title: Structure of bradavidin-C-terminal residues act as intrinsic ligands.
abstract: Bradavidin is a homotetrameric biotin-binding protein from Bradyrhizobium japonicum, a nitrogen fixing and root nodule-forming symbiotic bacterium of the soybean. Wild-type (wt) bradavidin has 138 amino acid residues, whereas the C-terminally truncated core-bradavidin has only 118 residues. We have solved the X-ray structure of wt bradavidin and found that the C-terminal amino acids of each subunit were uniquely bound to the biotin-binding pocket of an adjacent subunit. The biotin-binding pocket occupying peptide (SEKLSNTK) was named "Brad-tag" and it serves as an intrinsic stabilizing ligand in wt bradavidin. The binding of Brad-tag to core-bradavidin was analysed by isothermal titration calorimetry and a binding affinity of ∼25 µM was measured. In order to study the potential of Brad-tag, a green fluorescent protein tagged with Brad-tag was prepared and successfully concentrated from a bacterial cell lysate using core-bradavidin-functionalized Sepharose resin.
authors.affiliation: Institute of Biomedical Technology, University of Tampere, Tampere University Hospital, Tampere, Finland.
authors.forename: Jenni
authors.initials: J
authors.lastname: Leppiniemi
authors.forename: Toni
authors.initials: T
authors.lastname: Grönroos
authors.forename: Juha A E
authors.initials: JA
authors.lastname: Määttä
authors.forename: Mark S
authors.initials: MS
authors.lastname: Johnson
authors.forename: Markku S
authors.initials: MS
authors.lastname: Kulomaa
authors.forename: Vesa P
authors.initials: VP
authors.lastname: Hytönen
authors.forename: Tomi T
authors.initials: TT
authors.lastname: Airenne
mesh_terms: D000345:Affinity Labels / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D001710:Biotin / Q000378:metabolism; D020369:Bradyrhizobium; D002352:Carrier Proteins / Q000737:chemistry* / Q000378:metabolism*; D018360:Crystallography, X-Ray; D008024:Ligands; D008958:Models, Molecular; D008969:Molecular Sequence Data; D011487:Protein Conformation; D055550:Protein Stability; D021122:Protein Subunits / Q000737:chemistry / Q000378:metabolism; D013379:Substrate Specificity
chemical_list: D000345:Affinity Labels; D002352:Carrier Proteins; D008024:Ligands; D021122:Protein Subunits; C501304:bradavidin protein, Bradyrhizobium japonicum; D001710:Biotin