Search research articles

ABOUT JoVE

Overview Leadership Blog JoVE Help Center

AUTHORS

Publishing Process Editorial Board Scope & Policies Peer Review FAQ Submit

LIBRARIANS

Testimonials Subscriptions Access Resources Library Advisory Board FAQ

RESEARCH

JoVE Journal Methods Collections JoVE Encyclopedia of Experiments Archive

EDUCATION

JoVE Core JoVE Business JoVE Science Education JoVE Lab Manual Faculty Resource Center Faculty Site

Terms & Conditions of Use

Related Concept Videos

Conservative Site-specific Recombination and Phase Variation

Conservative Site-specific Recombination and Phase Variation

Because the DNA segments are cut and reorganized in a direction-specific manner, site-specific recombination has emerged as an efficient genetic engineering technique. Flippase and Cyclization recombinases or Flp and Cre, respectively, are two members of the tyrosine recombinase family derived from bacteriophages, that are used to mediate site-specific DNA insertions, deletions, and targeted expression of proteins in mammalian cell lines.
The recognition sites for Cre recombinase called LoxP...

You might also read

Related Articles

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

Sort by

Same author

hexABC seeking the physical code of DNA.

Nature communications·2026

Same author

Prognostic impact of pleural effusion in acute heart failure and its link to diuretic therapy.

The American journal of medicine·2026

Same author

The expanding role of protease therapeutics (2012-2026): from replacement therapies to immune system modulation and beyond.

The Biochemical journal·2026

Same author

The NMR Exchange Format (NEF): Specification and Applications.

bioRxiv : the preprint server for biology·2026

Same author

Efficient exploration of peptide libraries using active learning with AlphaFold-based screening.

bioRxiv : the preprint server for biology·2026

Same author

Beyond Classical Force Fields: Physics-Driven Assessment of the Grappa Machine-Learned Force Field on the FoldBind Dataset.

Chemphyschem : a European journal of chemical physics and physical chemistry·2026

Same journal

Layered social competition coordinates reproductive hierarchy formation in ants.

bioRxiv : the preprint server for biology·2026

Same journal

Combination epigenetic-targeted therapy increases the immunogenicity of poorly immunogenic sarcomas.

bioRxiv : the preprint server for biology·2026

Same journal

Loss of LanC-like proteins delays post-injury regeneration of aging skeletal muscles.

bioRxiv : the preprint server for biology·2026

Same journal

Integrative Transfer Network: Deep Transfer Learning Across Populations and Prediction Targets.

bioRxiv : the preprint server for biology·2026

Same journal

Confidence-supported label-free metabolic imaging with FPhaS phase autofluorescence microscopy.

bioRxiv : the preprint server for biology·2026

Same journal

Sequence-encoded autoinhibition couples mRNA decapping activity to phase separation.

bioRxiv : the preprint server for biology·2026

See all related articles

Search research articles

Related Experiment Video

Updated: May 22, 2025

Fast Enzymatic Processing of Proteins for MS Detection with a Flow-through Microreactor

Fast Enzymatic Processing of Proteins for MS Detection with a Flow-through Microreactor

Published on: April 6, 2016

PERRC: Protease Engineering with Reactant Residence Time Control.

Sage Nelson¹, Jokent Gaza², Seyednima Ajayebi¹

¹Department of Chemical Engineering, University of Florida, Gainesville, 32611, USA.

Biorxiv : the Preprint Server for Biology

|March 17, 2025

Summary

This summary is machine-generated.

Researchers engineered a protease with enhanced specificity using Protease Engineering with Reactant Residence Time Control (PERRC). This method precisely controls evolution stringency, creating a new protease variant (TEVESNp) with a 65-fold preference for its target substrate.

Keywords:

High-Throughput Screening Protease Engineering Tobacco Etch Virus (TEV) protease Yeast Surface Display

More Related Videos

In Vitro Directed Evolution of a Restriction Endonuclease with More Stringent Specificity

In Vitro Directed Evolution of a Restriction Endonuclease with More Stringent Specificity

Published on: March 25, 2020

Time-resolved ElectroSpray Ionization Hydrogen-deuterium Exchange Mass Spectrometry for Studying Protein Structure and Dynamics

Time-resolved ElectroSpray Ionization Hydrogen-deuterium Exchange Mass Spectrometry for Studying Protein Structure and Dynamics

Published on: April 17, 2017

Related Experiment Videos

Last Updated: May 22, 2025

Fast Enzymatic Processing of Proteins for MS Detection with a Flow-through Microreactor

Fast Enzymatic Processing of Proteins for MS Detection with a Flow-through Microreactor

Published on: April 6, 2016

In Vitro Directed Evolution of a Restriction Endonuclease with More Stringent Specificity

In Vitro Directed Evolution of a Restriction Endonuclease with More Stringent Specificity

Published on: March 25, 2020

Time-resolved ElectroSpray Ionization Hydrogen-deuterium Exchange Mass Spectrometry for Studying Protein Structure and Dynamics

Time-resolved ElectroSpray Ionization Hydrogen-deuterium Exchange Mass Spectrometry for Studying Protein Structure and Dynamics

Published on: April 17, 2017

Area of Science:

Biotechnology
Molecular Biology
Protein Engineering

Background:

Proteases with engineered specificity are valuable for therapeutics and biotechnology.
Broad substrate specificity of natural proteases limits their applications.
Engineering protease specificity is challenging due to residual activity on native substrates.

Purpose of the Study:

To develop a novel platform for engineering protease specificity.
To create an orthogonal protease variant with high substrate selectivity.
To demonstrate the utility of the platform in constructing protein circuits.

Main Methods:

Developed Protease Engineering with Reactant Residence Time Control (PERRC) platform.
PERRC utilizes endoplasmic reticulum (ER) retention sequence strength and residence time.
Adjusted counterselection to selection substrate ratios for precise evolution control.

Main Results:

Evolved an orthogonal tobacco etch virus protease variant (TEVESNp).
TEVESNp shows a 65-fold preference for a single-amino-acid-altered substrate (ENLYFES) over its parent (ENLYFQS).
Demonstrated TEVESNp's function in bacterial orthogonal protein circuit construction.

Conclusions:

PERRC enables precise engineering of protease specificity.
TEVESNp is the first engineered orthogonal protease with high selectivity based on a single amino acid difference.
Molecular dynamics simulations revealed functional active site rearrangements.