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Hormone phage: an enrichment method for variant proteins with altered binding properties
1Department of Protein Engineering, Genentech, Inc., South San Francisco, California 94080.
Proteins
|January 1, 1990
Summary
This study developed a novel phage display system for human growth hormone (hGH). The system successfully displayed functional hGH, enabling efficient enrichment and isolation of hGH variants.
Area of Science:
- Biotechnology
- Molecular Biology
- Protein Engineering
Background:
- Filamentous phage display systems are valuable for protein engineering and antibody selection.
- Displaying large proteins like human growth hormone (hGH) presents challenges due to protein folding and epitope accessibility.
Purpose of the Study:
- To engineer a phage display system capable of presenting functional human growth hormone (hGH).
- To validate the system's utility for isolating hGH variants with altered receptor binding properties.
Main Methods:
- A gene fusion was constructed between human growth hormone (hGH) and the M13 gene III protein.
- The fusion construct was cloned into a phagemid vector and expressed on the surface of filamentous phage particles.
- Phage particles displaying the hGH-gene III fusion were selected using hGH receptor-coated beads.
Main Results:
- The hGH-gene III fusion protein was correctly folded and immunologically active, confirmed by monoclonal antibody binding.
- Phage particles displaying the hGH fusion were significantly enriched (>5000-fold) after affinity selection.
- The system demonstrated effectiveness in isolating specific hGH variants.
Conclusions:
- The developed phage display system enables the functional presentation of human growth hormone (hGH).
- This system is a powerful tool for isolating novel hGH receptor binding mutants.
- The approach is potentially applicable to displaying other large proteins with complex epitopes for selection purposes.