tata binding protein
TRANSCRIPT
Transcription in Eukaryotes
• Eukaryotic RNA polymerases, unlike their
bacterial counterparts, are incapable of binding
by themselves to their respective promoters
• Eukaryotic RNA polymerases rely on proteins
called transcription factors to show them the
way
• Two classes: general transcription factors and
gene-specific transcription factors (activators)
• General transcription factors combinewith RNA polymerase to form apreinitiation complex
• The assembly of preinitiationcomplexes involving polymerase IIis quite complex
Structure and Function of TFIID
TFIID contains several subunits
– TATA-box binding protein (TBP)
• Highly evolutionarily conserved
• Binds to the minor groove of the TATA box
–Saddle-shaped TBP lines up with DNA
–Underside of the saddle forces open the
minor groove
–The TATA box is bent into 80° curve
– TBP-associated factors (TAFs) specific for class II
TBP [TFIID] function
• Binds TATA - main sequence recognition event during
Binds a variety of different TATA-like sequences
– A slow binding reaction
– minor groove contact
– binds as monomer
• Affinity of TBP for TATA contributes to promoter strength
• Binds also several other polypeptides– activators (Sp1, Tax1, E1A)
– TAFs (dTAF110, dTAF40)
– GTFs (TFIIB, TFIIA)
– inhibitors
• TBP = universal TF involved in all three pol syst.– TBP i SL1, TFIID, TFIIIB
DNA
Other factors
N
TBP versus TFIID
• Subunit-structure– TFIID = TBP + multiple TAFs
– mammalian TFIID: 750 kDa (II), 300 kDa (III) and 200 kDa (I)
– TBP only a small core in the TFIID complex• human 38 kDa, yeast 27 kDa, Arabidopsis 22 kDa
– TBP = N-term divergent domain + C-term. conserved domain
• C-term domain 180aa
• Carries all essential functions
• N-term domain divergent
TAFs
TBP
N
TBPs saddle-structure
DNA
protein
Concaveinside
Convexsurface
3D: saddle-structure• Twofold symmetry - form of a saddle.
• Concave inside binds DNA in minor groove through a 10-stranded antiparallel -sheet
• Convex surface binds other GTFs via 4 -helixes• loop (“stirrup”) on each side with Phe side-
chains intercalating in DNA
TBPs effect on DNA
• DNA-structure is distorted upon TBP binding– DNA severely bended, unwinded and distorted
– DNA shaped by TBP´s -sheet
– The intercalating Phe-residues contributes to kink
• Effect?– Upstream and downstream elements brought closes together
– incompatible with nucleosome structure
.. but this wayNot like this
A Two-Step Mechanism of TBP
Binding to DNA• First step
– Full-length TBPWT first binds to TATA box to form an unbent TBP-TATA box complex.
• Second step– Then, this unbent complex
slowly forms the bent TBP-TATA box complex.
– TFIIB can directly recognize the unbent and/or bent TBP-TATA-complexes to form the bent TBP-TATA box complex.
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The Versatility of TBP
• Genetic studies have demonstrated TBP
mutant cell extracts are deficient in:
– Transcription of class II genes
– Transcription of class I and III genes
• TBP is a universal transcription factor required
by all three classes of genes
• Required in transcription of at least some
genes of Archaea, single-celled organisms
lacking nuclei