Measuring Diacylglycerol Kinase-θ Activity and Binding

B. Tu-Sekine; D. M. Raben

doi:10.1016/bs.mie.2016.09.002

Measuring Diacylglycerol Kinase-θ Activity and Binding

B. Tu-Sekine, D. M. Raben

School of Medicine

Research output: Chapter in Book/Report/Conference proceeding › Chapter

Abstract

This section provides detailed protocols for the analysis of a mammalian diacylglycerol kinase, DGKθ, including an activity assay, a kinetic analysis, preparation of small unilamellar vesicles, and a vesicle pulldown assay. The goal of this section is to provide an overview of the unique challenges inherent in the study of an interfacial enzyme such as DGKθ and to outline methods useful for analysis. We include a short tutorial on selecting lipids for forming the interface since this is critical for a successful in vitro assay, and lipids are important regulators of this enzyme. The general principles can be applied to the study of other interfacial enzymes.

Original language	English (US)
Title of host publication	Methods in Enzymology
Publisher	Academic Press Inc.
Pages	231-253
Number of pages	23
DOIs	https://doi.org/10.1016/bs.mie.2016.09.002
State	Published - 2017

Publication series

Name	Methods in Enzymology
Volume	583
ISSN (Print)	0076-6879
ISSN (Electronic)	1557-7988

Keywords

Diacylglycerol kinase
Interfacial binding
Interfacial enzymology
Liposomes

ASJC Scopus subject areas

Biochemistry
Molecular Biology

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10.1016/bs.mie.2016.09.002

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title = "Measuring Diacylglycerol Kinase-θ Activity and Binding",

abstract = "This section provides detailed protocols for the analysis of a mammalian diacylglycerol kinase, DGKθ, including an activity assay, a kinetic analysis, preparation of small unilamellar vesicles, and a vesicle pulldown assay. The goal of this section is to provide an overview of the unique challenges inherent in the study of an interfacial enzyme such as DGKθ and to outline methods useful for analysis. We include a short tutorial on selecting lipids for forming the interface since this is critical for a successful in vitro assay, and lipids are important regulators of this enzyme. The general principles can be applied to the study of other interfacial enzymes.",

keywords = "Diacylglycerol kinase, Interfacial binding, Interfacial enzymology, Liposomes",

author = "B. Tu-Sekine and Raben, {D. M.}",

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AB - This section provides detailed protocols for the analysis of a mammalian diacylglycerol kinase, DGKθ, including an activity assay, a kinetic analysis, preparation of small unilamellar vesicles, and a vesicle pulldown assay. The goal of this section is to provide an overview of the unique challenges inherent in the study of an interfacial enzyme such as DGKθ and to outline methods useful for analysis. We include a short tutorial on selecting lipids for forming the interface since this is critical for a successful in vitro assay, and lipids are important regulators of this enzyme. The general principles can be applied to the study of other interfacial enzymes.

KW - Diacylglycerol kinase

KW - Interfacial binding

KW - Interfacial enzymology

KW - Liposomes

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BT - Methods in Enzymology

PB - Academic Press Inc.

ER -

Measuring Diacylglycerol Kinase-θ Activity and Binding

Abstract

Publication series

Keywords

ASJC Scopus subject areas

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