DNA-binding determinants of the alpha subunit of RNA polymerase: novel DNA-binding domain architecture.
The Escherichia coli RNA polymerase alpha-subunit binds through its carboxy-terminal domain (alpha CTD) to a recognition element, the upstream (UP) element, in certain promoters. We used genetic and biochemical techniques to identify the residues in alpha CTD important for UP-element-dependent transcription and DNA binding. These residues occur in two regions of alpha CTD, close to but distinct from, residues important for interactions with certain transcription activators. We used NMR spectroscopy to determine the secondary structure of alpha CTD, alpha CTD contains a nonstandard helix followed by four alpha-helices. The two regions of alpha CTD important for DNA binding correspond to the first alpha-helix and the loop between the third and fourth alpha-helices. The alpha CTD DNA-binding domain architecture is unlike any DNA-binding architecture identified to date, and we propose that alpha CTD has a novel mode of interaction with DNA. Our results suggest models for alpha CTD-DNA and alpha CTD-DNA-activator interactions during transcription initiation.
Gaal
T
T
Department of Bacteriology, University of Wisconsin, Madison 53706, USA.
Ross
W
W
Blatter
E E
EE
Tang
H
H
Jia
X
X
Krishnan
V V
VV
Assa-Munt
N
N
Ebright
R H
RH
Gourse
R L
RL
D000595:Amino Acid Sequence; D001483:Base Sequence; D004247:DNA / Q000378:metabolism; D012321:DNA-Directed RNA Polymerases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics; D009682:Magnetic Resonance Spectroscopy; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016296:Mutagenesis; D011401:Promoter Regions, Genetic; D011487:Protein Conformation; D012335:RNA, Ribosomal / Q000235:genetics / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000235:genetics; D014158:Transcription, Genetic; D001616:beta-Galactosidase / Q000235:genetics
D012335:RNA, Ribosomal; D011993:Recombinant Fusion Proteins; D004247:DNA; D012321:DNA-Directed RNA Polymerases; D001616:beta-Galactosidase