title: BAFF/BLyS receptor 3 comprises a minimal TNF receptor-like module that encodes a highly focused ligand-binding site.
abstract: BAFF/BLyS, a member of the tumor necrosis family (TNF) superfamily of ligands, is a crucial survival factor for B cells. BAFF binds three receptors, TACI, BCMA, and BR3, with signaling through BR3 being essential for promoting B cell function. Typical TNF receptor (TNFR) family members bind their cognate ligands through interactions with two cysteine-rich domains (CRDs). However, the extracellular domain (ECD) of BR3 consists of only a partial CRD, with cysteine spacing distinct from other modules described previously. Herein, we report the solution structure of the BR3 ECD. A core region of only 19 residues adopts a stable structure in solution. The BR3 fold is analogous to the first half of a canonical TNFR CRD but is stabilized by an additional noncanonical disulfide bond. BAFF-binding determinants were identified by shotgun alanine-scanning mutagenesis of the BR3 ECD expressed on phage. Several of the key BAFF-binding residues are presented from a beta-turn that we have shown previously to be sufficient for ligand binding when transferred to a structured beta-hairpin scaffold [Kayagaki, N., Yan, M., Seshasayee, D., Wang, H., Lee, W., French, D. M., Grewal, I. S., Cochran, A. G., Gordon, N. C., Yin, J., Starovasnik, M. A, and Dixit, V. M. (2002) Immunity 10, 515-524]. Outside of the turn, mutagenesis identifies additional hydrophobic contacts that enhance the BAFF-BR3 interaction. The crystal structure of the minimal hairpin peptide, bhpBR3, in complex with BAFF reveals intimate packing of the six-residue BR3 turn into a cavity on the ligand surface. Thus, BR3 binds BAFF through a highly focused interaction site, unprecedented in the TNFR family.
authors.affiliation: Department of Protein Engineering, Genentech, Inc., One DNA Way, South San Francisco, California 94080, USA.
authors.forename: Nathaniel C
authors.initials: NC
authors.lastname: Gordon
authors.forename: Borlan
authors.initials: B
authors.lastname: Pan
authors.forename: Sarah G
authors.initials: SG
authors.lastname: Hymowitz
authors.forename: JianPing
authors.initials: J
authors.lastname: Yin
authors.forename: Robert F
authors.initials: RF
authors.lastname: Kelley
authors.forename: Andrea G
authors.initials: AG
authors.lastname: Cochran
authors.forename: Minhong
authors.initials: M
authors.lastname: Yan
authors.forename: Vishva M
authors.initials: VM
authors.lastname: Dixit
authors.forename: Wayne J
authors.initials: WJ
authors.lastname: Fairbrother
authors.forename: Melissa A
authors.initials: MA
authors.lastname: Starovasnik
mesh_terms: D000595:Amino Acid Sequence; D053264:B-Cell Activating Factor; D053265:B-Cell Activation Factor Receptor; D001665:Binding Sites; D003545:Cysteine / Q000737:chemistry; D006801:Humans; D066298:In Vitro Techniques; D008024:Ligands; D008565:Membrane Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D008958:Models, Molecular; D008969:Molecular Sequence Data; D019906:Nuclear Magnetic Resonance, Biomolecular; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D018124:Receptors, Tumor Necrosis Factor / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012996:Solutions; D014409:Tumor Necrosis Factor-alpha / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism
chemical_list: D053264:B-Cell Activating Factor; D053265:B-Cell Activation Factor Receptor; C120197:BLyS receptor; D008024:Ligands; D008565:Membrane Proteins; D018124:Receptors, Tumor Necrosis Factor; D011994:Recombinant Proteins; D012996:Solutions; C435926:TNFRSF13C protein, human; C505719:TNFSF13B protein, human; D014409:Tumor Necrosis Factor-alpha; D003545:Cysteine