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Live-cell protein engineering with an ultra-short split intein.

Antony J Burton1, Michael Haugbro1, Eva Parisi1

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Split inteins enable protein semisynthesis in live cells. The novel VidaL split intein facilitates rapid protein trans-splicing for precise chemical modification and engineering of cellular proteins.

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chemical biologyintein splicingprotein engineeringprotein semisynthesis

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Area of Science:

  • Biochemistry
  • Molecular Biology
  • Protein Engineering

Background:

  • Split inteins are effective for in vitro protein modification.
  • Their application for in vivo protein semisynthesis in live cells remains limited.

Purpose of the Study:

  • To characterize the naturally split intein VidaL for its potential in live-cell protein engineering.
  • To demonstrate the utility of VidaL for in vitro and in vivo protein semisynthesis.

Main Methods:

  • Biochemical and structural characterization of the VidaL split intein.
  • Demonstration of protein trans-splicing in vitro and in mammalian cells.
  • Application in assembling multidomain proteins and modifying histone proteins.

Main Results:

  • VidaL exhibits rapid and efficient protein trans-splicing.
  • The shortest known N-terminal fragment of VidaL enables versatile protein engineering.
  • Successful semisynthesis of dual posttranslationally modified histones and modification/translocation of HP1α in live cells.

Conclusions:

  • The VidaL system is a powerful tool for precise chemical modification of cellular proteins.
  • It allows for spatial and temporal control over protein engineering in live cells.
  • VidaL expands the capabilities of protein semisynthesis for complex biological applications.