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Related Concept Videos

Restriction Enzymes01:11

Restriction Enzymes

Restriction enzymes are bacterial enzymes used to cut DNA in a sequence-specific manner. To cleave DNA, they bind to specific palindromic sequences called restriction sites. Such palindromic DNA sequences or inverted repeats are commonly found in regions of functional significance, such as the origin of replication, gene operator sites, and regions containing transcription termination signals.
The host bacteria protect their own genomic DNA from these enzymes by methylating these sites. Some...
Conservative Site-specific Recombination and Phase Variation02:53

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...

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Related Experiment Video

Updated: Jun 5, 2026

In Vitro Directed Evolution of a Restriction Endonuclease with More Stringent Specificity
09:16

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

Published on: March 25, 2020

Engineering enzyme specificity using computational design of a defined-sequence library.

Shaun M Lippow1, Tae Seok Moon, Subhayu Basu

  • 1Codon Devices, Inc., 99 Erie Street, Cambridge, MA 02139, USA. slippow@gmail.com

Chemistry & Biology
|December 21, 2010
PubMed
Summary
This summary is machine-generated.

Researchers developed a novel method combining computational design and enzyme screening to engineer highly active enzymes. This approach significantly enhanced glucose 6-oxidase for producing valuable chemicals from simple sugars.

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Protein WISDOM: A Workbench for In silico De novo Design of BioMolecules
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Area of Science:

  • Biotechnology
  • Enzyme Engineering
  • Synthetic Biology

Background:

  • Engineering enzymes with high activity and specificity is crucial for developing efficient biosynthetic pathways.
  • Current methods face limitations in creating enzymes for novel reactions.

Purpose of the Study:

  • To develop and validate a method combining structure-based computational protein design with library-based enzyme screening.
  • To engineer a glucose 6-oxidase enzyme for improved activity in a D-glucose to D-glucaric acid conversion pathway.

Main Methods:

  • A novel method was developed integrating computational protein design with library-based screening.
  • Inter-residue correlations identified by design were encoded into a defined-sequence library.
  • The approach was validated by engineering glucose 6-oxidase.

Main Results:

  • The engineered glucose 6-oxidase variant showed approximately 400-fold higher activity on glucose compared to the wild-type enzyme.
  • A highly active variant was identified after a single round of diversification and screening of 10,000 wells.
  • The method demonstrated rapid discovery of significantly improved enzymes.

Conclusions:

  • The combined computational design and library screening strategy is effective for engineering enzymes with enhanced activity and specificity.
  • This approach offers a broadly applicable tool for discovering new enzymes for engineered biological pathways.
  • The developed method accelerates the creation of biocatalysts for high-value molecule production.