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Updated: May 27, 2025

Genome-wide Protein-protein Interaction Screening by Protein-fragment Complementation Assay PCA in Living Cells
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Multiplex mapping of protein-protein interaction interfaces.

Jingxuan He1, Ling-Nan Zou1, Vidhi Pareek2

  • 1Department of Chemistry, The Pennsylvania State University, University Park, PA 16802, USA.

Biorxiv : the Preprint Server for Biology
|February 20, 2025
PubMed
Summary
This summary is machine-generated.

We developed SpARC-map, a novel method for identifying protein-protein interaction interfaces in vivo. This technique uses bacterial affinity selection and DNA sequencing to map interactions, aiding in structural biology and drug discovery.

Keywords:
protein complexprotein interaction interfaceprotein-protein interactionpurinosome

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Area of Science:

  • Biochemistry
  • Molecular Biology
  • Structural Biology

Background:

  • Identifying protein-protein interaction (PPI) interfaces is crucial for understanding cellular mechanisms.
  • Existing methods often require specialized equipment or in vitro complex reconstitution, limiting their accessibility.

Purpose of the Study:

  • To introduce SpARC-map, a novel, accessible in vivo method for mapping PPI interfaces.
  • To demonstrate the utility of SpARC-map in identifying known and novel protein interaction sites.

Main Methods:

  • SpARC-map employs in vivo affinity selection within a bacterial host.
  • High-throughput DNA sequencing is used to infer the location of PPI interfaces.
  • The method utilizes routine microbiology techniques without specialized instrumentation.

Main Results:

  • SpARC-map successfully identified the known interface in the p21-PCNA complex.
  • The method revealed novel interfaces within the purinosome complex, consistent with substrate channeling.
  • SpARC-map results provided constraints for machine learning-based protein structure prediction.

Conclusions:

  • SpARC-map is a versatile and sensitive method for mapping PPIs across a range of affinities.
  • The technique facilitates the study of complex biological systems like the purinosome.
  • SpARC-map has implications for structural biology, drug discovery, and computational protein modeling.