TY - JOUR
T1 - Structural insights into the asymmetric effects of zinc-ligand cysteine mutations in the novel zinc ribbon domain of human TFIIEα for transcription
AU - Okuda, Masahiko
AU - Tanaka, Aki
AU - Hanaoka, Fumio
AU - Ohkuma, Yoshiaki
AU - Nishimura, Yoshifumi
N1 - Funding Information:
This work was supported by a grant for Collaborative of Regional Entities for the Advancement of Technological Excellence (CREATE) from the Japan Science and Technology Agency, by the Project on Protein 3000, Transcription and Translation, and by Grants-in-Aid for Scientific Research from the Ministry of Education, Culture, Sports, Science and Technology.
PY - 2005/10
Y1 - 2005/10
N2 - The large subunit of TFIIE (TFIIEα) has a highly conserved zinc ribbon domain, which is essential for transcription. Recently, we determined the solution structure of this domain to be that of a novel zinc finger motif [Okuda et al. (2004) J. Biol. Chem. 279, 51395-51403]. On examination of the functions of four cysteine mutants of TFIIEα, in which each of four zinc-liganded cysteines was replaced by alanine, we found an interesting functional asymmetry; on a supercoiled template, the two C-terminal mutants did not show any transcriptional activity, however, the two N-terminal mutants retained about 20% activity. Furthermore, these two pairs of mutants showed distinct binding abilities as to several general transcription factors. To obtain structural insights into the asymmetry, here we have analyzed the structures of the four cysteine mutants of the zinc ribbon domain by CD and NMR. All four mutants possessed a characteristic partially folded structure coordinating with a zinc atom, despite the imperfect set of cysteine-ligands. However, they equilibrated with several structures including the random coil structure. Unexpectedly, the two N-terminal mutants mainly equilibrated with the random coil structure, while the two C-terminal ones mainly equilibrated with folded structures. The characteristic structure formation of each mutant was reversible, which totally depended on the zinc binding.
AB - The large subunit of TFIIE (TFIIEα) has a highly conserved zinc ribbon domain, which is essential for transcription. Recently, we determined the solution structure of this domain to be that of a novel zinc finger motif [Okuda et al. (2004) J. Biol. Chem. 279, 51395-51403]. On examination of the functions of four cysteine mutants of TFIIEα, in which each of four zinc-liganded cysteines was replaced by alanine, we found an interesting functional asymmetry; on a supercoiled template, the two C-terminal mutants did not show any transcriptional activity, however, the two N-terminal mutants retained about 20% activity. Furthermore, these two pairs of mutants showed distinct binding abilities as to several general transcription factors. To obtain structural insights into the asymmetry, here we have analyzed the structures of the four cysteine mutants of the zinc ribbon domain by CD and NMR. All four mutants possessed a characteristic partially folded structure coordinating with a zinc atom, despite the imperfect set of cysteine-ligands. However, they equilibrated with several structures including the random coil structure. Unexpectedly, the two N-terminal mutants mainly equilibrated with the random coil structure, while the two C-terminal ones mainly equilibrated with folded structures. The characteristic structure formation of each mutant was reversible, which totally depended on the zinc binding.
KW - CD
KW - NMR
KW - TFIIE
KW - Transcription
KW - Zn-binding
UR - http://www.scopus.com/inward/record.url?scp=28244449100&partnerID=8YFLogxK
U2 - 10.1093/jb/mvi138
DO - 10.1093/jb/mvi138
M3 - 学術論文
C2 - 16272138
AN - SCOPUS:28244449100
SN - 0021-924X
VL - 138
SP - 443
EP - 449
JO - Journal of Biochemistry
JF - Journal of Biochemistry
IS - 4
ER -