Small-Scale Secretory VHH Expression in Saccharomyces cerevisiae

Area of Science:

• Biotechnology
• Molecular Biology
• Immunology

Background:

• Single-domain antibodies (VHHs) are typically produced in Escherichia coli after isolation.
• Targeting VHH expression to the secretory pathway of Saccharomyces cerevisiae (baker's yeast) allows for secretion of correctly folded, soluble, disulfide-bonded, and N-glycosylated VHHs.

Purpose of the Study:

• To describe the small-scale production of VHHs in baker's yeast using both episomal and genomically integrated vectors.
• To enable the production of VHHs linked to the llama long hinge region for enhanced functionality.
• To facilitate the development of double antibody sandwich ELISAs for antigen detection.

Main Methods:

• Small-scale production of VHHs in baker's yeast using shaker flasks.
• Utilized both episomal vectors and vectors requiring genomic integration for VHH expression.
• Engineered VHHs linked to the llama long hinge region with a single cysteine residue.

Main Results:

• Successfully produced correctly folded, soluble, disulfide-bonded, and N-glycosylated VHHs secreted into the culture medium.
• Generated VHHs linked to the llama long hinge region, enabling partial dimerization via a single cysteine residue.
• Demonstrated the suitability of this format for developing double antibody sandwich ELISAs.

Conclusions:

• Baker's yeast is an effective system for producing functional, secreted VHHs.
• The VHH-hinge fusion protein format is ideal for constructing sensitive sandwich ELISAs.
• This method is applicable for detecting various antigens, including foot-and-mouth disease virus.