A fluorometric method for the assay of protein kinase activity

Protein kinases constitute one of the largest protein families in nature. Current methods to assay their activity involve the use of radioactive ATP or very expensive reagents. In this work, we developed a highly sensitive, cost-effective and straightforward protocol to measure protein kinase activi...

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Detalles Bibliográficos
Publicado: 2018
Materias:
Enzyme kinetics
Non-radioactive assay
Phosphorylation
Protein kinase
lactate dehydrogenase
protein kinase
pyruvate kinase
Article
enzyme activity
fluorometry
kinetic parameters
limit of detection
potato
priority journal
substrate concentration
Acceso en línea:https://bibliotecadigital.exactas.uba.ar/collection/paper/document/paper_00032697_v557_n_p120_Rojas
http://hdl.handle.net/20.500.12110/paper_00032697_v557_n_p120_Rojas
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Biblioteca Digital - Facultad de Ciencias Exactas y Naturales (UBA) de Universidad de Buenos Aires
id paper:paper_00032697_v557_n_p120_Rojas
record_format dspace
spelling paper:paper_00032697_v557_n_p120_Rojas2023-06-08T14:24:06Z A fluorometric method for the assay of protein kinase activity Enzyme kinetics Non-radioactive assay Phosphorylation Protein kinase lactate dehydrogenase protein kinase pyruvate kinase Article enzyme activity fluorometry kinetic parameters limit of detection potato priority journal substrate concentration Protein kinases constitute one of the largest protein families in nature. Current methods to assay their activity involve the use of radioactive ATP or very expensive reagents. In this work, we developed a highly sensitive, cost-effective and straightforward protocol to measure protein kinase activity using a microplate layout. Released ADP is converted into NAD+, which is quantified by its fluorescent properties after alkaline treatment (linear range 0–10 nmol ADP). To validate our protocol, we characterized a recombinant calcium-dependent protein kinase from potato. Overall, this tool represents a critical step forward in the functional characterization of protein kinases. © 2018 Elsevier Inc. 2018 https://bibliotecadigital.exactas.uba.ar/collection/paper/document/paper_00032697_v557_n_p120_Rojas http://hdl.handle.net/20.500.12110/paper_00032697_v557_n_p120_Rojas
institution Universidad de Buenos Aires
institution_str I-28
repository_str R-134
collection Biblioteca Digital - Facultad de Ciencias Exactas y Naturales (UBA)
topic Enzyme kinetics
Non-radioactive assay
Phosphorylation
Protein kinase
lactate dehydrogenase
protein kinase
pyruvate kinase
Article
enzyme activity
fluorometry
kinetic parameters
limit of detection
potato
priority journal
substrate concentration
spellingShingle Enzyme kinetics
Non-radioactive assay
Phosphorylation
Protein kinase
lactate dehydrogenase
protein kinase
pyruvate kinase
Article
enzyme activity
fluorometry
kinetic parameters
limit of detection
potato
priority journal
substrate concentration
A fluorometric method for the assay of protein kinase activity
topic_facet Enzyme kinetics
Non-radioactive assay
Phosphorylation
Protein kinase
lactate dehydrogenase
protein kinase
pyruvate kinase
Article
enzyme activity
fluorometry
kinetic parameters
limit of detection
potato
priority journal
substrate concentration
description Protein kinases constitute one of the largest protein families in nature. Current methods to assay their activity involve the use of radioactive ATP or very expensive reagents. In this work, we developed a highly sensitive, cost-effective and straightforward protocol to measure protein kinase activity using a microplate layout. Released ADP is converted into NAD+, which is quantified by its fluorescent properties after alkaline treatment (linear range 0–10 nmol ADP). To validate our protocol, we characterized a recombinant calcium-dependent protein kinase from potato. Overall, this tool represents a critical step forward in the functional characterization of protein kinases. © 2018 Elsevier Inc.
title A fluorometric method for the assay of protein kinase activity
title_short A fluorometric method for the assay of protein kinase activity
title_full A fluorometric method for the assay of protein kinase activity
title_fullStr A fluorometric method for the assay of protein kinase activity
title_full_unstemmed A fluorometric method for the assay of protein kinase activity
title_sort fluorometric method for the assay of protein kinase activity
publishDate 2018
url https://bibliotecadigital.exactas.uba.ar/collection/paper/document/paper_00032697_v557_n_p120_Rojas
http://hdl.handle.net/20.500.12110/paper_00032697_v557_n_p120_Rojas
_version_ 1768545717522006016

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