Transcript Chapter 21 (part 1) - Nevada Agricultural Experiment

Chapter 21 (part 1)
Transcription
Central Dogma
RNA Content of E. coli
Cells
type
Steady
State
Levels
Synthetic
Capacity
Stability
rRNA
83%
58%
High
tRNA
14%
10%
High
mRNA
3%
32%
Very Low
E. Coli RNA Polymerase
• RNA polymerase core
enzyme is a multimeric
protein a2,b, b’, w
• The b’ subunit is
involved in DNA
binding
• The b subunit contains
the polymerase active
site
• The a subunit acts as
scaffold on which the
other subunits
assemble.
• Also requires s-factor
for initiation –forms
holo enzyme complex
Site of DNA
binding and RNA
polymerization
General Gene Structure
5’
Promoter
Transcribed region
terminator 3’
Gene Promoters
Other s-Factors
• Standard genes – s70
• Nitrogen regulated genes – s54
• Heat shock regulated genes – s32
Transcriptional
Initiation
Closed complex
Open complex
Primer formation
Disassociation
of s-factor
Pausing induces termination
3’end tends to be AU
rich easily to disrupt
during pausing. Leads to
disassembly of RNA
polymerase complex
Rho Dependent Termination
• rho is an ATPdependent helicase
• it moves along RNA
transcript, finds the
"bubble", unwinds it
and releases RNA chain
Eukaryotic RNA
Polymerases
type
Location
Products
RNA polymerase I
Nucleolus
rRNA
RNA polymerase
II
Nucleoplasm
mRNA
RNA polymerase
rRNA, tRNA,
Nucleoplasm
III
others
Mitochondrial RNA
Mitochondrial gene
Mitochondria
polymerase
transcripts
Chloroplast RNA
polymerase
Chloroplast
Chloroplast gene
transcripts
Eukaryotic RNA
Polymerases
• RNA polymerase I,
II, and III
• All 3 are big,
multimeric proteins
(500-700 kD)
• All have 2 large
subunits with
sequences similar to b
and b' in E.coli RNA
polymerase, so
catalytic site may be
conserved
Eukaryotic Gene Promoters
• Contain AT rich concensus sequence
located –19 to –27 bp from transcription
start (TATA box)
• Site where RNA polymerase II binds
Transcription Factors
• TFAIIA, TFAIIB –
components of RNA
polymerase II holoenzyme complex
• TFIID – Initiation factor,
contains TATA binding
protein (TBP) subunit.
TATA box recognition.
• TFIIF – (RAP30/74)
decrease affinity to nonpromoter DNA