Transcription And It's Steps In Prokaryotes

Transcription:

Transcription is the process of synthesis of RNA using DNA as a template. The key enzyme is responsible for transcription is DNA-dependentRNA polymerase.

Structure of RNA polymerase:

RNA polymerase is a multisubunit enzyme. It consists of two parts i.e.σ factor and core enzyme. σ factor enables RNA polymerase to recognize promoter region of the DNA. Core enzyme consists of four polypeptide subunits, 2α, 1β and 1β' joined together byÑ peptide. It is the core enzyme that is responsible for 5' to 3' RNA polymerase activity. The σ factor and co-enzyme together compriseRNA polymerase holoenzyme. The process of transcription can be divided into following steps;

1. Initiation

2.Release of σ factor

3. Chain elongation

4. Termination

1.Initiation:

Initiation of transcription involves the binding of RNA polymerase holoenzyme to a specific region of the DNA called promoter region. The two conserved sequences found in E-coli promoter region are:-

1. Pribnow box:

This is a stretch of 6 nucleotides (5'-TATAAT-3') situated 8-10 nucleotides upstream of transcription start site.

1. -35 Sequences:

A second conserved sequence (5'-TTGACA-3') is found about nucleotides upstream of the transcription start site. The RNA polymerase holoenzyme binds to the promoter site with the help of its σ-factor and initiates its transcription activity.

Release of σ-factor:

Once the promoter region has been recognized by the RNA polymerase holoenzyme with the help of σ-factor, RNA polymerase begins to synthesize a transcript of DNA sequence. Theσ-factor is released as soon as RNA polymerase synthesize few nucleotides complementary to the template DNA strand and then leaves the core enzyme to the template DNA.

3.Chain elongation:

- Unlike DNA polymerase, does not require a primer.-DNA-dependent RNA polymerase requires all four types of nucleotides (ATP, UTP, GTP, CTP) as

-DNA-dependent RNA polymerase requires all four types of nucleotides (ATP, UTP, GTP, CTP) aspire-cursor in addition to DNA template.

-Only one of the two DNA strand serves as a template strand .

-The template strand is copied 3'-5' direction i.e new RNA strand is synthesized 5'-3' direction.

-Unlike DNA polymerase most of the RNA polymerase have no exonuclease activity,i.e they lack proofreading.

-During chain elongation phase of transcription, the growing end of new RNA chain base pairs temporarily with DNA template to form a short RNA-DNA hybrid duplex. The length of this hybrid duplex is around 8 base pairs. The RNA in this hybrid duplex peels off shortly after it's formation and DNA duplex reforms.

-To enable RNA polymerase to synthesize an RNA strand complementary to one of the two DNA strands, the DNA duplex must unwind over a short distance forming a transcription bubble. During transcription, The RNA polymerase generally separates 17 base pairs to form transcription bubble.

-The problem of supercooling is solved by DNA topoisomerase enzyme.

4.Termination:

The process of chain elongation of RNA continues until a terminal signal is reached. The termination may occur in two ways :

a.Rho (ρ)-dependent termination and

b.Rho(ρ)-independent termination

a.Rho (ρ)-dependent termination:

ρ-dependent transcription termination depends on a protein known as ρ-factor. ρ-factor binds to the GC-rich region near the 3' end of the newly synthesized RNA and migrates behind the RNA polymerase enzyme and at the termination site it causes RNA to leave the DNA template from RNA-DNA hybrid duplex as it has helicase-like activity. This process isATP-requiring and terminates transcription process. The DNA duplex is then reformed.

b.Rho (ρ)-independent termination:

Two specific conditions;

a.The mRNA transcript must be able to form stable hairpin like structure (secondary) which slows down the progress of RNA polymerase enzyme.

b.RNA transcript must possess the string of 'U' at the three prime end which is complementary to a stretch of 'A' in the template DNA. The bonding of A-U is weak and hence facilitates the separation of newly synthesized RNA from its template DNA strand as the double helix of DNA zips off behind RNA polymerase.

References:

(DL and MM)