title: Structural properties of the DNA-bound form of a novel tandem repeat DNA-binding domain, STPR.
abstract: Fibroin-modulator-binding protein 1 (FMBP-1) is a predicted transcription factor of the silkworm fibroin gene. The DNA-binding domain of FMBP-1 consists of four almost perfect tandem repeats of 23 amino acids each (R1-R4), and is referred to as the score and three amino acid peptide repeat (STPR) domain. This characteristic domain is conserved in eukaryotes, but the DNA-binding mode is not known. In this study, the structural properties of the DNA-bound form of the STPR domain were characterized. The combined experiments indicated that the STPR domain bound to the DNA duplex with a 1:1 binding ratio. The specific DNA caused considerable changes in the thermal unfolding profile and the digestion pattern of the STPR domain. These data suggested that the domain adapts a quite rigid helix-rich structure in the DNA-bound state, even though it moves flexibly in the absence of DNA. Furthermore, mutual induced-fit conformational change was also observed in DNA. Finally, we determined the DNA-binding surface of the STPR third repeat (R3) by alanine scanning mutagenesis; a particular site, composed of hydrophobic and hydrophilic residues, was identified. Notably, the substitution of Arg-9 in R3 with alanine residue, which is located in the middle of the surface, drastically abolished the alpha-helix-inducing and DNA-binding abilities. From these results, we predicted the DNA-binding mode of the STPR domain.
authors.affiliation: Department of Biological Sciences, Graduate School of Science, Hokkaido University, Sapporo 060-0810, Japan.
authors.forename: Shin
authors.initials: S
authors.lastname: Saito
authors.forename: Takuya
authors.initials: T
authors.lastname: Yokoyama
authors.forename: Tomoyasu
authors.initials: T
authors.lastname: Aizawa
authors.forename: Kyosuke
authors.initials: K
authors.lastname: Kawaguchi
authors.forename: Takeshi
authors.initials: T
authors.lastname: Yamaki
authors.forename: Daisuke
authors.initials: D
authors.lastname: Matsumoto
authors.forename: Tatsuro
authors.initials: T
authors.lastname: Kamijima
authors.forename: Masakatsu
authors.initials: M
authors.lastname: Kamiya
authors.forename: Yasuhiro
authors.initials: Y
authors.lastname: Kumaki
authors.forename: Mineyuki
authors.initials: M
authors.lastname: Mizuguchi
authors.forename: Sigeharu
authors.initials: S
authors.lastname: Takiya
authors.forename: Makoto
authors.initials: M
authors.lastname: Demura
authors.forename: Keiichi
authors.initials: K
authors.lastname: Kawano
mesh_terms: D000596:Amino Acids; D000817:Anilino Naphthalenesulfonates / Q000378:metabolism; D000818:Animals; D012831:Bombyx / Q000737:chemistry*; D002942:Circular Dichroism; D004247:DNA / Q000378:metabolism*; D057927:Hydrophobic and Hydrophilic Interactions; D019476:Insect Proteins / Q000737:chemistry* / Q000378:metabolism; D050505:Mutant Proteins / Q000378:metabolism; D010446:Peptide Fragments / Q000378:metabolism; D010447:Peptide Hydrolases / Q000378:metabolism; D011485:Protein Binding; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D013050:Spectrometry, Fluorescence; D020080:Tandem Repeat Sequences; D017186:Titrimetry
chemical_list: D000596:Amino Acids; D000817:Anilino Naphthalenesulfonates; D019476:Insect Proteins; D050505:Mutant Proteins; D010446:Peptide Fragments; C027132:1-anilino-8-naphthalenesulfonate; D004247:DNA; D010447:Peptide Hydrolases