Yeast surface display has emerged as a powerful and versatile technology for antibody discovery and protein engineering, playing a critical role in therapeutic development. By leveraging the eukaryotic machinery of yeast, this platform ensures proper protein folding and display, enabling the isolation of high-affinity binders through robust library screening.
Technology Overview
(i) Display Mechanism
The system utilizes the yeast Aga1p and Aga2p mating proteins. The antibody or target protein is genetically fused to Aga2p, which binds to Aga1p on the cell wall via disulfide bonds, securing the protein on the surface.
(ii) Key Advantages
As a eukaryote, yeast enables complex protein folding and glycosylation, ensuring consistent display of diverse proteins beyond antibodies, such as fibronectins and T cell receptors (TCRs).
(iii) Library Construction
We employ high-efficiency yeast electroporation to generate display libraries with sizes reaching up to 10⁹, providing exceptional diversity for hit discovery.
Antibody Discovery Workflow
The process of isolating novel antibodies via yeast display follows a structured sequence from antigen preparation to clonal selection.
Antigen Preparation
(i) Production
Target genes are synthesized, cloned into expression vectors, and expressed in systems like E. coli or mammalian cells. Proteins are then purified using affinity chromatography.
(ii) Labeling
Antigens are labeled with biotin or fluorophores to facilitate detection during screening. Biotinylation allows binding to streptavidin-conjugated magnetic beads or fluorescent dyes.
Library Construction
(i) Gene Cloning
Antibody genes (from immunized animals, naive libraries, or synthetic sources) are cloned into display vectors (e.g., pYD1). These vectors fuse the antibody to Aga2p under a galactose-inducible promoter and include detection tags like c-myc.
(ii) Transformation
The vector library is transformed into S. cerevisiae strains (e.g., EBY100) via high-efficiency electroporation and selected on SD-CAA medium to create a diverse yeast library.
Library Screening
(i) Magnetic Bead Enrichment
Induced yeast libraries are first incubated with biotinylated antigen and streptavidin magnetic beads. This step rapidly captures antigen-binding cells, removing non-binders and enriching the target population.
(ii) Flow Cytometry Sorting
Enriched pools are stained with fluorescent antigens and tag-specific antibodies. Dual-parameter sorting isolates yeast cells exhibiting both high display levels and strong antigen binding. Multiple rounds of sorting are performed to select clones with the highest affinity and specificity.
Summary
By combining high-diversity library construction with a dual screening strategy of magnetic enrichment and flow cytometry sorting, yeast surface display provides a rapid and reliable platform for isolating high-performance antibodies for therapeutic and diagnostic applications.
Alpha Lifetech has been continuously developing and refining its yeast surface display platform, enabling surface display of proteins and various antibodies. In addition, we provide comprehensive services including antigen preparation, animal immunization, downstream antibody affinity detection, and Antibody affinity maturation.