title: Molecular basis of transcription initiation in Archaea.
abstract: Compared with eukaryotes, the archaeal transcription initiation machinery-commonly known as the Pre-Initiation Complex-is relatively simple. The archaeal PIC consists of the TFIIB ortholog TFB, TBP, and an 11-subunit RNA polymerase (RNAP). The relatively small size of the entire archaeal PIC makes it amenable to structural analysis. Using purified RNAP, TFB, and TBP from the thermophile Pyrococcus furiosus, we assembled the biochemically active PIC at 65ºC. The intact archaeal PIC was isolated by implementing a cross-linking technique followed by size-exclusion chromatography, and the structure of this 440 kDa assembly was determined using electron microscopy and single-particle reconstruction techniques. Combining difference maps with crystal structure docking of various sub-domains, TBP and TFB were localized within the macromolecular PIC. TBP/TFB assemble near the large RpoB subunit and the RpoD/L "foot" domain behind the RNAP central cleft. This location mimics that of yeast TBP and TFIIB in complex with yeast RNAP II. Collectively, these results define the structural organization of the archaeal transcription machinery and suggest a conserved core PIC architecture.
authors.affiliation: Department of Chemistry, City College of the City University of New York, NY, USA. sdecarlo@ccny.cuny.edu
authors.forename: Sacha
authors.initials: S
authors.lastname: De Carlo
authors.forename: Shih-Chieh
authors.initials: SC
authors.lastname: Lin
authors.forename: Dylan J
authors.initials: DJ
authors.lastname: Taatjes
authors.forename: Andreas
authors.initials: A
authors.lastname: Hoenger
mesh_terms: D019843:Archaeal Proteins / Q000737:chemistry / Q000378:metabolism*; D020285:Cryoelectron Microscopy; D018360:Crystallography, X-Ray; D019641:DNA, Archaeal / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012321:DNA-Directed RNA Polymerases / Q000737:chemistry / Q000378:metabolism*; D046529:Microscopy, Electron, Transmission; D008958:Models, Molecular; D046912:Multiprotein Complexes / Q000737:chemistry / Q000378:metabolism; D009690:Nucleic Acid Conformation; D011485:Protein Binding; D020836:Protein Structure, Quaternary; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D021122:Protein Subunits / Q000737:chemistry / Q000378:metabolism; D020027:Pyrococcus furiosus / Q000235:genetics* / Q000378:metabolism; D035181:TATA-Box Binding Protein / Q000737:chemistry / Q000378:metabolism; D035581:Transcription Factor TFIIB / Q000737:chemistry / Q000378:metabolism*; D014158:Transcription, Genetic / Q000235:genetics*
chemical_list: D019843:Archaeal Proteins; D019641:DNA, Archaeal; D046912:Multiprotein Complexes; D021122:Protein Subunits; D035181:TATA-Box Binding Protein; D035581:Transcription Factor TFIIB; C106466:transcription factor B, Archaea; D012321:DNA-Directed RNA Polymerases