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

Caspases01:24

Caspases

Caspase, a family of cysteine proteases, serve as effectors in apoptosis. The ced3 gene in C.elegans was first identified to be involved in apoptosis. This gene encodes the ced-3 caspase that is similar to the interleukin-1-beta converting enzyme or ICE in mammals. In addition to apoptosis, caspases also function in the inflammatory response. Inflammatory caspases are essential in activating pro-inflammatory cytokines that recruit immune cells and block the replication of pathogens inside cells.
The Extrinsic Apoptotic Pathway01:17

The Extrinsic Apoptotic Pathway

The extrinsic apoptotic pathway is initiated when extracellular death-inducing signals, such as specific cytokines, activate the death receptors expressed on the cell surface. The immune cells involved in this pathway are natural killer cells (NK cells) and cytotoxic T-lymphocytes. NK cells are critical in innate immune response, while cytotoxic T-lymphocytes are associated with adaptive immune response. These cells recognize specific receptors expressed on the altered cells and activate...
The Intrinsic Apoptotic Pathway01:31

The Intrinsic Apoptotic Pathway

Internal cellular stress, such as cellular injury or hypoxia, triggers intrinsic apoptosis. The B-cell lymphoma 2 (Bcl-2) family of proteins are the primary regulators of the intrinsic apoptotic pathway. For example, during DNA damage, checkpoint proteins, such as Ataxia Telangiectasia Mutated (ATM protein) and Checkpoints Factor-2 (Chk2) proteins, are activated. These proteins phosphorylate p53 which further activates pro-apoptotic proteins, such as Bax, Bak, PUMA, and Noxa, and inhibits...
Mechanism of Lamellipodia Formation01:31

Mechanism of Lamellipodia Formation

Cells migrating in response to external stimuli form lamellipodia, which are thin membrane protrusions supported by a mesh of linked, branched, or unbranched actin filaments. These actin filaments interact with myosin motor proteins, creating the dynamic actomyosin complex within the cytoskeleton. Contractility, or the ability to generate contractile stress, is inherent to the actomyosin complex. It helps cells detect the stiffness of the surrounding ECM and exert contractile force for...
Allosteric Proteins-ATCase01:19

Allosteric Proteins-ATCase

Binding sites linkages can regulate a protein's function.  For example, enzyme activity is often regulated through a feedback mechanism where the end product of the biochemical process serves as an inhibitor.
Aspartate transcarbamoylase (ATCase) is a cytosolic enzyme that catalyzes the condensation of L-aspartate and carbamoyl phosphate to  N-carbamoyl-L-aspartate. This reaction is the first step in pyrimidine biosynthesis. UTP and CTP, the end products of the pyrimidine synthesis pathway,...
cAMP-dependent Protein Kinase Pathways01:25

cAMP-dependent Protein Kinase Pathways

Cyclic Adenosine Monophosphate (cAMP) is an essential second messenger that activates protein kinase A (PKA) and regulates various biological processes. A single epinephrine molecule binds to GPCR and activates several heterotrimeric G proteins, each stimulating multiple adenylyl cyclase, amplifying the signal, and synthesizing large numbers of cAMP molecules. Small changes in cAMP concentration affect PKA activity. The binding of four cAMP molecules induces a conformational change in PKA,...

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

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

Biochemistry·2025
Same author

Orthogonal RNA-regulated destabilization domains for three-color RNA imaging with minimal RNA perturbation.

Nature methods·2025
Same author

An NMR sample preparation case study: Considerations for the self-destructive protease caspase-6.

PloS one·2025
Same author

A Robust Fluorogenic Substrate for Chikungunya Virus Protease (nsP2) Activity.

bioRxiv : the preprint server for biology·2025
Same author

A robust fluorogenic substrate for chikungunya virus protease (nsP2) activity.

Protein science : a publication of the Protein Society·2025

Related Experiment Video

Updated: Jun 22, 2026

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

L2' loop is critical for caspase-7 active site formation.

Witold A Witkowski1, Jeanne A Hardy

  • 1Department of Chemistry, University of Massachusetts Amherst, Amherst, Massachusetts 01003, USA.

Protein Science : a Publication of the Protein Society
|June 17, 2009
PubMed
Summary

The L2' loop is crucial for caspase-7 enzyme stability and catalytic function. Specific mutations reveal how this loop facilitates substrate binding and enzyme activation, essential for controlling caspase activity.

More Related Videos

Exploring Caspase Mutations and Post-Translational Modification by Molecular Modeling Approaches
05:56

Exploring Caspase Mutations and Post-Translational Modification by Molecular Modeling Approaches

Published on: October 13, 2022

Measuring Composition of CD95 Death-Inducing Signaling Complex and Processing of Procaspase-8 in this Complex
07:17

Measuring Composition of CD95 Death-Inducing Signaling Complex and Processing of Procaspase-8 in this Complex

Published on: August 2, 2021

Related Experiment Videos

Last Updated: Jun 22, 2026

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

Exploring Caspase Mutations and Post-Translational Modification by Molecular Modeling Approaches
05:56

Exploring Caspase Mutations and Post-Translational Modification by Molecular Modeling Approaches

Published on: October 13, 2022

Measuring Composition of CD95 Death-Inducing Signaling Complex and Processing of Procaspase-8 in this Complex
07:17

Measuring Composition of CD95 Death-Inducing Signaling Complex and Processing of Procaspase-8 in this Complex

Published on: August 2, 2021

Area of Science:

  • Biochemistry
  • Molecular Biology
  • Enzymology

Background:

  • Caspase active sites feature four mobile loops involved in substrate binding.
  • The L2' loop plays a key role in forming the active-site loop bundle and maintaining inactive enzyme conformation via cross-dimer hydrogen bonds.

Purpose of the Study:

  • To investigate the role of L2' loop residues in caspase-7 catalytic function and enzyme stability.
  • To elucidate the structural and functional differences between active and inactive caspase conformations.

Main Methods:

  • Site-directed mutagenesis of L2' loop residues.
  • Enzyme activity assays.
  • Analysis of crystal structures of wild-type and mutant caspases.

Main Results:

  • Active site binding significantly stabilizes the caspase complex.
  • Mutation P214A shows conformational flexibility, being unstable ligand-free but stable when bound to substrate.
  • Residues K212 and I213 are essential for substrate binding and catalysis; mutation I213A leads to conformational rearrangement to accommodate substrate.

Conclusions:

  • The L2' loop is critical for stabilizing the substrate-binding groove and achieving native catalytic efficiency, despite not directly contacting the substrate.
  • Caspase-7 exists in distinct conformations in the absence and presence of substrate, regulated by the L2' loop.
  • Understanding L2' loop dynamics is key to controlling caspase function.