title: Reconstitution of bacterial expressed human CD94: the importance of the stem region for dimer formation.
abstract: Human CD94 is a subunit of the disulfide-linked, heterodimeric natural killer (NK) cell surface receptor CD94/NKG2. This receptor, a member of the C-type lectin superfamily, participates in regulating NK cell directed lysis through interaction with the major histocompatibility antigen HLA-E. Two forms of CD94 were expressed using a bacterial expression system and refolded in vitro. One form, residues 34-179, designated S34, corresponds to the entire extracellular region of the receptor, including a 23-residue stem region, and the other, residues 51-179, designated E51, corresponds only to the putative carbohydrate recognition domain of the receptor. The refolded full-length S34 protein existed as a noncovalent dimer initially but formed an interchain disulfide bond upon storage for several months. In contrast, the stemless construct, E51, existed largely as a monomeric form. The stem region of S34, residues 34-56, is sensitive to proteolysis and its absence results in dissociation of the dimer. This suggests that the residues in the stem region of CD94 help to stabilize the dimeric conformation.
authors.affiliation: Structural Biology Section, Office of The Scientific Director, National Institute of Allergy and Infectious Diseases, National Institutes of Health, 12441 Parklawn Drive, Rockville, Maryland 20852, USA.
authors.forename: J C
authors.initials: JC
authors.lastname: Boyington
authors.forename: A N
authors.initials: AN
authors.lastname: Raiz
authors.forename: A G
authors.initials: AG
authors.lastname: Brooks
authors.forename: A
authors.initials: A
authors.lastname: Patamawenu
authors.forename: P D
authors.initials: PD
authors.lastname: Sun
mesh_terms: D000595:Amino Acid Sequence; D015703:Antigens, CD / Q000737:chemistry*; D019281:Dimerization; D004220:Disulfides / Q000737:chemistry; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000737:chemistry / Q000378:metabolism; D008401:Gas Chromatography-Mass Spectrometry; D006801:Humans; D002479:Inclusion Bodies / Q000737:chemistry / Q000378:metabolism; D037181:Lectins, C-Type; D008562:Membrane Glycoproteins / Q000737:chemistry*; D008969:Molecular Sequence Data; D055654:NK Cell Lectin-Like Receptor Subfamily C; D051920:NK Cell Lectin-Like Receptor Subfamily D; D017510:Protein Folding; D020673:Protein Renaturation; D017434:Protein Structure, Tertiary; D011971:Receptors, Immunologic / Q000737:chemistry*; D011975:Receptors, Mitogen / Q000737:chemistry*; D055607:Receptors, Natural Killer Cell; D020539:Sequence Analysis, Protein
chemical_list: D015703:Antigens, CD; D004220:Disulfides; C527054:KLRC1 protein, human; C497869:KLRD1 protein, human; D037181:Lectins, C-Type; D008562:Membrane Glycoproteins; D055654:NK Cell Lectin-Like Receptor Subfamily C; D051920:NK Cell Lectin-Like Receptor Subfamily D; D011971:Receptors, Immunologic; D011975:Receptors, Mitogen; D055607:Receptors, Natural Killer Cell