DNA transcription
DNA transcription vs DNA replication
Transcription Replication
Does not require a primer DNA Polymerase needs an RNA Primer
Not all of the DNA is transcribed
(Large part of the genome is never transcribed and The whole genome is replicated - mitosis
made into RNA)
Only 1 strand of the DNA is transcribed by RNA polymerase Both strands of the DNA are replicated by DNA Polymerase
Similarities
Addition of nucleotides to 3’ end of the growing chain
Direction of growth: 5' to 3’
Uses the DNA template
Phases: Initiation, Elongation, Termination
RNA Polymerase subunits
• Mechanism: (NMP)n + NTP →(NMP)n+1 + PPi
• DNA dependent
• Active site has Mg2+ → second Mg2+ is brought in with new nucleotides in the
nucleotide addition and released with pyrophosphate
• Subunit composition: α2ββ'σ → holoenzyme (only this enzyme capable of initiating
transcription)
• Initiates transcription only at promoter region: conserved sequence at -35 and -10
region → recognised by subunit σ
• σ initiates transcription and dissociates →leaves the core enzyme ( α2ββ') which
carries out elongation
• Active site = β and β'
σ - transcription initiation
1. Recognition of primer by subunit σ
• Promoter region: conserved sequence at -35 and -10 region (bound by RNA
Pol but not made into RNA)
• Strong promoters have UP-element: strongly stimulate transcription by
providing an additional specific interaction between the RNA Pol and DNA eg)
the sequences directing the expression of rRNA
2. Role of subunit σ
• Has high affinity for DNA
• Reduces core enzyme's affinity to DNA → allows holoenzyme migrate along
DNA until promoter is found and bind specifically
• For regulated gene expression, different σ factors enable binding to different
promoters
• alpha helix of σ binds to the major groove of the DNA around -10 region and
allow DNA melting → transition from closed to open complex
• The alpha helix (helix-turn helix motif) is involved in recognition of -10 region
that contains aromatic AAs that can interact with bases on the non-template
strand = stabilises the melted DNA
• The favourable binding interaction between ssDNA and σ: 2 bases in the non-
template strand are flipped out and inserted into the pockets of σ protein
where favourable interactions stabilise the unwound state of the promoter
region
E.coli promoters
• Many different bacterial promoters reveal high degree of variation but all contain related
sequences → "Consensus Nucleotide Sequence"
• "average" the most common nucleotide found at each position in the collection of
promoters
• -35 and -10 sequences
DNA transcription vs DNA replication
Transcription Replication
Does not require a primer DNA Polymerase needs an RNA Primer
Not all of the DNA is transcribed
(Large part of the genome is never transcribed and The whole genome is replicated - mitosis
made into RNA)
Only 1 strand of the DNA is transcribed by RNA polymerase Both strands of the DNA are replicated by DNA Polymerase
Similarities
Addition of nucleotides to 3’ end of the growing chain
Direction of growth: 5' to 3’
Uses the DNA template
Phases: Initiation, Elongation, Termination
RNA Polymerase subunits
• Mechanism: (NMP)n + NTP →(NMP)n+1 + PPi
• DNA dependent
• Active site has Mg2+ → second Mg2+ is brought in with new nucleotides in the
nucleotide addition and released with pyrophosphate
• Subunit composition: α2ββ'σ → holoenzyme (only this enzyme capable of initiating
transcription)
• Initiates transcription only at promoter region: conserved sequence at -35 and -10
region → recognised by subunit σ
• σ initiates transcription and dissociates →leaves the core enzyme ( α2ββ') which
carries out elongation
• Active site = β and β'
σ - transcription initiation
1. Recognition of primer by subunit σ
• Promoter region: conserved sequence at -35 and -10 region (bound by RNA
Pol but not made into RNA)
• Strong promoters have UP-element: strongly stimulate transcription by
providing an additional specific interaction between the RNA Pol and DNA eg)
the sequences directing the expression of rRNA
2. Role of subunit σ
• Has high affinity for DNA
• Reduces core enzyme's affinity to DNA → allows holoenzyme migrate along
DNA until promoter is found and bind specifically
• For regulated gene expression, different σ factors enable binding to different
promoters
• alpha helix of σ binds to the major groove of the DNA around -10 region and
allow DNA melting → transition from closed to open complex
• The alpha helix (helix-turn helix motif) is involved in recognition of -10 region
that contains aromatic AAs that can interact with bases on the non-template
strand = stabilises the melted DNA
• The favourable binding interaction between ssDNA and σ: 2 bases in the non-
template strand are flipped out and inserted into the pockets of σ protein
where favourable interactions stabilise the unwound state of the promoter
region
E.coli promoters
• Many different bacterial promoters reveal high degree of variation but all contain related
sequences → "Consensus Nucleotide Sequence"
• "average" the most common nucleotide found at each position in the collection of
promoters
• -35 and -10 sequences
