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Related Experiment Video

Updated: Jun 26, 2025

Synthesis of an Intein-mediated Artificial Protein Hydrogel
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Novel protein ligase based on dual split intein.

Bing Lei1, Suyang Wang1, Xiaomeng Zhang1

  • 1College of Biological Science and Medical Engineering, Donghua University, Shanghai, 201620, PR China.

Biochemical and Biophysical Research Communications
|May 16, 2024
PubMed
Summary

Researchers developed a novel protein ligation system using split inteins for efficient protein engineering. This system demonstrates robust protein trans-splicing, offering new tools for creating complex proteins.

Keywords:
Controllable trans-splicingProtein ligaseProtein ligationSplit-inteinTrans-splicing

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Split-BioID &#8212; Proteomic Analysis of Context-specific Protein Complexes in Their Native Cellular Environment
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Split-BioID — Proteomic Analysis of Context-specific Protein Complexes in Their Native Cellular Environment

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Area of Science:

  • Biochemistry
  • Molecular Biology
  • Protein Engineering

Background:

  • Inteins are enzymes that catalyze protein splicing, excising themselves from precursor proteins.
  • Protein splicing typically occurs spontaneously with inteins covalently linked to exteins.
  • Split inteins offer potential for protein ligation and engineering applications.

Purpose of the Study:

  • To develop a novel protein ligation system utilizing two atypical split inteins.
  • To demonstrate the efficacy of this system in protein trans-splicing.
  • To explore optimization strategies and reaction mechanisms for intein-based ligation.

Main Methods:

  • Construction of fusion proteins with large segments of S1 and S11 split inteins.
  • Fusion of small intein segments to N-extein and C-extein sequences.
  • Demonstration of splicing activity in E. coli and in vitro.
  • Optimization of in vitro protein trans-splicing conditions.

Main Results:

  • A novel split intein system was successfully developed without cross-reactivity.
  • Protein trans-splicing was demonstrated in both cellular and cell-free systems.
  • In vitro splicing efficiency reached approximately 15% with various extein sequences.
  • Optimization strategies for in vitro trans-splicing were explored.

Conclusions:

  • The developed split intein system provides a novel method for protein ligation.
  • This approach expands the toolkit for trans-splicing-based protein engineering.
  • The findings offer new insights into intein-mediated protein ligase applications.