a tool to study kinase-substrate phosphorylation. Not only does the transfer of a thiophosphate group represent a unique modification amid the phosphoproteome, but it can also be stable to phosphatase activity. However, detection of this species is complicated due to the similar chemical reactivity of thiophosphate and proteinaceous thiols. Here, we describe a novel method for detection of protein thiophosphorylation utilizing the thiol-ene reaction. By first chemoselectively capping protein thiols through radical chemistry, kinase-catalyzed thiophosphorylation can be visualized specifically.

AB - The adenosine triphosphate (ATP) analogue adenosine 5′-O-(3-thiotriphosphate) (ATPγS) has been applied as a tool to study kinase-substrate phosphorylation. Not only does the transfer of a thiophosphate group represent a unique modification amid the phosphoproteome, but it can also be stable to phosphatase activity. However, detection of this species is complicated due to the similar chemical reactivity of thiophosphate and proteinaceous thiols. Here, we describe a novel method for detection of protein thiophosphorylation utilizing the thiol-ene reaction. By first chemoselectively capping protein thiols through radical chemistry, kinase-catalyzed thiophosphorylation can be visualized specifically.

KW - ATPγS

KW - Kinase

KW - Phosphoprotein

KW - Substrate

KW - Thiol capping

KW - Thiol-ene

KW - Thiophosphorylation

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Thiol-ene-enabled detection of thiophosphorylation as a labeling strategy for phosphoproteins

Abstract

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