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Continuous directed evolution of proteins with improved soluble expression.

Tina Wang1,2, Ahmed H Badran1,2, Tony P Huang1,2

  • 1Merkin Institute of Transformative Technologies in Healthcare, Broad Institute of Harvard and MIT, Cambridge, MA, USA.

Nature Chemical Biology
|August 22, 2018
PubMed
Summary
This summary is machine-generated.

We developed soluble expression phage-assisted continuous evolution (SE-PACE) to rapidly enhance protein expression. This system improves protein yield and activity, demonstrating broad applicability for protein engineering.

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Area of Science:

  • Biotechnology
  • Molecular Biology
  • Protein Engineering

Background:

  • Improving soluble protein expression is crucial for biotechnology and therapeutic applications.
  • Existing methods for protein evolution can be time-consuming and may not optimize for expression levels.
  • Phage-assisted continuous evolution (PACE) is a powerful tool for protein evolution but requires adaptation for soluble expression optimization.

Purpose of the Study:

  • To develop a novel system, SE-PACE, for rapidly evolving proteins with enhanced soluble expression.
  • To enable simultaneous selection for improved expression and retained protein activity.
  • To demonstrate the versatility of SE-PACE across different protein types, including antibody fragments, MBP, and APOBEC1.

Main Methods:

  • Development of SE-PACE utilizing a PACE-compatible AND gate with a split-intein pIII.
  • Application of SE-PACE for directed evolution of antibody fragments, maltose-binding protein (MBP), and APOBEC1.
  • Creation of an activity-independent SE-PACE variant for correcting folding defects and improving expression.

Main Results:

  • SE-PACE rapidly evolved antibody fragments with over 5-fold improvement in expression yield while maintaining binding activity.
  • Corrected folding-defective MBP variants and evolved APOBEC1 variants with superior expression properties.
  • Evolved APOBEC1 variants enhanced the expression and apparent activity of Cas9-derived base editors.

Conclusions:

  • SE-PACE is an effective system for accelerating the evolution of proteins with improved soluble expression.
  • The SE-PACE system can be adapted for both activity-dependent and activity-independent protein evolution.
  • This technology holds significant potential for diverse protein engineering applications in biotechnology and medicine.