alpha - amanitin
Non-competitive inhibitor of Poll II
Binds bridge helix
halts trxn by blocking translocation and elongation
TB & broad spectrum antibiotic
Blocks exit channel to halt trxn
What rxn does RNA polymerase catalyze?
3’-OH nuc. attack on 5’-phosphate
What are the 5 steps in the trxn cycle? (both bact. and euk)
- Binding –> closed complex
- Open complex
- Initial phosphodiester linkage
Where does RNA polymerase bind?
to the promoter (think tata box)
How does the open complex form?
RNA polymerase “melts” the DNA to form a trxn bubble
How does the initial linkage form?
polymerase catalyzes initial phosphodiester bond between the two initial rNTP’s
In what direction does RNA pol. advance along the DNA?
3’ –> 5’
-so the complimentary new RNA is being built 5’ –> 3’
Which steps in trxn are “heavily” regulated?
- Initiation (and its 3 substeps)
2. Elongation (pauses allow for regulation)
What are the 4 DNA dependent RNA polymerases?
- Pol I
- Pol II
- Pol III
Function of RNA pol I ?
Function of RNA pol II?
Function of Pol III ?
tRNA, U6 snRNA
Where does U6 snRNA function?
Is part of the splicing machinery
What is special about mitochondrial RNA?
Is one small subunit and one small helper protein. Most others are multiprotein complexes.
Specific sequence in the DNA that directs trxn of adjacent genes
- variable from gene to gene
- commonality is TATA box
Where is the TATA box and why T’s and A’s?
10 - 30 bp upstream from start site
A-T easier to melt
4 components of the RNA polymerase II pre-initiation complex.
- TFIID - binds TATA box (guided by TBP = tata binding protein)
- TFIIH - contains XPB, XPD = helicase activity
- contains CD47 = phosphorylates C-term. domain of pol II
- contains p44
- Mediator - protein-protein contact during initiation
- pol II holoenzyme
What does TBP bind and where?
in the minor groove of the TATA box
signals assembly of the pre-initiation complex
TFIIH has two major functions. What are they (broadly)?
- trxn - initiation complex
2. DNA repair (helicase activity)
Name 3 clinical syndromes caused by mutations in TFIIH subunits.
- Xeroderma Pigmentosum
- Cockaynes Syndrome
To help you remember Trichothiodystrophy, Tricho mean hair and (remember trichotelomania?)
What is a classic clinical presentation of TTD?
Brittle hair - lacks sulpher (thio)