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PIONEER: A Periplasmic Display Platform for Synthetic Biology-Based Screening of Genetically Encoded Protein
Jacob B Rowe1, Kyutae Lee1, Daniel G Isom1,2,3,4
1Department of Molecular and Cellular Pharmacology, University of Miami Miller School of Medicine, Miami, FL, USA.
Biorxiv : the Preprint Server for Biology
|November 24, 2025
Summary
We developed PIONEER, a yeast-based platform for studying biomolecules like proteins and nanobodies. This system enables function-first screening of membrane proteins by assessing activity through signaling, advancing drug discovery.
Area of Science:
- Biochemistry
- Molecular Biology
- Yeast Genetics
Background:
- Periplasmic display in yeast is an emerging technique for biomolecule screening.
- Existing methods have limitations in studying membrane protein function and ligand interactions.
Purpose of the Study:
- To present PIONEER, a modular yeast platform for periplasmic display of peptides, proteins, and nanobodies.
- To enable direct interrogation of membrane protein function and facilitate function-first screening.
Main Methods:
- Optimizing secretion signals and display scaffolds for stable ligand retention and autocrine signaling in *Saccharomyces cerevisiae*.
- Utilizing yeast periplasmic space for localized display and functional assessment of biomolecules.
- Applying the platform to human G protein-coupled receptors (GPCRs) for activity profiling and regulator classification.
Main Results:
- PIONEER successfully enables detection of surface expression and ligand activity profiling for GPCRs.
- The system distinguishes nanobody regulators, including antagonists and conformational stabilizers.
- Functional effects and chaperone activity of intrabodies can be differentiated using this platform.
Conclusions:
- PIONEER is a versatile platform for function-first screening of membrane and soluble proteins.
- It supports the assessment of biomolecule function through signaling, not just binding.
- The platform is adaptable for diverse targets and scalable for AI-driven protein design validation.
