Jove
Visualize
Contact Us
JoVE
x logofacebook logolinkedin logoyoutube logo
ABOUT JoVE
OverviewLeadershipBlogJoVE Help Center
AUTHORS
Publishing ProcessEditorial BoardScope & PoliciesPeer ReviewFAQSubmit
LIBRARIANS
TestimonialsSubscriptionsAccessResourcesLibrary Advisory BoardFAQ
RESEARCH
JoVE JournalMethods CollectionsJoVE Encyclopedia of ExperimentsArchive
EDUCATION
JoVE CoreJoVE BusinessJoVE Science EducationJoVE Lab ManualFaculty Resource CenterFaculty Site
Terms & Conditions of Use
Privacy Policy
Policies

Related Concept Videos

You might also read

Related Articles

Articles linked to this work by shared authors, journal, and citation graph.

Sort by
Same author

Optimized detection of caspase-6 activation in a murine inflammation model to inform neurodegenerative disease therapies.

PloS one·2026
Same author

Incorporation of Functional Proteins on Cellular Surfaces via Artificial Cell-Derived Vesicles (ACDVs) for Plasma Membrane Reprogramming.

Journal of the American Chemical Society·2026
Same author

Polymeric Lysosome-Targeting Chimeras (PolyTACs): Extracellular Targeted Protein Degradation without Co-Opting Lysosome-Targeting Receptors.

Journal of the American Chemical Society·2026
Same author

Complex Coacervate Emulsions as a Strategy to Stabilize Enzymes for Catalysis in Organic Solvents.

ACS macro letters·2025
Same author

HER2 Antibody-Nanogel Conjugates: A High-Capacity, Antibody-Sparing Alternative to Antibody-Drug Conjugates.

Nano letters·2025
Same author

Inhibition of Zika Virus Protease by Modulating NS2B-NS3 Interactions.

Biochemistry·2025
Same journal

Macromolecular crowding inhibits degradation of alpha-synuclein amyloid fibrils induced by cathepsins and MMP9.

Protein science : a publication of the Protein Society·2026
Same journal

Sequence-encoded differences in the conformational ensembles of CITED transcriptional activation domains impact coactivator binding.

Protein science : a publication of the Protein Society·2026
Same journal

The phospholipid biosynthesis enzyme PlsB contains three distinct domains for membrane association, lysophosphatidic acid synthesis, and dimerization.

Protein science : a publication of the Protein Society·2026
Same journal

Structural basis of ligand selectivity in FAD/NAD(P)H-dependent dehydrogenases: insights from trypanothione reductase and type II NADH dehydrogenase.

Protein science : a publication of the Protein Society·2026
Same journal

Achieving protease substrate-specific inhibition by mAb dual functional selections.

Protein science : a publication of the Protein Society·2026
Same journal

How important are quantum mechanical effects in controlling biological functions: Enzymes, electron transfer and bird navigation.

Protein science : a publication of the Protein Society·2026
See all related articles

Related Experiment Video

Updated: Dec 1, 2025

Lighting Up the Pathways to Caspase Activation Using Bimolecular Fluorescence Complementation
08:47

Lighting Up the Pathways to Caspase Activation Using Bimolecular Fluorescence Complementation

Published on: March 5, 2018

9.3K

Tracking exogenous intracellular casp-3 using split GFP.

Francesca Anson1, Pintu Kanjilal1, S Thayumanavan1,2

  • 1Department of Chemistry, University of Massachusetts, Amherst, Massachusetts, USA.

Protein Science : a Publication of the Protein Society
|November 9, 2020
PubMed
Summary
This summary is machine-generated.

Researchers developed a novel fluorescent tag for caspase-3 (a protease) that only activates in the cytosol, enabling precise tracking of protein delivery and avoiding issues with endosomal entrapment. This technology aids in studying protein therapeutics and cellular processes.

Keywords:
apoptosiscaspasecaspase-3intracellular protein deliverynanogelsplit GFP

More Related Videos

In Vivo Biosensor Tracks Non-apoptotic Caspase Activity in Drosophila
13:21

In Vivo Biosensor Tracks Non-apoptotic Caspase Activity in Drosophila

Published on: November 27, 2016

9.4K
Detecting Anastasis In Vivo by CaspaseTracker Biosensor
20:16

Detecting Anastasis In Vivo by CaspaseTracker Biosensor

Published on: February 1, 2018

9.6K

Related Experiment Videos

Last Updated: Dec 1, 2025

Lighting Up the Pathways to Caspase Activation Using Bimolecular Fluorescence Complementation
08:47

Lighting Up the Pathways to Caspase Activation Using Bimolecular Fluorescence Complementation

Published on: March 5, 2018

9.3K
In Vivo Biosensor Tracks Non-apoptotic Caspase Activity in Drosophila
13:21

In Vivo Biosensor Tracks Non-apoptotic Caspase Activity in Drosophila

Published on: November 27, 2016

9.4K
Detecting Anastasis In Vivo by CaspaseTracker Biosensor
20:16

Detecting Anastasis In Vivo by CaspaseTracker Biosensor

Published on: February 1, 2018

9.6K

Area of Science:

  • Biotechnology and Molecular Biology
  • Cellular and Subcellular Delivery Systems
  • Protein Engineering and Imaging

Background:

  • Effective cytosolic protein delivery is crucial for therapeutics, genetic modification, and research tools.
  • Current fluorescent tagging methods often cause non-specific interactions, high background noise, or alter protein structure.
  • Distinguishing cytosolic from endosomally entrapped proteins is challenging due to fluorophore properties and cargo complexity, especially for proteases prone to self-proteolysis.

Purpose of the Study:

  • To develop a functionally silent and reliable tagging technology for proteases, specifically caspase-3.
  • To create a method for sensitive visualization and accurate localization of cytosolic protein delivery.
  • To enable reliable differentiation between cytosolic and endosomal protein localization and track cargo translocation.

Main Methods:

  • Engineered a caspase-3 variant fused with the 11th strand of Green Fluorescent Protein (GFP).
  • This tagged caspase-3 remains enzymatically active and structurally similar to wild-type caspase-3.
  • Fluorescence is generated only when the tagged caspase-3 encounters cytosolic GFP strands 1-10, signaling non-endosomal localization.

Main Results:

  • The developed tagged caspase-3 variant retains wild-type enzymatic activity and structural integrity.
  • The system effectively signals non-endosomal localization with low false positive rates.
  • Demonstrated the ability to track the translocation of the tagged protease cargo within the cell.

Conclusions:

  • The tagged caspase-3 cytosolic reporter provides a reliable method for screening cytosolic versus endosomally-entrapped proteins.
  • This technology facilitates the study of protein delivery, charge-property relationships in cellular escape, and protease therapeutic applications.
  • Lays the groundwork for precise investigations into the number of caspases required for inducing apoptotic cell death.