week_9_lecture_1

Also extensive interactions between the phosphate

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Unformatted text preview: MORE REVERSE TRANSCRIPTS MADE TRANSCRIPTS TBP-DNA complex: TBP binds to and distorts DNA using a β sheet inserted into the minor groove A:T bases are favoured because they are more readily distorted to allow the initial opening of the minor groove There are also extensive interactions between the phosphate backbone and basic residues in the β sheet TBP: TATA-BINDING PROTEIN TBP: TBP • • • • Weaver 2002 Molecular Biology Ed. 2 TBP binds DNA minor groove forming “saddle” on DNA duplex ­ bends DNA (similar to CAP­cAMP) 80˚ at TATA box • TFIID consists of: TATAbinding protein (TBP) & 11 binding TBP-associated factors (TAFs) (TAFs) Different TAFs for different Different promoters promoters TFIID typically has a mass of TFIID ~800 kDa ~800 TBP “commitment factor” TBP binds RNAP II first & consigns promoter to be transcribed Bend in DNA caused by TBP Bend may serve to bring transcription factors and RNA polymerase closer together polymerase TBP/TATA BOX INTERACTIONS TBP/TATA huTBP huTBP produced in vaccina virus virus …or in or bacteria bacteria Weaver 2002 Molecular Biology Ed. 2 • Footprinting: TBP binds specifically to adenovirus major Footprinting: late TATA box (TBP is the only general transcription factor that makes sequence-specific DNA contacts) factor (b) no TBP binding to mutant TATA box (TTATCAT) (c) 180 a.a. C-terminal TBP region binds TATA box RNAP INITIATION COMPLEX RNAP • RNAP II initiation requires general RNAP factor complex assembly in defined order: order: • TFIID assembly (TBP and TAFs) ---> TFIID TFIIA ---> TFIIB ---> TFIIF TFIIA (2 subunits) ---> RNAP II ---> (DNA melting) ---> TFIIE ---> TFIIH & (J) ---> (promoter TFIIE clearance) clearance) Footprinting shows increased DNA Footprinting length covered as each subunit added: TFIIA -increased protection upstream TFIIB -protection downstream of TATA -protection box box • • • TAFs bind to DNA elements at the promoter, some have structural homology to histones TFIIB (one protein): crystal structure shows specific base contacts with the major groove upstream of BRE and the minor groove downstream of TATA Asymmetric binding of TFIIB to the TBP-TATA complex accounts for the asymmetry in the rest of assembly in th...
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