Bach1, a heme-dependent transcription factor, reveals presence of multiple heme binding sites with distinct coordination structure.
The mammalian transcription factor Bach1 functions as a repressor of the enhancers of heme oxygenase-1 (HO-1) gene (Hmox-1) by forming heterodimers with the small Maf proteins such as MafK. The transcription of Hmox-1 is regulated by the substrate of HO-1, heme. Heme induces expression of Hmox-1 in part by inhibiting the binding of Bach1 to the enhancers and inducing the nuclear export of Bach1. A dipeptide motif of cysteine and proline (CP motif) in Bach1 is essential for the heme-mediated regulation. In this study, we show that five molecules of heme bind to Bach1 by the heme-titration assay. The Bach1-heme complex exhibits an absorption spectrum with a major Soret peak at 371 nm and Raman band at 343 cm(-1) in high amounts of heme and a spectrum containing the major Soret peak at 423 nm at low heme concentrations. The spectroscopic characterization indicates that Bach1 has two kinds of heme-binding sites with different coordination structures. Mutagenesis studies have established that four molecules of heme bind to the cysteine residues of four CP motifs in the C terminus of Bach1. These results raise the possibility that two separated activities of Bach1, DNA-binding and nuclear export, are regulated by heme binding at the different CP motifs of Bach1 respectively, but not by cooperative heme-binding.
Hira
Shusuke
S
Institute of Multidisciplinary Research for Advanced Materials, Tohoku University, Sendai, Japan.
Tomita
Takeshi
T
Matsui
Toshitaka
T
Igarashi
Kazuhiko
K
Ikeda-Saito
Masao
M
D001483:Base Sequence; D050976:Basic-Leucine Zipper Transcription Factors / Q000737:chemistry / Q000378:metabolism*; D001665:Binding Sites; D017931:DNA Primers; D051856:Fanconi Anemia Complementation Group Proteins / Q000737:chemistry / Q000378:metabolism*; D006418:Heme / Q000378:metabolism*; D006801:Humans; D010957:Plasmids; D016133:Polymerase Chain Reaction; D011487:Protein Conformation
C102108:BACH1 protein, human; D050976:Basic-Leucine Zipper Transcription Factors; D017931:DNA Primers; D051856:Fanconi Anemia Complementation Group Proteins; D006418:Heme