Single-chain Fv phage display propensity exhibits strong positive correlation with overall expression levels

Area of Science:

• Biotechnology
• Molecular Biology
• Immunology

Background:

• Single chain variable fragments (scFvs) are crucial in research, diagnostics, and therapeutics.
• Phage display is a primary method for scFv discovery, influenced by various factors.
• Low thermodynamic stability of scFvs can lead to aggregation, impacting their utility.

Purpose of the Study:

• To investigate the relationship between scFv expression levels and their display efficiency on phage.
• To understand how scFv expression influences protein localization and yield in E. coli.
• To identify factors affecting scFv success in phage display library screening.

Main Methods:

• Characterization of 15 anti-tetanus toxin scFvs.
• Analysis of the correlation between overall scFv expression and display propensity.
• Assessment of soluble scFv localization in periplasmic and extracellular fractions.

Main Results:

• A strong positive correlation (Rho = 0.88) was observed between scFv expression and display propensity.
• Display propensity, expression, and localization were clone-specific, even with high sequence homology.
• No correlation was found between non-fused scFv expression and its soluble periplasmic or extracellular localization.

Conclusions:

• ScFv phage display propensity is directly linked to overall expression, assuming no premature aggregation.
• Factors like codon bias and mRNA abundance indirectly influence scFv display.
• High-expression scFvs may not yield high soluble protein, necessitating careful clone selection from libraries.