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 Experiment Videos

Selection for improved subtiligases by phage display.

S Atwell1, J A Wells

  • 1Department of Protein Engineering, Genentech, Inc., 1 DNA Way, South San Francisco, CA 94080, USA.

Proceedings of the National Academy of Sciences of the United States of America
|August 18, 1999
PubMed
Summary
This summary is machine-generated.

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

Non-Invasive MRI of Blood-Cerebrospinal Fluid Barrier Function.

Nature communications·2020
Same author

CNS distribution, signalling properties and central effects of G-protein coupled receptor 4.

Neuropharmacology·2018
Same author

Tissue magnetic susceptibility mapping as a marker of tau pathology in Alzheimer's disease.

NeuroImage·2017
Same author

MRI of cerebral micro-vascular flow patterns: A multi-direction diffusion-weighted ASL approach.

Journal of cerebral blood flow and metabolism : official journal of the International Society of Cerebral Blood Flow and Metabolism·2016
Same author

Cacidases: caspases can cleave after aspartate, glutamate and phosphoserine residues.

Cell death and differentiation·2016
Same author

Application of neurite orientation dispersion and density imaging (NODDI) to a tau pathology model of Alzheimer's disease.

NeuroImage·2015

This study presents a phage display method to engineer enzyme activity by screening millions of variants. Researchers successfully enhanced subtiligase ligase activity and identified mutants with improved stability.

Area of Science:

  • Biochemistry
  • Molecular Biology
  • Protein Engineering

Background:

  • Engineering enzyme activity is hindered by complex structure-function relationships and challenges in screening numerous enzyme mutants.
  • Developing efficient methods for enzyme variant screening is crucial for advancing protein engineering.

Purpose of the Study:

  • To develop and demonstrate a product capture strategy using phage display for selecting improved enzymes from extensive mutant libraries.
  • To engineer subtiligase, a peptide ligase, for enhanced catalytic activity and stability.

Main Methods:

  • Utilized phage display to present a large library of subtiligase variants (>10^9 mutants) on phage particles.
  • Randomly mutated active site residues of subtiligase in defined groups to create diverse mutant libraries.

Related Experiment Videos

  • Employed a product capture strategy to selectively isolate variants that ligated a biotin peptide to their N termini.

    • Main Results:

    • Identified mutant subtiligases with significantly increased ligase activity compared to wild-type.
    • Discovered novel subtiligase mutants possessing enhanced stability and oxidative resistance, traits associated with wild-type subtilisin.
    • Demonstrated the efficacy of phage display for enzyme engineering when linked to a selectable catalytic event.

    Conclusions:

    • Phage display with product capture is a powerful strategy for directed evolution of enzymes with improved function.
    • This approach can simultaneously enhance catalytic activity and other desirable properties like stability.
    • The methodology provides a versatile platform for engineering various enzymes for biotechnological applications.