title: Cell surface display of functional human MHC class II proteins: yeast display versus insect cell display.
abstract: Reliable and robust systems for engineering functional major histocompatibility complex class II (MHCII) proteins have proved elusive. Availability of such systems would enable the engineering of peptide-MHCII (pMHCII) complexes for therapeutic and diagnostic applications. In this paper, we have developed a system based on insect cell surface display that allows functional expression of heterodimeric DR2 molecules with or without a covalently bound human myelin basic protein (MBP) peptide, which is amenable to directed evolution of DR2-MBP variants with improved T cell receptor (TCR)-binding affinity. This study represents the first example of functional display of human pMHCII complexes on insect cell surface. In the process of developing this pMHCII engineering system, we have also explored the potential of using yeast surface display for the same application. Our data suggest that yeast display is a useful system for analysis and engineering of peptide binding of MHCII proteins, but not suitable for directed evolution of pMHC complexes that bind with low affinity to self-reactive TCRs.
authors.affiliation: Department of Chemical and Biomolecular Engineering, University of Illinois at Urbana-Champaign, Urbana, IL 61801, USA.
authors.forename: Fei
authors.initials: F
authors.lastname: Wen
authors.forename: Dhruv K
authors.initials: DK
authors.lastname: Sethi
authors.forename: Kai W
authors.initials: KW
authors.lastname: Wucherpfennig
authors.forename: Huimin
authors.initials: H
authors.lastname: Zhao
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D016367:Baculoviridae / Q000235:genetics; D002460:Cell Line; D019020:Directed Molecular Evolution / Q000379:methods*; D005434:Flow Cytometry; D015802:HLA-DR2 Antigen / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics; D006801:Humans; D006825:Hybridomas; D016350:Leucine Zippers; D008565:Membrane Proteins / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics; D051379:Mice; D008969:Molecular Sequence Data; D004676:Myelin Basic Protein / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics; D015202:Protein Engineering / Q000379:methods*; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism*; D016415:Sequence Alignment; D018411:Spodoptera / Q000166:cytology / Q000378:metabolism / Q000821:virology; D013601:T-Lymphocytes / Q000378:metabolism
chemical_list: D015802:HLA-DR2 Antigen; D008565:Membrane Proteins; D004676:Myelin Basic Protein; D011994:Recombinant Proteins