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Parallel Screening Using the Chloroalkane Penetration Assay Reveals Structure-Penetration Relationships
Kaley M Mientkiewicz1, Leila Peraro1, Joshua A Kritzer1
1Department of Chemistry, Tufts University, 62 Talbot Avenue, Medford, Massachusetts 02155 United States.
ACS Chemical Biology
|July 5, 2021
Summary
Polycationic peptide cytosolic penetration depends on arginine residue patterns. A new high-throughput assay quantifies this penetration, revealing how arginine number, pattern, and stereochemistry influence cell entry.
Area of Science:
- Biochemistry
- Cell Biology
- Drug Delivery
Background:
- Polycationic peptides are crucial for drug delivery, but their ability to enter the cytosol is key.
- Previous methods often measured total cell uptake, not specific cytosolic penetration.
- Understanding arginine residue influence is vital for designing effective peptide therapeutics.
Purpose of the Study:
- To develop and validate a high-throughput screening platform for measuring peptide cytosolic penetration.
- To investigate the impact of arginine residue number, patterning, and stereochemistry on cytosolic entry.
- To optimize peptide design for enhanced intracellular delivery.
Main Methods:
- Adaptation of the chloroalkane penetration assay for high-throughput screening.
- Quantitative analysis of peptide libraries with varying arginine content and arrangement.
- Automation-ready platform for efficient peptide screening.
Main Results:
- The developed assay accurately and quantitatively measures cytosolic penetration.
- Identified specific patterns of arginine residues that enhance cytosolic entry.
- Demonstrated the influence of arginine number and stereochemistry on penetration efficiency.
Conclusions:
- The high-throughput chloroalkane assay is effective for studying peptide cytosolic penetration.
- Arginine residue characteristics significantly modulate peptide entry into the cytosol.
- This platform facilitates the rational design of cell-penetrating peptides for therapeutic applications.
