title: Sortase-mediated modification of αDEC205 affords optimization of antigen presentation and immunization against a set of viral epitopes.
abstract: A monoclonal antibody against the C-type lectin DEC205 (αDEC205) is an effective vehicle for delivery of antigens to dendritic cells through creation of covalent αDEC205-antigen adducts. These adducts can induce antigen-specific T-cell immune responses or tolerance. We exploit the transpeptidase activity of sortase to install modified peptides and protein-sized antigens onto the heavy chain of αDEC205, including linkers that contain nonnatural amino acids. We demonstrate stoichiometric site-specific labeling on a scale not easily achievable by genetic fusions (49 distinct fusions in this report). We conjugated a biotinylated version of a class I MHC-restricted epitope to unlabeled αDEC205 and monitored epitope generation upon binding of the adduct to dendritic cells. Our results show transfer of αDEC205 heavy chain to the cytoplasm, followed by proteasomal degradation. Introduction of a labile dipeptide linker at the N terminus of a T-cell epitope improves proteasome-dependent class I MHC-restricted peptide cross-presentation when delivered by αDEC205 in vitro and in vivo. We also conjugated αDEC205 with a linker-optimized peptide library of known CD8 T-cell epitopes from the mouse γ-herpes virus 68. Animals immunized with such conjugates displayed a 10-fold reduction in viral load.
authors.affiliation: Whitehead Institute for Biomedical Research and Massachusetts Institute of Technology, Cambridge, MA 02142, USA.
authors.forename: Lee Kim
authors.initials: LK
authors.lastname: Swee
authors.forename: Carla P
authors.initials: CP
authors.lastname: Guimaraes
authors.forename: Sharvan
authors.initials: S
authors.lastname: Sehrawat
authors.forename: Eric
authors.initials: E
authors.lastname: Spooner
authors.forename: M Inmaculada
authors.initials: MI
authors.lastname: Barrasa
authors.forename: Hidde L
authors.initials: HL
authors.lastname: Ploegh
mesh_terms: D000595:Amino Acid Sequence; D019881:Aminoacyltransferases / Q000378:metabolism*; D000818:Animals; D017951:Antigen Presentation; D015703:Antigens, CD / Q000737:chemistry / Q000235:genetics / Q000276:immunology* / Q000378:metabolism*; D000956:Antigens, Viral / Q000737:chemistry / Q000235:genetics / Q000276:immunology*; D001426:Bacterial Proteins / Q000378:metabolism*; D018414:CD8-Positive T-Lymphocytes / Q000276:immunology; D003546:Cysteine Endopeptidases / Q000378:metabolism*; D018984:Epitopes, T-Lymphocyte / Q000737:chemistry / Q000235:genetics / Q000276:immunology*; D006566:Herpesviridae Infections / Q000276:immunology / Q000517:prevention & control; D015395:Histocompatibility Antigens Class I / Q000378:metabolism; D007114:Immunization; D018796:Immunoconjugates / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D037181:Lectins, C-Type / Q000737:chemistry / Q000235:genetics / Q000276:immunology* / Q000378:metabolism*; D051379:Mice; D008807:Mice, Inbred BALB C; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D008822:Mice, Transgenic; D015778:Minor Histocompatibility Antigens; D008969:Molecular Sequence Data; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism; D015202:Protein Engineering; D011956:Receptors, Cell Surface / Q000737:chemistry / Q000235:genetics / Q000276:immunology* / Q000378:metabolism*; D018144:Rhadinovirus / Q000235:genetics / Q000276:immunology; D014412:Tumor Virus Infections / Q000276:immunology / Q000517:prevention & control; D022223:Vaccines, Subunit / Q000235:genetics / Q000276:immunology; D014765:Viral Vaccines / Q000235:genetics / Q000276:immunology
chemical_list: D015703:Antigens, CD; D000956:Antigens, Viral; D001426:Bacterial Proteins; C093569:DEC-205 receptor; D018984:Epitopes, T-Lymphocyte; D015395:Histocompatibility Antigens Class I; D018796:Immunoconjugates; D037181:Lectins, C-Type; D015778:Minor Histocompatibility Antigens; D011956:Receptors, Cell Surface; D022223:Vaccines, Subunit; D014765:Viral Vaccines; D019881:Aminoacyltransferases; C120542:sortase A; D003546:Cysteine Endopeptidases; D046988:Proteasome Endopeptidase Complex