Prokaryotic Transcription

Transcription in Prokaryotes

• Prokaryotic transcription uses a single RNA polymerase.
• In E. coli, RNA polymerase is a 465 kD complex.
• It has multiple subunits: 2 α, 1β, 1β', 1 ω & 1σ.
• The RNA polymerase holoenzyme recognizes promoter sequences on DNA.
• Subunits are essential for assembly and activation by regulatory proteins.
• The polymerase is accurate, with approximately one error in 10,000 bases.
• This error rate is acceptable due to many transcripts produced from each gene.

Objectives

• Transcription in prokaryotes
• Properties of promoters in prokaryotes and eukaryotes
• Comparison of transcription in prokaryotes and eukaryotes

Stages of Transcription

• RNA polymerase holoenzyme binds to promoter sites.
• Polymerization initiates.
• Chain elongation occurs.
• Transcription terminates.

Properties of Promoters

• Promoters are typically 40 base pairs long.
• They are located on the 5' side of the transcription start site.
• Two consensus sequence elements:
  • The -35 region: consensus TTGACA. Sigma subunit binds here.
  • The Pribnow box (-10): consensus TATAAT. Ideal for unwinding.

Properties of Eukaryotic Promoters

• Eukaryotic gene promoter sequences typically lie between 45 to 55 bases.
• They include consensus sequences like the CAAT box (GGCCAATCT) and the TATA box (TATAAAA).

1- Initiation of Polymerization

• RNA polymerase has two binding sites for NTPs.
• The initiation site prefers ATP and GTP (most RNAs start with a purine at the 5' end).
• The elongation site binds the next incoming NTP.
• The 3'-OH of the first NTP attacks the alpha-phosphate of a second NTP, forming a phosphodiester bond.
• When 6-10 nucleotides have been added, the sigma subunit dissociates, completing initiation.
• Rifamycin and rifampicin inhibit transcription by binding to RNA polymerase.

2- Chain Elongation

• Core polymerase (without sigma subunit) elongates the RNA chain.
• Elongation rate is 20-50 bases per second, slower in G-C rich regions.
• Topoisomerases aid by relieving supercoiling ahead of and behind the polymerase.

3- Chain Termination

• Termination occurs by two mechanisms:
  • Rho-dependent: ATP-dependent helicase unwinds the RNA transcript, causing termination.
  • Rho-independent: Inverted repeat sequences transcribed into RNA create a hairpin loop, leading to termination. (Hair pin loop often followed by several uracils).

Termination Strategies in Bacteria

• Rho-dependent termination: Rho protein binds to the RNA and moves to the termination site, causing termination.
• Rho-independent termination: Sequence in DNA that forms the termination hairpin, along with a series of U's (often 4-9) in the mRNA triggers detachment of the polymerase.

Rho-Protein

• Rho is an essential protein for prokaryotic transcription termination.
• It unwinds the RNA-DNA hybrid helix.

Comparison of Transcription in Prokaryotes and Eukaryotes

Explore the transcription process in prokaryotes, focusing on RNA polymerase structure and function in E. coli. This quiz covers the stages of transcription, the properties of promoters, and compares prokaryotic and eukaryotic transcription mechanisms.