Yeast surface display for protein engineering and characterization

Abstract

Yeast surface display is being employed to engineer desirable properties into proteins for a broad variety of applications. Labeling with soluble ligands enables rapid and quantitative analysis of yeast-displayed libraries by flow cytometry, while cell-surface selections allow screening of libraries with insoluble or even as-yet-uncharacterized binding targets. In parallel, the utilization of yeast surface display for protein characterization, including in particular the mapping of functional epitopes mediating protein-protein interactions, represents a significant recent advance.

Figure 1

A selection of proteins successfully displayed as Aga2p fusions on the surface of yeast. Top row, left to right: human epidermal growth factor [31^••] (1JL9), human interleukin-2 [61] (2B51), single-chain antibody fragment 4m5.3 [62] (1X9Q), green fluorescent protein [63] (1EMA), human α_L integrin inserted domain [19] (1LFA), and human fibronectin [18] (1FNA). Bottom row, left to right: West Nile Virus envelope protein [32^•] (2I69), human EGF receptor ectodomain [27] (1NQL), and human MHC class II HLA-DR4αβ in complex with peptide [64] (2SEB). The PDB IDs for the structures shown are noted in parentheses. This figure was generated using Swiss-Pdb Viewer [65].

Figure 2

Comparison of numbers of studies employing phage or yeast display.

Figure 3

Comparison of dissociation constants determined by yeast surface display, K_d YSD, and other methods, K_d other. scFv binding to fluorescein; K_d other determined by fluorescence quenching. scFv binding botulinum neurotoxin type A; K_d other determined by surface plasmon resonance (SPR) [15]. scFv binding to carcinoembryonic antigen (CEA); K_d other determined by titration of soluble scFv against mammalian-cell-surface-expressed CEA (Michael M Schmidt, unpublished results). scFv binding to hen egg white lysozyme (HEL); K_d other determined by fluorescence quench titration. scFv binding to p53 peptides; K_d other determined by SPR. scFv binding to EGF; K_d other determined by SPR. scFv binding to heparin-binding EGF; K_d other determined by SPR. scFv binding to xeroderma pigmentosum-complementing protein group A; K_d other determined by SPR. Fibronectin binding to HEL; K_d other determined by equilibrium competition titration using purified fibronectin mutants [18]. Fibronection binding to maltose-binding protein; K_d other determined by SPR [17^••]. scTCR binding staphylococcal enterotoxin C3; K_d other determined by SPR. scTCR binding toxic shock syndrome toxin-1; K_d other determined by SPR [21]. Figure modified from Figure 8 of Lipovsek et al. [18].