Biochemistry 2280A Study Guide - Final Guide: Lac Operon, Pribnow Box, Consensus Sequence
30 Nov 2015
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11/24/2015 5:03:00 PM
2280 Topic 23
Why is there n RNA step in gene expression
1. the RNA step provides an amplification which contributes to gene
expression – RNA allows for genes to have some more transcripts than
others – couldn’t do if just DNA→Protein
2. Since RNA can be degraded, expression of a gene can be stopped quickly
- then can turn back on again
3. RNA provides additional opportunities to regulate gene expression
• RNA processing
• RNA export from the nucleus
Transcription
• 1st step of gene expression and for many the most frequently
regulated
• Prokaryotic Transcription
o Gene and promoter structure
o RNA polymerase
o Mechanism of transcription
o Regulation of transcription
▪ TRO operon
▪ lac operon
o Promoter: DNA sequence required to initiate transcription of
gene or operon
o Terminator: DNA sequences required to stop transcription
o Operon: A set of genes transcribed from a single promoter
and thus expressed from a common RNA
Bacterial Operon Structure
• Start site of transcription is +1, base upstream is -1, other side is
+2
• Different operons = different protein coding genes
• All expressed from common mRNA – operon
What are the key features of a bacterial promoter?
• 2 Key sequences
o -10: – 10 base pairs upstream of +1
o -35: - 35 base pairs upstream of +1
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o How did people fio these existed? – Looked at a bunch of
promoters – are there common sequence elements in
different E.coli promoters? – if yes have functional promoters
o What sequence patterns emerge? – noticed TA rich sequence
(pribnow box) at -10 site
▪ Most often T at first position most seconds were A
▪ Slightly differs at initiation site
▪ Same at -35 (rich in certain base pairs)
o How to determine if -35 sequence is important
▪ Mutate and see what changes – how would it effect
transcription
o When describing the -10 as TATAAT, or -35 as TTGACA you
are referring to……..consensus sequence
o Consensus sequence – Most frequent base at each position in
a group of functionally related DNA elements
o No elements identical to consensus – based on frequency
Bacterial RNA Polymerase
• The enzyme that makes RNA transcripts using DNA as a template
and nucleoside triphosphates (NTPs) as substrates
• Others that make RNA in cell but only one that uses DNA as
template is this
• Multi-subunit enzyme
The core enzyme can make RNA but does not recognize promoters
• The promoter specificity of RNA polymerase is determined by the
sigma subunit
• RNA polymerase + sigma subunit = RNA Polymerase holoenzyme
• Sigma allows the core enzyme to recognize promoter – KEY ROLE
• Sigma contacts the -10 and -35 sequences
o Makes space specific contact – physical with these regions
• Core enzyme does not make base specific contact
Steps in the Initiation of transcription
1. RNAP holoenzyme binds the promoter
• Start with RNA polymerase with sigma subunit
• Holoenzyme + sigma binds to promoter
• Closed complex – when RNA polymerase is bound
2. RNA polymerase unwinds the DHA strands around the start site
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