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Related Concept Videos

Protein Dynamics in Living Cells01:19

Protein Dynamics in Living Cells

Different fluorescence-based techniques are used to study the protein dynamics in living cells. These techniques include FRAP, FRET, and PET.
Fluorescent recovery after photobleaching (FRAP) is a fluorescent-protein-based detection technique used to quantify protein movement rates within the cell. This method exposes a small portion of the cell to an intense laser beam. The laser beam causes permanent photobleaching of the fluorophore-tagged proteins in the exposed region. As the bleached...

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Related Experiment Video

Updated: Jun 11, 2026

Assaying Protein Kinase Activity with Radiolabeled ATP
08:05

Assaying Protein Kinase Activity with Radiolabeled ATP

Published on: May 26, 2017

Real-time fluorogenic kinase assay using protein as substrate.

Hongye Sun1, Karen Mitchell, Linda Lee

  • 1Life Technologies Corporation, Foster City, CA 94404, USA. hongye.sun@lifetech.com

Analytical Biochemistry
|July 6, 2010
PubMed
Summary
This summary is machine-generated.

A novel real-time fluorogenic kinase assay uses myelin basic protein (MBP) and a dye-labeled lipopeptide. Phosphorylation by kinases alters MBP

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Characterization at the Molecular Level using Robust Biochemical Approaches of a New Kinase Protein
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Characterization at the Molecular Level using Robust Biochemical Approaches of a New Kinase Protein

Published on: June 30, 2019

Related Experiment Videos

Last Updated: Jun 11, 2026

Assaying Protein Kinase Activity with Radiolabeled ATP
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Published on: May 26, 2017

Visualizing Protein Kinase A Activity In Head-fixed Behaving Mice Using In Vivo Two-photon Fluorescence Lifetime Imaging Microscopy
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Characterization at the Molecular Level using Robust Biochemical Approaches of a New Kinase Protein
11:23

Characterization at the Molecular Level using Robust Biochemical Approaches of a New Kinase Protein

Published on: June 30, 2019

Area of Science:

  • Biochemistry
  • Molecular Biology
  • Enzyme Assays

Background:

  • Kinase assays are crucial for drug discovery and biological research.
  • Existing methods may lack real-time monitoring or require complex procedures.
  • Myelin basic protein (MBP) is a known substrate for various kinases.

Purpose of the Study:

  • To develop a novel, real-time fluorogenic kinase assay.
  • To utilize myelin basic protein (MBP) as a substrate in this assay.
  • To validate the assay's performance with known kinases and inhibitors.

Main Methods:

  • A noncovalent complex of MBP and a negatively charged, dye-labeled lipopeptide was formed.
  • The fluorescence of the complex was monitored in the presence of kinase and ATP.
  • Phosphorylation-induced changes in complex affinity and fluorescence were measured.

Main Results:

  • The assay demonstrated a fivefold fluorescence increase upon kinase activity.
  • Kinetic parameters (apparent K(m)(ATP)) were comparable to literature values.
  • Inhibitor constants for staurosporine were consistent with established data.

Conclusions:

  • The developed fluorogenic assay provides a sensitive and real-time method for studying kinase activity.
  • This assay is suitable for evaluating kinase kinetics and inhibitor potency.
  • The assay's reliability is confirmed by its performance with multiple kinases and a known inhibitor.